Report scientific 4a parte by IEO Istituto Europeo di Oncologia

Research activities

Clinical Research

The last year has seen a gradual introduction of a disease-orientated Task Force structure to replace the previous Working Groups at the Institute. These new initiatives have been formed to ensure that Molecular Medicine scientists can integrate seamlessly into the clinical multidisciplinary groups. Also, each Task Force has now got dedicated infrastructural support in the form of statistics and data management. Each Task Force will have a rotating Chairman and vice chairs, plus a professional secretary. The purpose of their meetings is to brainstorm, then formulate new research translational projects, interdigitated with clinical protocols. Funding will be the Task Forces’ own responsibility, together with monitoring and publication of results. The Task Force should also adopt the responsibility of training in research methodology, the clinical and non-clinical research fellows working on the relevant disease site, irrespective of their professional discipline. Inevitably the evolution of the new structure means that the original examplar, the Breast Cancer Task Force, is extremely mature and very productive, whereas several others are still settling down. However, a large number of new projects have been planned and are described in the following pages. Here we are also reporting the summary of some of the results already obtained grouping them either for organ pathologies or for transversal research programmes. One chapter deals with the IEO Tumour Registry. A speciﬁc chapter has been added on Biobank which will be soon in full operation.

Each research chapter had a small Writing Committee composed of scientists often not entirely involved in the described research activities. We are grateful to all of them for their efforts to sintetically describe the many activities developed in IEO with the aim of integrating laboratory and clinical research.

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Identiﬁcation and validation of new molecular targets for drug development

Successful cancer treatment requires an understanding of the cascade of molecular lesions that underlie the acquisition of the tumor phenotype and that can therefore constitute meaningful therapeutic targets or diagnostic and prognostic indicators. The search for these molecular lesions has received great impetus over the last decades, and we now have an ever expanding catalog of genetic and epigenetic alterations that are being associated to various types of cancer and to various stages of its development. In parallel there has been a conceptual reframing of cancer as a developmental problem, which is enabling to position the list of molecular lesions within a pathological history of the disease and a hierarchy of cell lineages. This conceptual reframing of cancer allows then to test experimentally, thanks to new cancer models and to novel high-throughput technologies, how the various cell compartments, the genetic and epigenetic lesions that affect them, and the range of cancer cell-host interactions ultimately shape the tumor phenotype. Consistent with this conceptual and experimental paradigm, and in line with the objective to investigate cancer as a complex disorder of development gone awry, we have focused on: i) the alterations in genetic and epigenetic regulatory networks; ii) the actual aberrations in cell division; iii) the emerging links between cancerous growth and the molecular environment of the host; and iv) the predictive relevance of molecular markers for clinical prognosis. This structure is meant to follow our reﬁned understanding of the disease, in which the derangement of growth and differentiation programs (through genetic and epigenetic mechanisms) leads to abnormal cell division in the context of the host tissue and its microenvironment, enabling a molecular dissection that is a prerequisite for the experimental validation of therapeutic interventions. I. Genetic and epigenetic deregulation 1. Probing the cancer networks with genome wide approaches (Myriam Alcalay) The main focus of the lab is to understand transcriptional networks involved in oncogenesis by exploiting the integrated use of high-throughput technologies, which

represent powerful tools for the discovery and analysis of genetic networks underlying cancer. We are focusing on two main lines of research: a. Molecular signature of Acute Myeloid Leukemia We previously analyzed specific expression profiles associated to acute myeloid leukemia (AML) in different model systems: • Expression of oncogenic transcription factors PML/RAR, AML1/ETO and PLZF/RAR in U937 cells (1) • AML bearing NPM mutations (2) • Specific cellular subpopulations purified from the bone marrow of PML/RAR or AML1/ETO leukemic mice (unpublished). On the basis of the results obtained from these studies, we initiated the functional characterization of a specific target gene, Pirin (PIR), which is strongly downregulated by oncogenic fusion proteins in U937 cells, with the aim to assess its role in oncogenesis. Pirin is a novel protein identified as an interactor of NFI/CTF1 transcription factor. It is highly conserved in evolution and is ubiquitously expressed in human tissues with nuclear localization. The function of Pirin in mammalian cells remains largely unknown. Some reports suggest a possible role as a tumor suppressor gene in humans. Our studies of Pirin expression/function in hematopoiesis and leukemia showed that: • PIR expression is significantly repressed in a large proportion of AML, regardless of subtype or underlying karyotipic abnormalities. • PIR expression increases during in vitro myeloid differentiation of primary hematopoietic precursor cells • ablation of PIR in the U937 cell line or in murine primary hematopoietic precursor cells results in impairment of terminal myeloid differentiation • knock-down of PIR in U937 cells results in increased expression of genes associated to the early phases of hematopoiesis, in particular HOXA genes. Taken together, our results suggest that PIR is required for terminal myeloid maturation, and its downregulation may contribute to the differentiation arrest associated to AML.

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These results have been recently published (3). Deregulated expression of Pirin may also be involved in other tumor types. In fact, Tissue Microarray analysis (TMA) revealed that Pirin expression is down-regulated in approximately 50% of melanomas. In a subset of melanoma samples where the protein is instead expressed at high levels, it is however de-localized to the cytoplasm (4). Loss of function experiments in melanocytic cells showed that PIR is a negative regulator of cellular senescence, and may have a relevant role in melanoma progression (5). b. The PRDM gene family in cancer PRDM5 is as a transcriptional repressor that behaves as a putative tumor suppressor in different types of cancer. In previous work, we showed that the tumor suppressor Prdm5 negatively modulates both canonical Wnt/b-catenin and planar cell polarity (PCP) Wnt pathways during zebrafish development, suggesting that inactivation of PRDM5 may represent a mechanism of activation of Wnt signaling in human tumors (6). We found that PRDM5 physically interacts with nucleophosmin (NPM), a nucleolar protein that shuttles between the nucleus and the cytoplasm and that acts both as proto-oncogene and as tumor suppressor. NPM is the most frequently mutated gene in AML. In zebrafish, both genes are strongly expressed during embryogenesis. We found that the effects of prdm5 and npm1 targeted knock-down are similar and resemble an impairment of morphogenetic movements during gastrulation due to a block of PCP Wnt pathway, accompanied by a reduction of the cephalic region. In agreement, overexpression of both prdm5 and npm1 causes a “dorsalized” phenotype. Functional studies using loss and gain of function approaches showed that both prdm5 and npm1 increase dkk1 expression at early stages of development, thus repressing canonical Wnt/b-catenin pathway in addition to the PCP pathway. Expression of the NPM mutant isolated from human AML in zebrafish embryos results in a phenotype similar to that obtained through knock-down experiments, suggesting the mutant protein acts as a dominant negative and may exert its oncogenic activity at least partly through activation of Wnt signaling. Further studies focusing more specifically on the effects of NPM mutant in early hematopoiesis are currently ongoing. 2. Myc-induced senescence: a new rationale for targeting Cdk2 kinase activity in cancer (Bruno Amati) As already summarized in last year’s report, our work had uncovered a unique role of the cyclin-dependent kinase 2 (Cdk2) in suppressing a senescence program induced by the c-myc oncogene. This work was recently published (Campaner et al., 2010). Activation of the c-myc gene product (the Myc protein) in cultured Cdk2-/- mutant cells caused cellular senescence, an effect not seen in control

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wild-type cells. This effect also ocurred in vivo in myctransgenic Cdk2-/- mice, and was associated with a delay in B-cell lymphomas. Pharmacological inhibition of Cdk2 selectively induced senescence in cells over-expressing Myc, suggesting that pharmacological targeting of Cdk2 might be selectively effective in Myc-driven tumors. Characterisation the novel Ras-family member RasL11a, a In a parallel project, we characterised the function of RasL11b, a Ras super-family protein of unknown function. Our work led to the unexpected finding that RasL11a is located in the nucleolus, is chromatin-associated and modulates pre-ribosomal RNA (pre-rRNA) synthesis (Pistoni et al., 2010). As determined by immunofluorescence, RasL11a co-localized with the RNA polymerase I-specific transcription factor UBF and, like UBF, marked the active subset of rDNA repeats (also called Nucleolar Organizers, or NORs) on mitotic chromosomes. In cells, RasL11a existed in stable complexes with UBF and, as shown by Chromatin immunoprecipitation, distributed along the rDNA transcription unit. Over-expression of RasL11a enhanced pre-rRNA levels in cells, while RasL11a knockdown had the opposite effect. Our data imply that RasL11a acts in concert with UBF to facilitate initiation and/or elongation by RNA Polymerase I. We speculate that the role of RasL11a in this context is to modulate pre-rRNA synthesis in response to specific upstream signals, the nature of which remains to be determined. 3. Mapping the epigenome of cancer cells (Saverio Minucci) Epigenetic alterations are linked to the establishment and maintenance of the cancer phenotype. Importantly, those alterations are potentially reversible, since they do not involve genetic mutations in the underlying DNA sequence. Epigenetic therapies are based on the identification of drugs able to interfere with the activity of enzymes responsible for the epigenetic alterations occurring in tumor cells: new drugs have recently been approved for use in cancer patients, validating clinically this strategy. Unfortunately, however, clinical responses are not always consistent, and do not parallel closely the results observed in preclinical models. The main reasons for this failure are: a) the use of epigenetic drugs in a modality of treatment similar to traditional chemotherapy, not based on a targeted approach; b) the lack of biomarkers for directing therapeutic choices, due at least in part to the lack of appropriate technologies for epigenome profiling from cancer samples, that are just starting to emerge; c) the use of inappropriate preclinical models, mainly based on cell lines where epigenetic drift has occurred as a consequence of long term in vitro culture. Our work is based on the postulate that epigenetic therapies have to be directed against specific functional epigenetic alterations present in cancer cells. In order to identify those

alterations, and investigate their mechanistical and biological consequences, we have developed novel technologies for the study of the epigenome, and adapted their use to preclinical models of leukemias and solid tumors based almost exclusively on the use of primary cells, not undergoing epigenetic drifts. In our studies, we are attempting to generate new tools for mapping epigenetic alterations in biologically relevant cells. We have developed and applied a novel largescale approach based on the use of restriction enzymes (RE) to probe and sequence DNA elements, which are accessible in chromatin, termed NA-Seq (Nuclease Accessible sitesSequencing). With our test we can obtain highly reliable results (90% accuracy in prediction of the epigenetic state) with relatively few cells (Gargiulo et al. Developmental Cell 16, 466-481, 2009). We have applied initially this technique to normal hematopoietic differentiation, and then we have extended the study to leukemic cells. From these studies, we have derived notions on the regulatory circuitry employed in distinct cellular states, and in tumor cells, that we are starting to analyze systematically with the dual goal of improving the molecular understanding of cancer speciﬁc regulatory networks, and to identify novel markers/targets for translational/clinical research. This strategy has provided relevant biological information, with a potential impact on clinical approaches to epigenetic therapies. We intend therefore to further pursue: a) the development of innovative technologies for the study of the epigenome, with a particular emphasis on the miniaturiazation of approaches to use small amounts of patient-derived cells; b) the optimization of preclinical models of acute myeloid leukemias and breast cancer, with an emphasis on the feasibility of studies on the epigenome of cancer initiating and stem cells; c) to explore mechanistically the basis of epigenetic alterations in leukemias and breast cancer, with the concurrent goal of identifying biomarkers of those alterations; d) to monitor the response to epigenetic drugs (alone, or in combination), to identify the molecular determinants of sensitivity/resistance to therapy, and provide a rational basis for the use of epigenetic drugs in stratiﬁed patient populations. While we will mainly use in our studies drugs that are clinically available (for the immediate transfer of relevant information to the patients), or at an advanced stage of clinical experimentation, we plan to explore the potential of novel small molecules modulating the activity of epigenetic enzymes (such as histone demethylases), for which currently there are no available drugs with acceptable properties. 4. Molecular mechanisms of cancer and ageing (Pier Giuseppe Pelicci) The last decade of cancer research has established the concept of cancer stem cells (CSCs) as a subpopulation of cells within a tumor that are entirely responsible for tumori-

genesis. They are called cancer stem cells because, like the stem cells present in normal tissues of the body, they can produce further cells like themselves and also differentiate to provide various different cell types. The cancer stem cell hypothesis provides an appealing perspective through which to examine cancer initiation, recurrence, and metastasis, at the same time arousing strong expectations that targeting cancer stem cells would allow effective tumor eradication. However, it seems that tumour-initiating cells might be resistant to many conventional cancer therapies, which might explain the limitations of these drugs in curing human malignancies. Furthermore, the study of these cancer-driving cells within tumours has been slow because it has been hard to identify them unequivocally, separate them out and study them in the laboratory environment. The focus of our group in the last few years has switched toward the characterization of normal and cancer stem cells (SCs), particularly with regard to the mechanisms of self-renewal regulation studied in vitro and in vivo in preclinical and clinical models of leukemia and breast cancer (mouse models/patientderived primary tumor cells). In this context, the year 2009 has been particularly rewarding for our research activities as our results, defining a number of SC and CSC-specific biological properties and underlying molecular mechanisms, have been published in two of the most important and high impact peer reviewed journals: Nature (Viale et al., Nature 2009) and Cell (Cicalese et al., Cell, 2009). In brief, these were our main findings: i) Cancer stem cells show increased replicative potential, due to the upregulation of the cell-cycle inhibitor p21, which is critical in preventing excess DNA-damage accumulation and functional exhaustion of leukemic SCs; this observation suggests that p21 or the p21-pathways could be molecular targets for therapeutic intervention. ii) Increased frequency of symmetric self-renewing divisions in ErbB2-mammary CSCs is due to p53 inactivation; of particular therapeutic interest is our observation that pharmacological re-activation of p53 correlates with restoration of asymmetric divisions in CSCs and tumor growth reduction, without significant effects on additional cancer cells. We are now extending our investigation on the above molecular targets and on any novel molecular signal or pathway which might be essential to sustain CSC. Indeed, other results of our group, although still preliminary, suggest that symmetric divisions and extended self-renewal in mammary CSCs are due to activation of the c-myc oncogene, as a direct consequence of the loss of function of p53, and that these are imposed by c-myc to CSCs as part of a more general role of this oncogene in supporting a fully competent SC-phenotype. Furthermore, our preliminary results suggest that mammary stem cells also respond to accumulating DNA damage by activating a p53-independent and p21-dependent DNA-repair pathway, and that this response in mammary SCs might also require

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up-regulation of c-myc, which in turn regulates modes of cell division, so that the observed damage-induced expansion of mammary SCs is regulated by the p53-myc axis. Although p21-upregulation and loss-of p53 functions appear critical for the maintenance of the transformed phenotype in leukemias and mammary tumors, respectively, yet they are not sufficient to induce transformation, thus we are also investigating mechanisms of tumor progression of pre-tumoral SCs. In particular, regarding the mechanism of leukemogenesis we are trying to dissect the role of leukemia-associated oncogenes in hematopoietic SCs. These cells are prone to the accumulation of secondary mutations but do not possess growth advantage, resulting in the maintenance of a pool of preleukemic hematopoietic SCs. On the basis that the acquisition of these secondary mutational events might transform the hematopoietic SCs in highly proliferating leukemic SCs, we have been trying to identify putative cooperators of the oncogene PML/RAR in leukemogenesis by high-throughput sequencing of viral integration sites from 48 PML/RAR-dependent MLV infected leukemias. During 2009: i) we identified 264 putative cooperators; ii) we tested 44 putative cooperating genes by gene expression profiling and validated 29/44 genes (65,9%). The validated targets include amongst other known oncogenes as Myc (and Evi1, Myb, Gfi1) and new candidate cancer genes as Rreb1, Adamts2, Cux1; iii) we then tried to identify cellular pathways affected by viral integration by Gene Ontology of 132 targets for which literature data are available and found that 51% of targets controls cellular proliferation, both as positive and negative regulators, and, interestingly, 20% of targets controls stem cell development. Finally, recently it has become apparent that chromatin regulation plays a critical role in determining the fate of stem cells and their descendants. Several studies have also suggested that there is a direct correlation between epigenetic modifications, such as histone methylation, histone acetylation and DNA methylation, and gene expression in disease-relevant cells, including cancer cells. Indeed the epigenetic deregulation in CSCs shows a correlation between genetic mutations and epigenetic changes, as shown for the fusion protein PML-RAR in leukemia or in the epigenetic inactivation of DNA repair genes that have an impact on the stability of the genome. Furthermore, failure to correctly regulate chromosomal DNA replication can lead to uncontrolled growth and genome rearrangements, which are critical features of the transformed phenotype. Our efforts to investigate new mechanisms of chromatin alterations (in particular by leukemia-associate fusion proteins) and de-regulation of DNA replication through the use of genome wide highthroughput analyses, is allowing a comparative genomic analysis of normal tissue and cancer samples, looking for putative chromatin-based biomarkers and the identification

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of new epigenetic drugs. The characterization of cancer stem cells and of the underlying mechanisms of their biological properties is expected to alter the design of clinical research programs and to improve the way we assess the efficacy of novel anticancer drugs. Indeed, the group of Prof Di Fiore (Pece et al., 2010), with whom we have a long standing and productive collaboration, has recently showed that the heterogeneous phenotypical and molecular traits of human breast cancers are a function of their CSC content, so that breast cancers can be stratified on the basis of their biological characteristics (poorly differentiated G3 versus well differentiated G1), in addition suggesting that the p53 gene may be responsible for the variation of CSCs found in different human tumours. Many molecular pathways that are known to be relevant to normal stem cell biology have also been involved in CSC biology (Notch,Wnt, Shh, Bmi1, Pten) but the dividing line between the normal (physiological) and the pathological implementation of these pathways is still unclear. Thus, gaining insight into the biology of CSCs will hopefully allow designing targeted drugs which will not interfere with normal pathways. Although much information has been obtained by studying human samples, mouse model systems are critical to provide the basis for the development of novel CSC-based anti-cancer therapies and new methods for assessing treatment efficacy. The challenge of our group for the next coming years is to develop mouse models tailored to exploit these new areas; we are now improving our model systems and developing new ones to evaluate whether activation of critical checkpoint pathways in stem cells leads to altered tissue homeostasis, organism ageing and increased cancer incidence. Recent research into the mechanism of ageing, cancer and stress has established the biological links between these processes, but their nature remains controversial. Ageing and cancer are regulated by common preserved pathways in which the p53 protein discussed above has a central role. We have focused on the characterization of a novel signaling pathway which involves the p53 tumor suppressor, the p66Shc lifespan determinant and reactive oxygen species (ROS) as signaling molecules, to examine the relationship between checkpoint competence, ageing and the ageing-associated high risk of cancer. Thus, we have been studying p66shc role in normal SC self-renewal: our preliminary results suggest that primary mammary cells from the long-living p66KO mouse model contain increased numbers of SCs compared to WT primary mammary cells from both young and old mice. Analysis of mammary gland morphology during ageing from WT and p66KO mice (3, 10 and 22 month old) showed also marked morphological differences. In particular, mammary glands from p66KO mice of any age were characterized by an increase in ramifications and terminal end buds, in line with the increase in SC frequencies observed in transplantation experiments. Our preliminary data on p66KO cells

also suggest that the protein Sirt1, a histone-deacetylase that functions as life span determinant in flies and worms and is known to modulate p53 activity, may be a candidate mediator of the effect of p66Shc on p53. Thus, mechanistically, p66Shc might regulate the function of p53 on mitotic signature genes through an oxidative stress-dependent effect on Sirt1, by affecting its protein levels, its stability and/or its localization. The physiological cellular outcomes might then be increased cellular senescence, increased apoptosis and/or inhibition of autophagy that, by irreversibly altering tissue architecture, would lead to ageing. Notably, another important physiological role of the p66Shc signalling pathway concerns the regulation of metabolism and the development of the adipose tissue: p66Shc-generated oxidative signals regulate the threshold of sensitivity to insulin in adipocytes, and the development of fat tissue in vivo, suggesting that the regulated generation of ROS has evolved to control energy conservation. Moreover, the findings of reduced adiposity and increased overall insulin sensitivity in p66Shc-null mice might have important implications for the effect of p66Shc on lifespan. We are now starting a new research project aimed at addressing the relation between cancer and obesity, and, more generally, at studying molecular determinants of environmental cancer risk. In summary, the discovery of cancer stem cells opens new directions in the fight against cancer. It will affect the design of clinical research programs and likely improve the way we assess the efficacy of novel anticancer drugs. In this context, the research activity of our group is trying to define the biological and molecular mechanisms that control survival and proliferation of CSCs through studies spanning from regulation of cellular division and genomic stability, damage repair and checkpoint activation to control of DNA transcription and replication by chromatin remodeling. 5. Histone methylation in lineage commitment and carcinogenesis (Giuseppe Testa) The focus of our lab is on the epigenetic mechanisms that enable lineage commitment and their aberrations in cancer. The goal is to elucidate the chromatin regulatory mechanisms that underlie cell fate transitions in both the physiologic and the pathophysiologic setting. Specifically we focus on the methylation of histone H3 on lysine tails 4 and 27, mediated by, respectively, the Trithorax (Trx) and Polycomb (PcG) protein families and antagonized by recently identified histone lysine demethylases (HDMs). These modifications are emerging as key regulators for the programming of genomes that underlies the establishment and maintenance of differentiated cell states. Not surprisingly, aberrations in these pathways are also being recognized as important determinants or modulators of tumors, hinting at common regulatory circuits that preside over stem cell physiology and that are perturbed or hijacked in oncogenesis. Hence,

we pursue two complementary lines of research that investigate PcG and Trx function in: i) the physiology of genome programming during differentiation; and ii) the aberrant genome programming that accompanies tumorigenesis. i) Polycomb and trithorax in the physiology of genome programming The connection between histone lysine methylation and developmental fate became apparent with the realization that Ezh2, a member of the Polycomb group (PcG) of proteins first discovered in the fly as stable repressors Hox genes, catalyzes the trimethylation of histone H3 on lysine 27 (H3K27me3) while Trx (and its mammalian homologs of the Mll family), identified in the fly as a stable activator of Hox genes, catalyzes the trimethylation of histone H3 on lysine 4 (H3K4me3). In ES cells and some adult stem cells most PcG target genes are kept in a repressed state but poised for activation by a bivalent chromatin signature that features both H3K4me3 and H3K27me3. Upon differentiation many of these bivalent domains are resolved and their genes become either completely active (marked solely by H3K4me3) or definitely repressed (marked solely by H3K27me3) in a lineage specific fashion. Contrary to the long held assumption that histone lysine methylation (HLM) was irreversible and that this irreversibility underlined lineage stability, research over the last years revealed the existence of histone lysine demethylases (HDMs) as key effectors of the dynamic regulation of HLM, suggesting that the establishment and maintenance of cell lineages involves a regulated process of addition and removal of methyl marks. In particular, following the identification of Jmjd3 as a histone H3 lysine 27 demethylase (De Santa et al. 2007), we determined that it is required for the neural commitment of ES cells (Burgold et al. PLoS One 2008). It is noteworthy that much of the current knowledge of global changes in chromatin and cell fate transitions, including the seminal discovery of bivalent domains, derives from a comparison between ES and NS cells, points respectively of departure and arrival for neural commitment, with similar ‘steady-state’ levels of Jmjd3 and several other HDMs. Our focus on the early stages of neural commitment allowed us to identify Jmjd3 as a H3K27 demethylase that is required for the onset of differentiation, underscoring the importance of defining the precise developmental windows and epigenetic switches that enable cell fate transitions. Our finding that Jmjd3 regulates key neural genes at the onset of neurulation by binding to their promoters and resolving their bivalent domains affords now the possibility of modulating H3K27me3 levels throughout neurogenesis and assess the relevance of their dynamics both in vitro and in vivo. o this end we generated over the last year several tools that enable a precise and flexible modulation of the PcG axis during both fetal and adult neurogenesis. In fact, sevearal lines of convergent evidence

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indicate that the dynamics of H3K27me3 and PcG action are not limited to the exit from the pluripotent state. In the transition from ES cells to neural progenitors to terminally differentiated neurons, for example, a recent analysis showed that at each stage new gene subsets acquire or lose the PcG and the DNA methylation mark, revealing an unexpected degree of plasticity in PcG recruitment and suggesting that H3K27me3 is not a simple switch between pluripotency and commitment but rather accompanies the entire process of fate acquisition. Consistenly with this dynamics, we determined that Jmjd3 is progressively upregulated during neural development and we are currently probing its speciﬁc function in corticogenesis. ii) Polycomb in the aberrant genome programming of cancer This line of research investigates the relevance of H3K27 methylation in the pathological counterpart of the experimental system described above, namely the aberrant or blocked differentiation that characterizes gliomagenesis. In fact, consistent with the role of PcG in lineage choices, alterations in H3K27me3 are likely to be early events in the cascade of epigenetic aberrations of cancer, particularly the hypermethylation of CpG promoters that is an important mechanism of tumor suppressor inactivation. Convergent lines of evidence indicate that CpG hypermethylation in cancer cells is the result of an instructive process through which altered developmental programs determine aberrant hypermethylation at multiple loci. Throughout development de novo DNA methylation proceeds in an instructive fashion and the majority of genes that are hypermethylated in cancers are pre-marked by H3K27me3 in ES cells. The recent observation that most epithelial cancers share a core transcriptional signature with ES cells corroborates this model and suggests that the PcG-dependent gene expression program that orchestrates development in normal cells is hijacked in cancer cells as the main template for cancer DNA methylation. Finally, the oncogenic role of H3K27me3 imbalances is underscored by the direct involvement of various PcG members in human and experimental tumors, of which the two best characterized examples are Ezh2 and Bmi1, the component of Polycomb repressive complex 1 (PRC1) that stimulates ubiquitination of histone H2A lysine 119 (H2AK119) following H3K27me3 by Ezh2. Hence, this line of research in the lab explores the proposition that loss of the physiologic regulation centered around H3K27me3 is important for the initiation and/or maintenance of tumors, combining the conditional modulation of this epigenetic axis in advanced models gliomas and glioblastomas that recapitulate faithfully the pathogenesis of the corresponding human tumors.

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II. Aberrations of cell division 6. The spindle assembly checkpoint as a therapeutic target in cancer (A. Musacchio) Our group is interested in the molecular mechanism of the spindle assembly checkpoint (SAC) and of its interaction with kinetochores. Kinetochores are large protein scaffolds that mediate the interaction of the mitotic chromosomes to the mitotic spindle. It is through this interaction that the replicated chromosomes are equally parted during mitosis. The SAC proteins are recruited to mitotic kinetochores to monitor the correct attachment and alignment of chromosome to the mitotic spindle, and they arrest cell cycle progression until the attachment process has been successfully completed. Errors in the attachment process in a mother cell will affect the distribution of chromosomes to the daughter cells, causing them to become aneuploid or polyploidy, and the SAC protects from aneuploidy. Chromosome instability is particularly frequent in cancer cells, but the underlying causes have remained largely unclear. It was initially proposed that a compromised SAC in cancer cells might cause chromosome instability by preventing cells to monitor the state of kinetochore-microtubule attachment. However, several subsequent studies failed to support this idea. Rather than abrogated, the SAC appears to be usually functional in cancer cells, and a growing body of evidence indicates that the impairment of the SAC is generally lethal for cancer cells. Recently, it has been shown that the presence of supernumerary centrosomes, a very frequent feature in tumors, is a sufficient cause for chromosome instability. When entering mitosis, cells with supernumerary centrosomes first face the formation of a multipolar spindle. In the majority of cell divisions, multipolarity is suppressed by a pathway involving the HSET motor and the SAC. The latter allows the additional time required for spindle bi-polarization in cells with multiple centrosomes relative to normal cells. These observations suggest the possibility that inhibition of the SAC will drive cells with multiple centrosomes prematurely out of mitosis, causing their death. To explore these ideas, we are active working on the identification of strategies for targeting the SAC. In 2009, we have thoroughly characterized as small molecule named Reversine. Early studies identified Reversine as an Aurora B inhibitor, but work in our laboratory finally demonstrated that Reversine is a potent and relatively selective MPS1 inhibitor. MPS1 is a crucial component of the SAC, and our studies are elucidating the mechanism of action of this kinase and its interaction with additional checkpoint kinases, including Aurora B, BUB1 and BUBR1.We are also exploring the complex interaction of the SAC with kinetochores. Specifically, we are studying the mechanism of recruitment of SAC proteins to the KMN network, a 10-protein complex that acts as a microtubule receptor at the kinetochore. We have

reconstituted an 11-subunit assembly containing the KMN network and an additional interaction partner, and are using this material in vitro to identify sites of phosphorylation that may be important for SAC recruitment. III. Host-tumour interactions 7. Chronic inflammation and cancer: transcriptional mechanisms as potential therapeutic targets (G. Natoli) Epidemiological and experimental data demonstrated a direct link between chronic inflammation and the development of several types of cancers. Moreover, cancers often contain an inflammatory infiltrate that is hijacked by tumor cells to promote angiogenesis, tissue invasion and cell proliferation. In vivo experiments in mouse models have demonstrated that modulation of tumor properties by inflammatory cells requires the transcription factors of the NF-kB family, which are essential to mount the transcriptional program underlying the inflammatory response. In turn, this has led to the suggestion that anti-NF-kB drugs may be used in tumor therapy. Drugs disabling the whole NF-kB signaling pathway are already being tested in clinical trials, but several concerns about their safety have been raised because of the many physiological responses in which NF-kB is required, most notably the defense against microbial infections. Conversely, therapies blocking the induction of subsets of inflammatory genes relevant to disease (and specifically cancer) pathogenesis regulated genes should provide more restricted and predictable effects, but the molecular bases for their design are not available. Our unit is interested in understanding the mutual relationships between the transcriptional response underlying the inflammatory response and the epigenome, namely the collection of chromatin modifications that maintain and propagate across mitosis the program of gene expression characteristic of a given cell type. On the one hand chromatin controls recruitment of transcription factors to inflammatory genes and generates tissue-specific and temporal profiles of inflammatory gene expression. On the other, inflammation affects epigenetic control in bystander tissue cells, thus leading to alteration of tissue differentiation in chronically inflamed tissues (e.g. intestinal metaplasia in chronic gastritis and Barrett’s esophagus in reflux esophagitis). Such alterations often precede and are associated with the development of malignant tumors. We have identified a number of enzymes acting on chromatin and regulated by inflammatory stimuli that may provide both a mechanistic link between inflammation and altered differentiation, as well as potential targets for pharmacological intervention. One such gene is Jmjd3, which is inducible by inflammatory stimuli and erases a histone covalent modification that controls cellular differentiation by keeping repressed the genes that specify alternative cell fates (trimethylation of histone H3 at lysine 27). Additional

enzymes involved in writing or erasing histone modifications were found to be regulated by inflammatory stimuli or anyway involved in inflammatory gene expression, including H3K4 demethylases. For each of these molecules we are defining the genomic distribution in inflammatory and bystander cells, as well as the consequences of their deletion, both in vitro and in mutant animal models. For Jmjd3, we have found that it is extensively associated with the transcription start sites of most active or poised genes in macrophages, and that it contributes to tune their regulation (being however dispensable for their induction, as indicated by the analysis of macrophages obtained from the Jmjd3-/mice we have generated). Most importantly, the effects we have seen are largely independent of H3K27me3 demethylation, suggesting the possibility that this hydroxylase has additional substrates that contribute to its biological functions. An additional area of research tackled by the laboratory related to the identification of genomic regulatory elements (enhancers) controlling inflammatory gene expression. This activity involved the extensive use of chromatin immunoprecipitation coupled with high-throughput sequencing and allowed us to determine a general organizational principle of these enhancers, which consists in the combination of binding sites for ubiquitous, stimulus-responsive TFs and binding sites for costitutive cell type-restricted and lineagedetermining TFs. This combination allows creating a cell type-specific context within which transcription of inflammatory genes is regulated, thus explaining variability among cell types in the inflammatory gene expression program induced by identical stimuli. IV. Predictive markers 8. Predictive value of p53 gene status in breast carcinoma patients treated with taxanes-based adjuvant chemotherapy (G. Viale) We have retrieved additional 178 tumor tissue samples from the patients enrolled in a randomized clinical trial evaluating the possible benefit of the addition of taxanes to anthracyclin-based adjuvant chemotherapy, reaching a total of 2176 evaluable patients. All the tumor samples have been centrally re-evaluated for the expression of estrogen (ER) and progesterone receptors (PgR), and of HER2 status. We have also immunostained for the p53 protein all the cases, looking for the nuclear accumulation of this protein that is associated with the occurrence of gene mutations. The immunohistochemical reaction has been performed using the D07 monoclonal antibody (Dako, Glostrup, Denmark), which recognizes an epitope shared by the wild-type protein and by its mutated forms. The detection step of the reaction has been carried on using an high-sensitive reagent (EnVision, Dako). Some 40% of the cases did show nuclear accumulation of the p53 protein. DNA has been extracted from 673

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cases, and sequencing of the p53 gene has been already performed for 521 cases. Mutations have been detected in 85/521 (16.3%) cases. The correlative analyses of mutational status with clinico-pathological variables are currently ongoing. 9. Use of primary human breast tumor cells as an ex-vivo model to evaluate the effects of HDAC inhibitors (HDACi) on different cellular phenotypes (S. Chiocchia, S. Minucci) Histone deacetylases (HDACs) are a class of modification enzymes that catalyze the removal of acetyl molecules from e-NH3 groups of lysines. The more abundant substrates are histones, but it is becoming more and more evident that they deacetylate a broad spectrum of proteins. Due to the deep involvement of HDACs in cellular proliferation, inhibitors of HDACs (HDACi) have been looked at as promising anti-cancer drugs since many years. In 2006 SAHA (commercially known as vorinostat) was the first HDACi to be FDA approved for treatment of advanced T-cell cutaneous lymphoma. We have been obtaining breast primary tumor specimens from the Surgical Division of the IEO and culturing them following established procedures. In parallel, a complete analysis in collaboration with IEO Pathology Division (Dr. Michele Masullo and Prof. Giuseppe Viale) is performed on the primary tumor immediately after excision, to establish global levels of histone acetylation and other tumor properties. The in vitro cultures are tested for their sensitivity to HDACi. Based on previous results, we expect that depending on the global acetylation level of histone H4tumors with low-grade of acetylation (“hypo-acetylated”) will respond to HDACi undergoing apoptosis, whereas tumors with high-grade of acetylation (“acetylated”) will be insensitive to HDACi. Gene expression profiles on these two classes of tumors (hypo-acetylated and acetylated) will then be performed to establish a signature for responsivness to treatment with HDACi.

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Biological Resource Center and Xenotransplantation Unit

In the next future we will assist important changes in the diagnostic and therapeutic ﬁeld, driven by biomedical research. The IEO bio bank (Biological Resource Center (IBRC)) is a prestigious element of our Institution and a necessary step for the realization of the Molecular Medicine program recently developed in our Campus. The IBRC, was established in order to collect tissue that following surgery is in excess of what is required for diagnosis and would otherwise be discarded. The Bio bank provides biospecimens prepared from a collection of blood and tissue samples, taken with consent from healthy individuals and patients diagnosed with cancer. Some information about the health and treatment of the donors is also collected and used for epidemiology studies. IEO, through the Bio bank, has the necessary infrastructure to provide high quality biospecimens required for biomarker discovery and xenotransplantation experiments. The advantage of transplanted human tumors in immunodeﬁcient mice (NOD / SCID), enables us to grow the tumor (which retains the morphology and biological features of the tumor of origin) and treat the animal with new drugs that will be available in the future, in order to directly dissect the pharmacological effects of drugs or combination therapies on tumor growth in vivo and design personalized therapies directed to single patients. This innovative and collaborative project will give researchers the unique opportunity to translate their frontline discoveries into practical, faster and more accurate diagnosis for cancer, which in turn will enable better treatment outcomes for patients.

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The Tumour Registry

The Tumour Registry (TR) was activated in March 2006 with the aim to collect data on all those consulting at the European Institute of Oncology (IEO), at risk of developing or already presenting with a tumour. It has actually become a supporting tool for the current practice as well as for epidemiological/basic research, guaranteeing a quick analysis of the IEO clinical activity and playing a key role in the production of scientific publications. Eligible to enter the Registry are all those coming to the IEO for consultation since its opening, with unique identification number (patients’ record) and at least one episode accessible from Institute’s intranet. A minimum data set of variables was defined and data entry was divided in 4 forms. On the first form personal data (i.e., sex, address, date of birth) and information on follow-up (i.e., date of last contact, date of last visit, vital status, cause of death) are recorded. The following types of record are assigned: 1. Visit only: a healthy individual comes to IEO for either visit or genetic counselling 2. Anamnesis: the patient, at the moment free of disease, reports on a tumour diagnosed in the past and already treated and cured. 3. Diagnosis only: diagnosis of tumour is made at IEO. The patient decides to be treated elsewhere. 4. Second Opinion: the patient or the patient’s parents come to IEO and ask for a second opinion on a diagnosis and/or a treatment proposed elsewhere. 5. Long: the patient receives at least one treatment at IEO. Detailed information on patient’s tumour(s) (i.e., date of diagnosis, morphology, topography, TNM staging) is recorded on the second form, together with some epidemiological information (i.e., familiarity, height and weight at diagnosis and smoking habits). The third form is dedicated to the treatment strategy, where every therapy is classified as administered or proposed. The fourth is dedicated to the chronology of events, in order to better describe the history of the disease and to take note of the tissues preserved in the Biobank of IEO.

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Sources of information for data collection are: 1. database of patients’ administrative data (personal information is automatically downloaded); 2. files accessible on intranet; 3. online databases (surgery, laboratory medicine); 4. patients’ clinical dossier digitalized and accessible on e-Paper. The Registry was implemented using the interface software Argos™, based on Oracle™ database system. The implementation, completed in February 2006, was managed in the division of Epidemiology and Biostatistics, with the help of the IT division. Although it was decided that data from the TR must not be directly accessible to researchers or clinicians, we have established a system to collect and answer requests. Data extrapolation and analysis are managed in the division of Epidemiology and Biostatistics on the advice of the Scientific Direction of IEO. For detailed information on the TR structure, materials and methods please refer to a recent paper published in the IEO Journal ecancermedicalscience: “Case mix at the European Institute of Oncology: first report of the Tumour Registry, years 2000-2002” [Botteri E, Iodice S, Maisonneuve P, Alfieri M, Burzoni N, Manghi L, Martinetti M, Montanari B, Albertazzi E, Bazolli B and Rotmensz N. doi: 10.3332/ecancer.2009.149] The First Four Years Of Activity After a 6 months pilot period, from March to August 2006, which involved the training of the operators, ad hoc improvements to the structure of the registry, data quality control and editing of the user guide, from September 2006 the data entering has been running at top speed. We started entering individuals who came for the first time to IEO in the year 2000 (dossier number CC00) in a sequential fashion. By February 2010, 117,645 individuals who visited IEO for the first time in the years 2000-2004 were entered in the Tumour Registry. These were not necessarily presenting

with a tumour. In Table 1 some characteristics of these individuals are reported.

Table 1. Characteristics of individuals a

Classiﬁcation

No. Patients (%)

< 20 years

1,800 (1.5)

20-34 years

13,812 (11.7)

35-49 years

36,190 (30.8)

50-64 years

41,263 (35.1)

65-79 years

22,644 (19.2)

> 80 years

1,936 (1.6)

Men

36,471 (30.1)

Women

81,174 (69.0)

Long

28,231 (24.0)

Second Opinion

36,235 (30.8)

Anamnesis

1,872 (1.6)

Diagnosis

1,285 (1.1)

Visit

50,022 (42.5)

Age

Sex

Type of record

Total a

117,645

Individuals who came to IEO for the ﬁrst time in the years 2000-2004

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Table 2. Tumors by site a Tumour site

Collectedb Overall

Total

Invasive tumors IEO diagnosis

IEO Surgery

Head and Neck

2173

1897

537

387

Lip

Tongue

378

360

158

143

Major salivary glands

412

212

Gum

Floor of mouth

Other and unspeciﬁed parts of mouth

326

302

Oropharynx

301

293

Nasopharynx

259

257

Hypopharynx

151

150

Other and other ill deﬁned sites Digestive organs and peritoneum c

182

174

11437

11222

1895

1106

Oesophagus

323

315

Stomach

2014

1993

373

272

Small intestine, including duodenum

197

192

Colon

4092

4001

649

441

Rectum, rectosigmoid junction and anus

1755

1729

351

264

Liver and intrahepatic bile ducts

869

852

124

Gallbladder and extrahepatic bile ducts

635

617

Pancreas

1352

1340

175

38 6

Retroperitoneum and peritoneum

Other and ill-deﬁned sites

104

Respiratory and intrathoracic organs

8961

8603

2071

1322

Nasal cavities, middle ear and accessory sinuses

Larynx

1053

934

290

262 1000

Trachea, bronchus and lung

7316

7116

1628

Pleura

341

327

Thymus, heart and mediastinum

197

173

Other and ill-deﬁned sites Bone, connective tissue, skin and breast Bone and articular cartilage Connective and other soft tissue

31269

25728

13024

8782

1639

1536

558

215

Skind

3775

3571

1571

979

Breast

25839

20609

10892

7587

Genitourinary organs

14289

11910

3005

2061

Cervix uteri

2408

998

369

269

Uterine corpus

1073

945

292

212

Ovary and other uterine adnexa

2131

1942

637

355

Other and unspeciﬁed female genital organs

483

341

119

Prostate

4348

4270

887

603

Testis

476

441

124

Bladder

1611

1320

255

204

Kidney

1501

1433

278

224

Other and ill-deﬁned sites

187

162

Other and unspeciﬁed sites

2664

2196

367

285

Penis and other male genital organs

Eye

Brain

1137

1120

Other and unspeciﬁed parts of nervous system

1374

966

319

277

Thyroid gland Other endocrine glands and related structure Lymphatic and haematopoietic tissue

112

2413

2390

829

Hodgkin lymphoma

437

435

141

Non-hodgkin lymphoma

1297

1293

553

Multiple myeloma

299

Leukemia

348

Other and unspeciﬁed

Non-skin melanoma

Malignant neoplasm without speciﬁcation of site Total

190 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

1274

1242

298

74560

65268

22045

14038

By February 2010, 74,560 tumours were entered (out of 68,480 individuals presenting with 1 or more tumours). A synthesis is reported in Table 2. Developments A recent paper entitled “Analysis of local and regional recurrences in breast cancer after conservative surgery” represents the ﬁrst published study conducted on the data of the TR [Botteri E, Bagnardi V, Rotmensz N, Gentilini O, Disalvatore D, Bazolli B, Luini A, Veronesi U. Annals of Oncology 2009; doi: 10.1093/annonc/mdp386]. It consisted of a multi stage analysis of local, regional and distant recurrences, performed on data of 2,784 women treated for early breast cancer by quadrantectomy and whole breast irradiation at IEO. Besides this paper, many other studies and projects based on the TR data are now arising from the collaboration of our and other divisions of the Hospital. The IEO TR has been proven functional and reliable in monitoring the activity of the Hospital, allowing extraction of data from any subpopulation with characteristics of interest. This structured and centralized Registry represents an important tool for our research-oriented Institution.

Among individuals who presented at IEO for the ﬁrst time in years 2000-2004; bIncluding all invasive and in situ tumors. Benign neoplasia or negative histology after radical surgery in IEO are also collected. cPolyps are not collected; dBenign nevi are not collected.

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Cancer Prevention and Genetics

Our current lines of research are focusing mostly on chemoprevention, pharmacogenomics and the effects of lifestyle changes in various study cohorts of subjects at higher risk of developing cancer in order to validate the most effective strategies to reach a tailored prevention. Scientiﬁc aims: 1. Chemoprevention in subjects at higher risk for ER negative breast cancer, like patients with endocrine non-responsive, resected breast cancer and DIN, and BRCA1 mutation carriers . In particular we are studying two classes of drugs, NSAIDs and Statins. 2. The possible role of raloxifene and very low dose tamoxifen in premenopausal women and exemestane and celecoxib in postmenopausal women, as chemopreventive agents in a pre-surgical model. 3. Optimizing the dose of tamoxifen and selecting the population who can beneﬁt most from tamoxifen as chemopreventive agent, through pharmacogenomics studies on Polyporphisms (SNPs), particularly on CYP2D6. 4. The role of aromatase inhibitors in prevention, evaluated in pre and post surgery settings. 5. The IGF system and sexhormones as cancer risk factors and surrogate intermediate biomarkers of response to chemopreventive agents. 6. Ductal lavage as a risk assessment tool and source of new biomarkers studies. 7. The role of budesonide as a putative chemopreventive agent in subjects at high risk for lung cancer (current and former heavy smokers). 8. A biomarkers study with allopurinol in subjects with adenoma of the colon. 9. Long term effects of radiotherapy in BRCA mutation carriers. 10. Chemoprevention of breast and ovarian cancer with Fenretinide in young healthy BRCA mutation carriers. 11. Diet and physical activity to prevent recurrences after standard treatment in women with invasive breast cancer. Ongoing and starting projects: 1. A randomized phase II prevention trial with anti-COX2 vs.

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statins in subjects at high risk for hormone non-responsive breast cancer 2. IBIS II - an international multi center study of tamoxifen vs anastrozole in post menopausal women with Ductal Carcinoma in Situ (DCIS) 3. IBIS II - an international multi center study of anastrozole vs placebo in post menopausal women at increased risk of breast cancer 4. The HOT Study: Hormone Replacement Therapy and low dose Tamoxifen. A phase III trial of breast cancer prevention with low dose tamoxifen in HRT users 5. Pre-surgical study: to investigate the biological activity of metformin in all breast cancer histotypes. The primary endpoint is the KI67 modulation; several other secondary endpoints will be evaluated to study the interaction of the glucose metabolism and breast cancer. 6. The role of radiotherapy in carriers of BRCA mutations after conservative surgery. 7. A phase III study with fenretinide (4-HPR) for primary prevention of breast cancer in subjects at high risk for familial/ hereditary breast cancer. 8. A randomized placebo-controlled phase III clinical trial with low dose tamoxifen (5 mg day) in women with ER positive intraepithelial neoplasia of the breast. 9. DIANA(DIet and ANdrogens)-5: randomized controlled trial to test the efﬁcacy of dietary change and physical activity to prevent or delay the recurrences in breast cancer patients estimated to be at higher risk based on their metabolic milieu.

Major achievements and publications Randomized biomarker trial of anastrozole or lowdose tamoxifen or their combination in subjects with breast intraepithelial neoplasia In the ATAC trial, the combination arm was inferior to anastrozole alone in terms of disease-free survival possibly due to an adverse pharmacokinetic interaction or a predominant estrogenic effect of tamoxifen under estrogen deprivation. We assessed whether the addition of a lower dose of tamoxifen

inﬂuenced anastrozole bioavailability and favorably modulated biomarkers. Anastrozole concentrations were not affected by the combination with low-dose tamoxifen, whereas endoxifen levels were lower in poor CYP2D6 metabolizers only. The combination arm had a positive modulation of biomarkers of bone metabolism (statistically signiﬁcant reduction of C-telopeptide and osteocalcin compared to anastrazole alone). Compared with anastrozole, the combination arm showed lower IGF-I/IGFBP-3 levels (-17% versus -9%; P = 0.004) and lower estradiol/SHBG and estrone sulfate reductions (-15%

versus -29% and -30% versus 38%, respectively). However, IGF-I/IGFBP-3 and estradiol/ SHBG did not decrease in poor CYP2D6 metabolizers. Endometrial thickness was not greater in the combination than in the anastrozole arm. These results suggest that with this low dosage of tamoxifen the combination may be beneﬁcial also to better balance the side effects of the single agents (ﬁgure 1).

Fig. 1) Effects of tamoxifen, anastrozole and their combination on drugs concentration and bone metabolism biomarkers.

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A randomized double-blind 2x2 trial of low-dose tamoxifen and fenretinide for breast cancer prevention of high risk premenopausal women Tamoxifen and fenretinide are active in reducing premenopausal breast cancer risk and work synergistically in preclinical models. We assessed their combination in a two-by-two biomarker trial. A total of 235 premenopausal women (pT1mic breast cancer 21, IEN 160 or 5-year Gail risk > 1.3% 54, respectively) were randomly allocated to either tamoxifen 5 mg/d, fenretinide 200 mg/d, their combination, or placebo. During the 2-year intervention, tamoxifen signiﬁcantly lowered IGF-I and mammographic density by 12% and 20%, respectively, fenretinide by 4% and 10% (not signiﬁcantly), their combina-

tion by 20% and 22%, with no evidence for a synergistic interaction. The annual rate of breast neoplasms (n = 48) was 3.5% + 1.0%, 2.1% + 0.8%, 4.7% + 1.3%, and 5.2% + 1.3% in the tamoxifen, fenretinide, combination, and placebo arms, respectively, with hazard ratios (HRs) of 0.70 (95% CI, 0.32 to 1.52), 0.38 (95% CI, 0.15 to 0.90), and 0.96 (95% CI, 0.46 to 1.99) relative to placebo (tamoxifen x fenretinide adverse interaction P = .03). Both single agents, particularly fenretinide, showed numerical reduction in annual odds of breast neoplasms (ﬁgure 2).

Fig. 2) Modulation of IGF-I and mammographic density by tamoxifen, fenretinide and their combination

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Ductal intraepithelial neoplasia: post-surgical outcome for 1267 women cared for in one single institution over 10 years This was an observational study, a preliminary analysis of the role of chemoprevention in patients with a hormoneresponsive intraepithelial neoplasia. A total of 309 patients with DIN received low-dose tamoxifen as part of institutional guidelines and were compared with 371 patients with DIN who received no systemic treatment after surgery. Women with estrogen receptor and progesterone receptor >50% DIN who were not treated had a higher incidence of breast events than women on tamoxifen [hazard ratio (HR) 1.76; 95% conﬁdence interval (CI) 1.00–3.12] or women with ER/PgR <50% DIN (HR

1.72; 95% CI 1.14–2.58). High ER and high PgR expression is a significant adverse prognostic indicator of DIN, and low-dose tamoxifen appears to be an active treatment. Women with low-expression ER or PgR DIN do not seem to benefit from tamoxifen. A definitive clinical trial is warranted. This observation has been confirmed in a subsequent publication with a longer follow-up and more selected population (available online) (figure 3).

Fig. 3) Low-dose tamoxifen to prevent recurrence of invasive and intraepithelial breast cancer in women with Er positive DIN

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Circulating endothelial mature cells and progenitors as biomarkers for the anti-angiogenic therapy of cancer The new class of anti-angiogenic drugs is among the few that have entered the clinical oncology arena and gained approval for the use in patients during the past decade. The anti-VEGF monoclonal antibody bevacizumab and three small molecules designed to target a variety of tyrosine kinases involved in angiogenesis (namely sunitinib, sorafenib and pazopanib) have been approved for the therapy of colo-rectal, lung, breast, kidney, liver and central nervous system cancer, but their clinical development is still hampered by relevant drawbacks (Kerbel, 2008). The progression-free and/or overall survival advantage observed in randomized clinical trials involving these drugs was limited to a few months, at the expenses of a significant economic cost. Considering the current budget constraints, and the presence of some potentially serious side effects in recipients, it seems mandatory to identify and use markers able to select patients for whom these drugs are likely to be effective, and at the cost of manageable side effects. The endothelial cell turnover has always been thought to be very slow compared with other tissues. This notwithstanding, cells with and endothelial morphology were found to circulate in the blood more than 35 years ago (Hladovec et al., 1973). In the following years, the endothelial nature of these cells was confirmed by immunohistochemistry (IHC) studies, and their enumeration by means of positive enrichment, IHC or flow cytometry (FC) indicated that circulating endothelial cells (CECs) are increased in a very wide spectrum of disorders encompassing vascular, autoimmune, infectious and ischemic diseases (Moldovan et al., 1994; Bertolini et al., 2006). Over the past ten years, increased CEC counts were observed in some cancer patients (Mancuso et al., 2001; Farace et al., 2007), and these cells were studied as surrogate biomarkers of angiogenesis and anti-angiogenic drug activity in preclinical models and medical oncology (Monestiroli et al., 2001; Shaked et al., 2005; Calleri et al., 2009). These studies also indicated that the endothelial phenotype was expressed by cells displaying a wide variety of different features (Blann et al., 2005; Bertolini et al., 2006). Some CECs had a phenotype compatible with terminally differentiated endothelial cells (EC), in some cases being apoptotic or necrotic and thus most likely derived from the turnover of vessel walls. Some other cells expressed progenitor-associated antigens in addition

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to endothelial antigens, and were considered as circulating endothelial progenitor (CEP) candidates. CEPs and CECs phenotype. There is a lack of consensus regarding the surface markers that identify CEPs and CECs, due to the lack of a marker that can unambiguosly identify these cells. By means of flow cytometry, the most widely used antigens to identify CEPs are CD34, VEGFR2 and CD133. Many investigators identify and enumerate CEPs by means of the coexpression of CD34 and VEGFR2 (Rosenzweig, 2005; Bertolini et al., 2006; Asahara et al.,1997; Shi et al., 1998), but these antigens are expressed also by mature CECs. The sole antigen that according to some authors is expressed on progenitors but not on mature endothelial cells is CD133. Unfortunately, in humans CD133 is expressed also by hematopoietic stem cells (HSCs), and other progenitors from neural and gastro-intestinal tissues (Mizrak et al., 2008). Moreover, the use of CD133 for CEP identification (Peichev et al., 2000; Rafii et al., 2002) has led to the isolation of cells that not all laboratories were able to differentiate in vitro and in vivo along the endothelial lineage (Yoder et al., 2007; Case et al., 2007). More specific markers are thus needed to distinguish CEPs from CECs and HSCs. Even more controversies exist for the enumeration of CEP in mice, because the expression and function of CD34 and CD133 antigens are less well characterized in mice than in humans. A candidate CEP phenotype in mice is CD45-, VEGFR2(Flk)+, CD117+ (Monestiroli et al., 2001; Shaked et al., 2005), and antibodies reacting with a particular configuration of CD144 are also used (Nolan et al., 2007; Gao et al., 2008). This controversy regarding CEP phenotype in mice has led in the past to opposing conclusions regarding CEP contribution to cancer vasculogenesis, with some studies indicating a crucial role for CEPs and some others failing to observe a relevant CEP contribution (reviewed in Madlambajan et al, 2009). Antigenic promiscuity between CECs and platelets has prompted the development and validation of new FC enumeration procedures were DNA staining reagents have allowed the enumeration and isolation of platelet-depleted, DNAcontaining cells with a EC phenotype (CD45-,CD31+CD146+) (Mancuso et al., 2009). Transmission electron microscopy (TEM) and RT-PCR of sorted CECs confirmed their endothelial

nature by virtue of the presence of EC-specific Weibel-Palade bodies and of RNA transcripts for the EC-specific gene VE-cadherin. TEM studies also offered an explanation of the controversies about CEC frequency in the blood. The majority of sorted CECs, in fact, were found to be apoptotic or necrotic cellular fragments, most likely lost at count after the cell processing involved in IHC enumeration. Along with apoptotic CECs, however, TEM showed the presence of small, viable and lymphoid-like cells that are compatible with a progenitor cell morphology (Fig.1). TEM will most likely be of help for the next crucial steps in CEC and CEP studies, ie to dissect the functions of candidate CEC and CEP subpopulations. Both these cell families, in fact, encompass subpopulations with different roles. Multiparametric FC has shown that among DNA+,CD45-,CD31+,CD146+ CECs there are some expressing other EC-related antigens such as CD143, CD144, VEGFR1, VEGFR2, VEGFR3, activation antigens such as CD105 (endoglin), and others (Bertolini et al., 2006). CEC number and viability in cancer and treatment Endothelial cell enumeration (both CECs and CEPs) has led to the observation that these cells are increased and are more viable in some types of cancer patient compared to healthy controls (Mancuso et al., 2001; Farace et al; 2007). In breast cancer, CECs and CEPs demonstrated a strong relationship with the Nottingham prognostic index (NPI), but only CECs positively predicted higher NPI scores and correlated with tumor invasiveness and size, possibly reflecting total tumor vascular volume (Goon et al., 2009). Possible explanations for these findings involve the angiogenic switch associated with cancer growth and the robust production of angiogenic growth factors such as VEGF, bFGF, HGF and many others by cancer cells and/or various host cells (Bertolini et al., 2006). The recent and unexpected finding of an autocrine loop in EC (Lee et al., 2007) is of particular interest, because it might be that the increase of viable CECs in the blood of cancer patients mirrors an aberrant vascular turnover/remodeling associated with high local levels of VEGF produced by cancer cells. Following the preclinical evidence that CEC count can be used as a surrogate marker of angiogenesis and of anti-angiogenic drug activity and to define the optimal biological dosage of anti-angiogenic drugs (Shaked et al., 2005; Shaked et al., 2005b), CEC number and viability have been measured in different clinical trials where cancer patients have been treated with anti-angiogenic therapies (Rabascio et al., 2004; Farace et al., 2007). An increase in the number of apoptotic CECs after 60 days of therapy was associated with a prolonged progression-free survival and overall survival in metastatic breast cancer patients treated with a doublet metronomic chemotherapy regimen (Mancuso et al., 2006). When the anti-VEGF antibody bevacizumab was added to metronomic chemotherapy for the therapy of metastatic breast cancer, patients

who had a clinical response (as well as the larger population of patients who had a clinical benefit from the treatment) had significantly greater baseline levels of viable CECs than patients who did not respond to therapy. Also, the number of apoptotic CECs before the beginning of therapy was associated with a prolonged progression-free survival (Dellapasqua et al., 2008). The finding that in long term responder patient at the time of progression there was a statistically significant decrease in CEC count might suggest a switch toward a different vascularization of the neoplastic lesions. It must be noted that the decrease in CEC number was associated with an increase in the level of VEGF-A and b-FGF when compared to baseline-values (Calleri et al., 2009). More pre clinical studies are now needed to understand whether such switch is toward the cooption of more stable vessels from the non-neoplastic tissue in an independent VEGF mechanism, or toward a new type of neoangiogenesis involving different vessel, more stable and with lower turnover (Adighibe et al., 2006; Bergers et al., 2008; Kilanski et al., 2009). In patients with renal cancer treated with the anti-angiogenic drug sunitinib, CEC kinetics were different in patients with clinical benefit compared to those with progressive disease (NordenZfoni et al., 2007). Taken together, these data suggest that the investigation of CEC number and viability by FC has potential for the stratification of cancer patients who are more likely to benefit from anti-angiogenic treatments (Bertolini et al., 2006; Schneider et al., 2005; Kerbel et al., 2002). CEPs in tumor grow and metastases There are different lines of research studying the role of CEP in early and late stage of cancer development and metastasis. In the late ‘90, (Asahara et al., 1997; Shi et al., 1998) demonstrated that human peripheral blood contains cells that are able to differentiate into endothelial cells. However, the first proof of the contribution of CEPs in tumor development was reported by Lyden et al (2001) in Id1 deficient mice. These mice have a severe CEP defect and show impaired angiogenesis, tumor growth, and progression of micrometastases to macrometastases, suggesting a critical role for these cells in angiogenesis-mediated tumor growth. More recently it was found that CEP generation depends on the ability of Id1 to restrain the expression of its target gene p21 (Ciarrocchi et al., 2007). Genetic ablation of p21 rescued CEP generation in these Id1 deficient mice, re-establishing normal tumor growth (Ciarrocchi et al., 2007). Since the first description of CEPs, their identification and role in tumor vasculature has often been debated (Gao et al., 2009). Understanding whether or not all types of tumor rely – at least in part – on CEP dependent vessel generation has been elusive, primarily because the relative contribution of CEP-derived vessels was found to be extremely variable in different preclinical models of cancer (Lyden et al., 2001, Ruzinova et al., 2003). Clinical studies in patients who received a gender-mismatched bone marrow

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transplant before cancer recurrence (Peters et al., 2005) indicated that CEP-derived vessels were indeed present, albeit at a low frequency (on average, 5% of all vessels). These apparently conflicting data can be explained by demonstrating that the recruitment of CEPs into tumor vasculature depends on the tumor grade (Ruzinova et al., 2003) and also by showing that CEPs are key contributors in the first steps of tumor vascularization, but – after the establishment of cancer vessels - their quantitative contribution to neoplastic angiogenesis is quantitatively less relevant as these cells become progressively diluted (Nolan et al., 2007). Also, very recent preclinical evidence indicates that the redundancy in cellular-derived mechanisms of neoplastic vascularization is much higher than anticipated, with multiple cellular populations of hematopoietic origin actively participating in addition to CEPs (Madlambayan et al., 2009). In clinical studies, increased CEPs were found in patients affected by lung (Dome et al., 2006), breast (Naik et al., 2008) and brain (Greenfield et al., 2009) cancer, as well as in lymphoma (Igreja et al., 2007), chronic lymphocytic (Gora-Tybor et al. 2009) and acute myeloid (Wierzbowska et al., 2008) leukemia patients. In some of these studies, high CEP levels correlated with a poor prognosis and/or survival (Dome et al., 2006; Wierzbowska et al., 2008; Greenfield et al., 2009). Three previously unrecognized and crucial roles for CEP in cancer have been recently suggested. Angiogenesismediated progression from micro- to lethal macro-metastasis is a leading cause of death in cancer patients. Using preclinical models of pulmonary metastasis, the Mittal lab found that tumors induce Id1 expression in CEPs, and Id1 suprression after metastatic colonization blocks CEP mobilization, cause angiogenesis inhibition, impaired pulmonary macrometastases, and increased survival of tumor-bearing animals (Gao et al., 2008; Shaked et al. (2006) found that treatment of tumor-bearing mice with vascular disrupting agents (VDAs, drugs which cause a rapid shutdown of the tumor vasculature flowed by extensive tumor hypoxia and necrosis), leads to an acute mobilization of CEPs, which home to the viable tumor rim which remains after VDA treatment and drive rebound revascularization and tumor regrowth/recovery after therapy. In another study, Shaked et al. (2008) found that some chemotherapeutics (and taxanes in particular) also induce a rapid CEP mobilization, most likely generation by the modulation of circulating SDF-1 levels. Prevention of this CEP spike by concurrent treatment with antiangiogenic drugs, by genetic manipulation strategies (eg undertaking treatment of tumors in Id mutant mice) or by anti-SDF-1 resulted in enhanced antitumor activity of the administered VDA or chemotherapeu-

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tic drug. These findings rise the possibility that therapeutic strategies aimed at reducing CEP mobilization might enhance the efficacy of anti-cancer therapies and – at the same time – reduces the risk of cancer metastases. CEC and CEP as tools to monitor minimal residual disease A number of investigators have suggested that vessel-lining ECs in tumors might express a specific antigenic profile. The St Croix lab compared gene expression patterns of ECs obtained from normal resting tissues, tumors, and regenerating liver (Seaman et al., 2007). They identified 25 transcripts overexpressed in tumor versus normal ECs, including 13 that were not found in the angiogenic endothelium of regenerating liver. Those EC tumor-specific antigens were primarily cell surface molecules of uncertain function. Should the tumor– specific expression of these antigens be confirmed in CECs, it would be possible – following an intuition of the late Judah Folkman (Folkman et al., 2004) - to monitor tumor-specific CECs in subjects at high risk for cancer development or recurrence after therapy. In contrast to cancer cells, tumorlining ECs have long been considered for many years a genetically stable host cell drug target (Kerbel, 1991). Preclinical and clinical evidence has recently challenged this dogma, and in hematological malignancies such as non-Hodgkin’s lymphoma, myeloma, chronic and acute leukemias and myelodysplastic syndromes there is evidence that ECs might share the same genetic abnormalities found in cancer cells (Gunsilius, 2003; Della Porta et al., 2008). Although it is still not clear whether these unexpected findings were due to a common cancer/EC progenitor, to cancer-to-EC trans-differentiation or to fusion between cancer and ECs (Bertolini et al., 2006), there are at least two related questions that deserve investigation. First, are oncogene-bearing CEC/CEPs potentially able to cause cancer recurrence, and, if so, should they be investigated as a new site for minimal residual disease after therapy? The second question stems from the recent findings of Shen et al (2008). These authors have found that that in some preclinical models of cancer, most blood vessels were derived from precancerous cancer stem cells (pCSCs). These pCSCs expressed VEGFR-2, and were much more potent in tumor vasculogenesis than the differentiated tumor monocytic cells from the same tumor. They also observed tumor-cell related vasculogenesis in human cervical cancer and breast cancers. These findings suggest that oncogene-bearing CEC/ CEPs might be one of the possible hidden identities of cancer stem cells, and suggest a possible explanation for resistance to anti-angiogenic drug therapy of cancer.

CEC/CEP in the clinic. It seems now possible to anticipate two separate fields of clinical investigation for CEPs and CECs. Some CEPs (along with other pro-angiogenic cells of hematopoietic and/or mesenchymal origin) seem to have a “catalytic” role (Seande et al., 2008) in promoting angiogenesis during tumor growth, in stimulating growth of micro- and macro-metastases and in rebound revascularization after certain therapies (Rafii et al., 2002; Nolan et al., 2007; Gao et al., 2008; Lyden et al., 2001; Ciarrocchi et al., 2007; Shaked et al, 2006; Kaplan et al., 2005). These cells are potentially promising targets for anticancer therapies and for adjuvant therapeutic strategies in patients at risk for cancer relapse. Also, one can try to exploit the tumor tropism of these cells for delivering anti-cancer drugs specifically at tumor site. Notably, these catalytic functions are in most cases associated with a pulsating presence of these cells in the blood and in the tissues, and thus it might be difficult to exploit measurement of these cells as biomarkers for selection and stratification of patients. CECs in most cases are apoptotic or necrotic cells, being released in circulation as a byproduct of vascular turnover, so they do not represent a “druggable” target for anticancer therapies. On the other hand, CEC presence in the blood seems less pulsating (ie more stable) and there is increasing evidence of their potential as surrogate biomarkers of cancer angiogenesis and of anti-angiogenic drug activity (Bertolini et al., 2006; Farace et al., 2007). Conclusions In the future, a great effort should be devoted to reach a consensus in the identification and enumeration of CECs and CEPs. Multi parametric FC seems to be a good tool to reach this goal, in spite of the lack of a single specific antigen/ marker able to identify these cells and of the rarity of them. Clinical data suggest that the identification and enumeration of CECs and CEPs might be useful to select the most appropriate therapy for patients who are candidates to antiangiogenic treatments, either as baseline or as dynamic markers. Furthermore, CECs and CEPs might be studied as escape markers in order to prevent the switch towards a disease refractory to anti-angiogenic therapies, alone or in association with chemotherapy (Calleri et al., 2009; Jain et al., 2009).

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Fig. 1: Rendering of the different cellular events with an endothelial phenotype enumerated by ﬂow cytometry, including CD45,CD31+CD146+ macroparticles from apoptotic or necrotic CEC fragments, DNA-containing CD45-CD31+CD146+ viable or apoptotic CECs, and lymphoid-like CEPs expressing CD133 and/or CD34 progenitor markers (from Mancuso and Bertolini, Microvascular Res 2010, modiﬁed).

Fig.2: Mechanisms for endothelial cell detachment and microparticle formation following vascular damage. A vascular insult, such as inﬂammation, might induce endothelial cell detachment from the intima and shedding of endothelial microparticle (EMP) from the activated endothelial cells. Ciculating CEC, endothelial cells (CEC) undergo apoptosis by anoikis (From Martin-Padura and Bertolini, Front Biosci 2009, modiﬁed)

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Fig.3: Role of CECs and CEPs in cancer. A. Circulating endothelial cells (CECs), with a mature phenotype, are increased during tumor progression. Endothelial progenitor cells (CEPs) home from the bone marrow to incorporate to the tumor neovessels. Other bone marrow-derived cells might participate in the process of tumor angiogenesis (see text), such as VEGFR1+-hematopoietic progenitor cells that, toghether with the CEPs, might initiate the pre-metastatic niche. B. Followig antiangiogenic treatment there is an increased of apoptotic CECs detaching from the tumor vessels. CEP mobilization is increased by high-dose chemotherapy and reduced by metronomic chemotherapy and angiogenic treatments (From Martin-Padura and Bertolini, Front Biosci 2009, modiﬁed).

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Circulating Tumour Cells

The objective of this group led by Franco Nolè and Maria Teresa Sandri is to understand the role of Circulating Tumor Cells (CTCs) in patients with different malignancies. The detection of circulating tumour cells (CTCs) in peripheral blood has been of interest for over a century. The existence of these cells fits nicely with the model of haematogenous spread in the development of metastatic disease and has the promise of providing a better understanding of the biology of metastasis. There are accumulating reports on the isolation and characterization of CTCs. The findings that CTCs can be found in patients before the primary tumor is detected, in a significant proportion of patients when a carcinoma recurs, and that CTCs persist in some patients after removal of the primary tumor, have been the impetus for continued studies of these tumor cells. Evidence that CTCs are derived from clones in the primary tumor suggests that they may reflect the tumor burden at all stages of tumor progression. Thus, in addition to a potential role in early diagnosis and prognostication, CTCs may play a major role in characterizing genetic and immunophenotypic changes with tumor progression, thereby helping to guide targeted therapy. A major problem in the field is that different techniques have been used to isolate and characterize CTCs and that different studies report conclusions sometimes difficult to compare. Recently a new technology has been developed with the purpose of detecting CTCs in blood with a standardized and reproducible approach. The system (CellSearch, Veridex LLC, Warren, NJ) involves a technique of mixing a blood sample with iron particles coated with an antibody that attaches to epithelial cells. The epithelial cells are then distinguished from leukocytes by antibodies that have been tagged with a fluorescent dye so that the cancer cells can be easily distinguished and counted. Since epithelial cells are not usually found in the blood, these cells are likely cancerous cells from the tumor. This strategy represents a step forward compared to the previous methods, and it could provide useful predictive and prognostic information with a simple, safe and repeatable blood drawing, avoiding invasive procedures such as bone marrow aspiration, which may provide limited patient compliance. Breast Cancer There are many potential clinical applications for CTCs in breast

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cancer including screening, predicting which patients with early stage disease will recur despite adjuvant therapy, monitoring for recurrence after adjuvant therapy, estimating prognosis in metastatic disease, predicting which drug is most likely to be efficacious for metastatic disease, and monitoring therapy for metastatic disease. However, good data only exist for establishing prognosis in metastatic breast cancer, and for monitoring therapy in metastatic breast cancer. A study by Cristofanilli et al, using a standardized method based on immunomagnetic enrichment and selection of circulating epithelial cells and on the use of multiple markers for positive identification of CTCs, demonstrated that CTCs are highly prognostic in 177 patients with metastatic breast cancer (MBC) who were beginning a new therapy (Cristofanilli M et al. N Engl J Med 2004). Multivariate analysis of clinical factors demonstrated that while clinical factors (including time to metastasis, HER2/neu status and type of therapy) still correlated to outcomes; the strongest predictors of progression-free and overall survival were the levels of CTCs at baseline and at the first follow-up visit. This study suggests that levels of CTCs may be predictive of outcome in patients with malignant breast cancer. These results have been confirmed in a prospective mono-institutional study, conducting in our Institution (F. Nolè … M.T. Sandri et al, Ann Oncol 2008), evaluating the prognostic significance of CTCs detection in advanced breast cancer patients. The study included 80 patients with histological diagnosis of breast cancer, evidence of metastatic disease from imaging studies, starting a new line of therapy and/or treated for the advanced disease with a maximum two lines of therapy. The CellSearch system was used to test for circulating tumor cell levels before starting a new treatment and after four, eight weeks and at the first clinical evaluation and every two months thereafter. At baseline, 49 patients were found to have ≥ 5 CTCs. The baseline number of CTCs were associated with progression-free survival (hazard ratio [HR] 2.5; 95% confidence interval [CI] 1.2–5.4). The risk of progression for patients with CTCs ≥ 5 at the last available blood draw was five times the risk of patients with 0–4 CTCs at the same time point (HR 5.3; 95% CI 2.8–10.4). At the last available blood draw, patients with rising or persistent CTCs ≥ 5 demonstrated a statistically significant higher risk of progression with respect to patients with CTCs < 5 at both blood draws (HR 6.4; 95%

CI 2.8–14.6). These results indicate that elevated CTCs levels measured at any time in the clinical course of a patient with metastatic breast cancer predict an imminent progression and that this analysis represents an additional step in the process of validating this method. In addition to enumeration, an exciting area of CTC research involves the phenotyping and expression profiling of CTCs. In this regard, the evaluation of CTCs could be considered as a real-time biopsy allowing, in most patients with metastatic breast cancer, the possibility to evaluate the possible change in tumor genotype and phenotype during the clinical course of the disease. Several studies demonstrated that the presence of a target antigen can be quantitatively analyzed using epithelial cells isolated from blood. In particular, there are some evidences of the possibility that HER-2 gene amplification can be acquired during progression of cancer demonstrating a phenotypic conversion from HER-2- to HER-2+ during the course of the treatment. Ongoing Projects on Circulating Tumor Cells in Breast Cancer Advanced Breast Cancer: • Phenotypic and functional characterization of Circulating Tumor Cells in patients with Advanced Breast Cancer This study is aimed to evaluate i) the concordance of HER-2/ neu expression between the primary tumor and CTCs, ii) the acquisition of HER-2/neu overexpression on CTCs in patients with HER-2/neu negative, advanced breast cancer, treated with chemotherapy during their clinical course, iii) the efficacy, in terms of response rate, to trastuzumab-based therapy, in patients showing the acquisition of HER-2/neu overexpression, and iv) a possible correlation between the acquisition of HER-2/ neu overexpression on CTCs and clinical outcome. This project is partially supported by AIRC and foresees some translational “tasks” aimed at the evaluation of immunophenotypical detection and functional characterization of cancer stem cells within the isolated CTC population, at the identification of new stem cell genes, by gene microarray analysis and of genes involved in resistance to apoptosis. Between June 2007 and October 2008, a total of 76 patients were enrolled in the trial and followed for CTCs enumeration and HER-2 phenotypic characterization (first and second part of the study). Fifty-nine pts had at least 1 CTC in 7.5ml of blood (prevalence 75%). Median CTCs number was 7/7.5 ml (range: 1- 4988). Concordance of HER2 phenotype between the primary tumor and CTCs in the basal sample was 86% (49/57 pts) and 82% (50/61) during the given treatment. The “acquisition of her-2/neu over-expression on CTCs during chemotherapy”, has been evaluated in the second part of the study. Fifteen pts with primary HER-2 positive tumor had at least 1/7.5 ml CTC in the blood sample and 42 with primary HER2 negative tumor had at least 1/7.5 ml CTC in the blood sample. These pts were followed during the treatment to assess a change in HER-2 phenotype.

Six patient (14%) with HER2 negative primary tumor acquired HER2 overexpression on CTCs prior to start the new chemotherapy regimen. Two pts with HER-2 positive disease in the primary tumor lost HER2 overexpression on CTCs prior to start the new chemotherapy regimen (E. Munzone, F. Nolè, MT Sandri et al, Ann Oncol 2009) Having obtained the approval of Ethics Committee, patients with acquired overexpression of HER-2/neu on CTCs will be treated with Trastuzumab. This part of the study will be conducted in 2010. • Evaluation of Reverting estrogen-receptor-negative phenotype in HER-2-overexpressing advanced breast cancer patients exposed to trastuzumab plus chemotherapy Biological features of endocrine responsiveness (expression of ER and/or PR) are important predictive and prognostic markers in breast cancers. About 40% of patients with ER-positive primary tumors that relapse after adjuvant endocrine therapy have recurrent tumors in which ER expression is lost. Progression to an ER-negative phenotype typically involves the constitutive over-expression of growth promoting genes that are normally regulated by estrogens, thereby leading to a loss of estrogen dependence, resistance to anti-estrogens, and a more aggressive phenotype overall. The re-expression of ER, therefore, in tumors in which ER has been lost or is not expressed could allow for endocrine therapy sensitivity. ER-negative tumors frequently over-express growth factor receptors such as EGFR or c-erbB2, as do many ER-negative breast cancer cell lines, suggesting that up-regulated growth factor signalling may provide an alternative growth stimulus. EGFR and c-erbB2 over-expression are also important prognostic indicators in breast cancer, independent of their inverse correlation with ER expression. This increased growth factor receptor expression and/or activation correlates with increased MAPK activity, both in tumors and in cell lines. Using cell lines that over-express constitutively active forms of Raf-1, MEK1, or c-erbB-2 and ligand-activatable EGFR, some authors have shown that the resultant hyperactivation of MAPK (ERK1/2) activity through these signaling pathways leads to the down-regulation of ER. This down-regulation is not a consequence of ligand-independent ER activation, and it is reversible in vitro. Abrogation of ERK activity using either pharmacologic inhibitors or dominant-negative ERK (dnERK) constructs reverses the ER down-regulation and restores ER activity. These data suggest that up-regulated growth factor signaling via MAPK is directly linked to loss of ER expression and generation of the ER-negative phenotype and that, at some stage in the progression pathway, the ER-negative phenotype may be not permanent. As a consequence, therapeutic targets enabling the restoration of ER expression may provide beneﬁcial treatment strategies for breast cancer patients. Both geﬁtinib (Iressa®), which inhibits the kinase activity of EGFR and trastuzumab (HerceptinTM), a monoclonal antibody against c-erbB2, have been shown to reduce MAPK activity. These data

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supported the hypothesis that advanced breast cancer patient with ER-negative tumors and c-erbB2 overexpressed could revert the ER-negative phenotype into positive after treatment with trastuzumab. On these bases, we planned a study aimed to to evaluate the ability of Trastuzumab containing therapy to revert ER-negative phenotype to ER positive in HER-2 overexpressing tumors. The study will be conducted in 2010. Operable Breast Cancer: Pilot Study This Project evaluates the role of CTC enumeration in patients with operable breast cancer and no known metastases. The entire project is divided in three parts: the first part is aimed at the evaluation of the prevalence and of the kinetics of CTCs in patients with operable breast cancer before and after any surgical procedure. The second part analyzes the presence of possible association between CTCs detection and clinicopathological characteristic, and the third part of the study will evaluate the “prognostic impact” of the presence of CTC before or after surgery, in terms of clinical recurrences, disease free-interval and overall survival. The presence of CTCs was evaluated in 56 breast cancer patients before surgery and again 5 days after surgery. In case of positivity of one of the perioperative samples, another sample was taken after 30 days. The presence of CTCs was assessed with the CellSearch System (Veridex, Warren, NJ). One to three CTCs were found in 16 (29%) of 56 patients before surgery, in 14 (30%) of 47 patients at day 5, and in 8 (30%) of 27 at day 30. No association with pathological characteristics was found, apart a borderline significant association between presence of CTCs at baseline and vascular invasion (P = 0.07). When we looked at concordance between CTCs at baseline and after day 5 (47 patients), we found 40% discordant samples (10 negative at baseline and positive at day 5, and 9 vice versa). This study provides evidence of the presence of CTCs in approximately 30% of patients with localized breast cancer both before and after surgery, with change from positive to negative and vice versa in 40% of cases. No association with the pathological variables was found, except for vascular invasion and presence of preoperative CTCs. Longterm follow-up will be required to understand the significance of these data. (Sandri MT, Zorzino L, Nolè F. et al, Ann Surg

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Oncol February 2010) Characterization of Circulating Tumor Cells Apart from counting CTC, in 2009 we started some projects aimed at the characterization of CTCs. In particular we focused on the possible detection among CTCs population, of cells presenting stemness feature, as identiﬁcation of cancer stem cells has fundamental implications for cancer biology in addition to clinical implications for cancer risk assessment, early detection, prognostication, and prevention. It is gaining more and more evidence the fact that cancer drugs based on tumor regression may have produced agents which kill differentiated tumor cells while sparing the small cancer stem cell population. According to the cancer stem cell model, tumors originate in either tissue stem cells or progenitor cells, through deregulation of the normally tightly regulated process of self-renewal. Self-renewal is the process by which stem cells generate progeny identical to themselves. Stem cells also differentiate to generate multipotent progenitors, which in turn give rise to committed progenitors and differentiated cells. Cancer stem cells share these properties with their normal counterparts: they have self-renewal capacity, driving tumorigenicity, recurrence and metastasis and they have the capacity to differentiate, albeit aberrantly, giving rise to a heterogeneous population of cancer cells. The differentiated cells constitute the bulk of the tumor, but they are not tumorigenic, due to their lack of self renewal capacity and limited proliferation potential. It has been previously demonstrated that that human breast cancers contain a cell population with stem cell properties, bearing the surface markers CD44+/ CD24-/lin-. Since then different markers have been investigate for their ability to identify the cancer stem cell: Among these, the aldehyde dehydrogenase 1 (ALDH1), a detoxifying enzyme responsible for the oxidation of intracellular aldehydes seems of particular interest. In vitro and in vivo experimental systems showed that normal and cancer human mammary epithelial cells with increased ALDH1 have stem/progenitor properties, and that in breast carcinomas high ALDH activity identiﬁes the tumorigenic cell fraction, capable of self-renewal and of generating tumors which recapitulate the heterogeneity of the parental tumor. Therefore it seems of utmost importance on one hand

to have drugs targeting this key population and on the other to have reliable methods to detect cancer stem cells. Tightly linked with stem cell properties, the epithelial to mesenchymal transition (EMT) seems to represent a very important step in disease progression: Several distinct traits have been conveyed by EMT, including cell motility, invasiveness, resistance to apoptosis, and some properties of stem cells. Many signal pathways are involved in the induction of EMT, such as transforming growth factor- , Wnt, Hedgehog, Notch, and nuclear factor- B. Finally, over the last few years, increasing evidence has shown that EMT plays an essential role in tumor progression and metastasis. Different markers have been proposed to speciﬁcally detect these cells. Among them Twist, Akt2 and PIk3a can be tested altogether. Taken these data altogether, we began to settle a system allowing the detection of the expression of these markers (ALDK1, Twist, Akt2 and PIk3a) on CTCs, as they may reveal to be better indicators of disease status than current imaging methods. This system is based on enrichment of CTC from 5ml blood followed by reverse transcription cDNA synthesis followed by 2 multiplex PCR assays, one to analyse Twist, PI3K , Akt2 and the second to analyse ALDH1. In 2010 we will test these assays in patients with locally advanced breast cancer undergoing neoadjuvant chemotherapy. Other malignancies Ongoing projects on Circulating Tumor Cells in Prostate Cancer We are focusing our attention in different clinical settings: 1.Patients with castration resistant tumor, with biochemical failure with no known metastases and patients with metastatic disease, non-responsive to hormonal ablation. In these patients the prevalence of CTCs will be assessed and both a possible predictive value of response to treatment and a prognostic impact of their baseline level on clinical outcome will be evaluated. Moreover the impact of quantitative changes in CTCs during chemotherapy will be studied. To date 45 of planned 54 pts have been enrolled. Preliminary results showed 70% prevalence of intact CTCs >5/7.5 mL. We found a signiﬁ-

cant correlation between CTCs >5 and a number of previous treatment >1 (p-value0.081). At the basal determination, CTCs show a positive correlation with PSA levels (p-value=0.022), alkaline phosphatase determination (p-value <0.001) and a negative correlation with Hb (p-value 0.087). A value of CTCs >20 after 6 weeks from the beginning of the therapy is correlated with a signiﬁcant decrease in progression free survival (p-value 0.037). CTCs seems to conﬁrm their prognostic and predictive value. Patients accrual will be completed in 2010 2. Patients with localized prostate cancer. During 2010 a project in patients with clinically localized prostate cancer, eligible for radical prostatectomy will be activated. In this population, we will evaluate the prognostic impact of the presence of CTC before any therapy in terms of correlation between presence of CTC, pathological stage, baseline PSA levels, Gleason score and Tumor Volume. Ongoing projects on Circulating Tumor Cells in colon cancer Some reports suggest a potential prognostic role for CTCs in patients affected by advanced colon cancer. In 2008 we started a study aimed at the evaluation of CTCs in patients undergoing curative surgery for advanced rectal cancer. These patients underwent CTCs determination before and after chemotherapy and surgery, and the association with Disease Free Survival and Overall Survival will be evaluated. Forty-one patients were enrolled; nowadays 26 were evaluable for biological parameters: among them (16M and 10F) underwent baseline sampling (t0), 8 pts completed CT/RT and therefore underwent subsequent two-evaluations pre-surgery (t1) and 7-days after surgery (t2). At t0 point, three pts presented 1 CTC (12%), one 2 CTCs (3.5%), one 27 CTCs (3.5%) while in twenty-one (81%) no CTCs were detected. At t1 and t2 none of the eight pts analyzed showed CTCs. CTCs ≥ 1 are present in 15% of our patients, but the sample is too small for clinical and/ or statistical analysis. The study recruitment is still open for achieving sample size objective.

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Virology

The objective of the group led by Maria Teresa Sandri and Mario Sideri is to understand, cure and prevent the process of virus-induced tumor development, and particularly to study the role of Human Papilloma Virus (HPV) in the development of different tumors like those of the head and neck, anus, vulva, and cervix. Infections with HPV are extremely common, but rarely cause serious primary disease in healthy individuals. Nevertheless, these viruses may persist after primary infection and cause tumour disease later. A cause-effect relationship exist between certain types of HPV, e.i. the oncogenic or “high risk HPV” genotypes, and cervical cancer, and the same types have now been associated with other malignancies, such as head and neck, anus and penis cancer. Nonetheless, a persistent infection is necessary but not sufﬁcient to lead to neoplasia development. Vaccines: a new strategy for prevention and cure? We still lack precise knowledge regarding the pathology of HPV. Why do these viruses sometimes persist? And why does virus persistence lead to tumour development and disease? It is obvious that the immune system plays a crucial role in the interplay between viruses and hosts, since tumour development and disease are more frequently observed in immunosuppressed individuals, like women infected with HIV who are more frequently affected by HPV-related diseases. Many studies have been performed to develop preventive and curative vaccines. In 2007 two preventive vaccines were introduced and are now available: they are both directed at the eradication of infection with HPV 16 and 18, which are known to cause about 70% of cervical cancer. Our group is involved in a study looking at the response to the vaccination of 18 years old adolescents living in the Milano and Monza area who are vaccinated with GARDASIL and followed for a ﬁve-year period. The primary endpoint is the incidence of viral infection, looking at the new epidemiology and prevalence of viral HPV genotypes in vaccinated girls. The project started in spring 2008 and almost 400 girls have been vaccinated so far. Cervix cancer screening and HPV The primary research of our group is focused on the role of

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HPV in cervix cancer. Pap smear screening has revolutionized cervix cancer screening since its introduction into the population. In Italy there is both organized and opportunistic screening: thanks to this approach the prevalence of cervix cancer has dramatically decreased and nowadays about 3500 new cases per year are diagnosed. In recent reports, however, there were observations suggesting an increase in cervix cancer incidence, especially of the adenocarcinoma of the cervix. Apart from those cases observed in women who do not regularly perform a screening test, the Pap test’s insensitivity and lack of specificity have been the main reasons for the remaining cases of cervix cancer despite the introduction of the screening. It is well known that cervical cancer is preceded by a long lag time of 15 to 20 years from early premalignant changes to invasive cancer. The etiologic role of human papillomavirus (HPV) in cervical cancer has been well established by scientific evidence, and more than 99% of these cancers can be attributed to HPV. (zur Hausen, Cancer Res 1989). Although not all women who are infected will develop cervical cancer because other co-factors play a fundamental role, a reasonable approach is represented by the direct search of the presence of the oncogenic HPV types in cervical samples, both in the screening population and the patients treated for a preneoplastic lesion. In particular in this last group of subjects, the persistence of HPV after the treatment during the follow-up would allow the selection of a group of patients bearing a higher chance of relapse and hence needing more intensive controls. On the contrary, the absence of HPV would identify those patients effectively cured by the conservative treatment, in whom intensive follow-up is not needed. The introduction of HPV testing and genotyping in screening requires information on the genotype distribution in positive pap smears and cervical pre-cancerous and cancerous lesions. Our group investigated this aspect in detail and a first paper was published (Sandri et al, J Med Virol. 2009); in a second paper submitted a different age distribution was observed between lesion associated with HPV 16-18 and lesions associated with other high risk HPV genotypes; a third paper, the largest Italian series published so far, investigated the distribution of HPV genotypes in invasive cervical cancers in Italy (Sideri et l, Vaccine 2009).

New and old diagnostic biomarkers New tests are now being proposed for the detection of the presence of the DNA of HPV in cervical samples. We think that it is very important to evaluate both the analytical and the clinical performance of the newly introduced test, and one of our aim is to present to the scientific community the advantage and disadvantage of the different methods. The results of the comparison of the two methods available have been the topic of a paper recently published (Sandri MT et al, J Clin Microbiol 2006). Apart from the presence of the infection with HPV, we are looking for other diagnostic tool which may help in a better characterization of the lesion, such as genotyping, specific mRNA for E6 and E7, viral load. Following the first published studies in which the relevance of bearing HPV 16 or 18 compared to the other oncogenic genotypes leads to a different risk of developing a pre-neoplastic lesion, we are studying the distribution of the different genotypes in patients with cervical lesions. In fact as it is viral persistence that is considered to be the true pre-cursor of neoplastic progression, DNA genotyping approaches designed to identify type specific persistence, could be more appropriate for identification of those individuals at greater risk of disease progression compared with a presence/absence test. Moreover the prevalence of the different oncogenic HPV in patients with cervical lesion is still not well known in the Italian population and we are studying the genotype distribution. A key information is related to the performance of the different genotyping systems: we performed a study in collaboration with a group of Marseille showing that two systems commercially available show similar performance (Halfon P, Sandri MT, JMV 2009). . A promising marker of progressing lesions is the determination of p16 overexpression. P16 identification by immunostaining can beconsidered as a marker, not only of HPV infection, butalso of activated expression of viral oncogenes and ofvirus-induced deregulation of the cell cycle. A large multi-institutional study was concluded in 2009 evaluating double staining on liquid based cytology specimens, in screening and triage of borderline lesions on 27.000 women. One of our objective is to better characterize women with HPV infection in order to build an algorithm allowing a tailored approach to the different situations. The use of biomarkers gives better positive predictive value as compared to HPV DNA testing alone. With improved diagnostic methods and triage in the detection of precursors and providing efficient treatment, the incidence of cervical cancer can further be reduced from its current incidence of 10/100000 women. Self collection for HPV testing Although women are encouraged by governments and health authorities to undertake regular Pap smears, the procedure involved in collecting a sample requires the intervention of a medical practitioner. It causes discomfort, is embarrassing, inconvenient, and conflicts with the personal beliefs of some

women. Even attending a doctor to discuss sexual matters can be confronting. Non participation in screening programmes is consequently higher among certain ethnic and religious subsections of the community, as well as women of poorer socioeconomic status and other women deemed at risk. Apart from this, the Pap smear result can be a false negative if the affected region of the cervix is missed (i.e., inadequate sampling), or the cytological interpretation is incorrect. If a woman is infected with HPV, the viral DNA will be present in cells that are shed from the surface epithelia of the cervix and vagina. Thus testing for HPV in such exfoliated cells is an alternative way of screening, as shown in a number of studies, with good results. In early work collection of exfoliated cells involved a cervico-vaginal lavage. A lavage can be self-administered, but its value is limited due to patient compliance. Subsequent studies found that tampon samples also contained an adequate amount of exfoliated cells for HPV detection. Such sampling can be carried out easily by the woman herself.. This can facilitate expanded screening of women and identify infected women without gynecological intervention. It would enable participation by women from geographic localities lacking gynecological services, as well as those who would otherwise be unable or unwilling to attend for a Pap smear. Even women who do attend for their regular Pap smear would likely prefer to take their own specimen, rather than visit their doctor. Self-collection increases uptake and could make up for loss of diagnostic accuracy, if any. Results from tampons correlate well with results from cervical scrapes. A recent meta-analysis concluded that self-collection may be an appropriate alternative for low resource settings or patients reluctant to undergo pelvic examination. Scrapes and biopsies allow detection of HPV in discrete lesions or sites sampled, so do not yield information representative of the entire cervical epithelium. Scrapes are signiﬁcantly less sensitive than cervicovaginal lavage specimens. Collecting a scrape requires, moreover, a trained health professional, whereas self-collection does not. Urine self-collected as a dry paper smear for ease of transport, storage and direct HPV PCR testing, gave results that matched those from a cervical scrape, and is an alternate potential sampling approach. A study has been now completed on more than 700 women attending opportunistic screening at IEO showing the accuracy of self sampling in the identiﬁcation of women with high grade cancer precursor lesions. As an extension of the concept of self-collection, a protocol has been suggested whereby a woman can purchase a tampon-based kit from a pharmacy or other suitable retail outlet, collect a specimen at her own convenience in private then either mail this to the clinical pathology laboratory or place it into a suitable receptacle, such as a collection box at the pharmacy or supermarket, the samples then being collected by a courier and transported to the laboratory that then does the HPV testing. The result would then be sent to her and/or a doctor she nominates on a form that accompanies the sampling kit.

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Breast Tumours

Ideas for clinical research in breast cancer derive mainly from the Task Force for breast cancer, the first multidisciplinary group set up at the IEO. The Main areas of clinical study are: 1) Sentinel Node Biopsy. Sentinel lymph node biopsy (SLNB) is to date a standard procedure for axillary staging in patients with early breast cancer. The fields of application of SLNB have been now enlarged and this procedure can be offered also in clinical scenarios in which it was previously considered contraindicated such as during pregnancy, in multicentric breast cancer, after neoadjuvant chemotherapy or previous breast surgery. A multicentric trial carried out within the International Breast Cancer Study Group (IBCSG) completed the accrual after randomizing 933 patients. The aim of this prospective randomized trial was to determine whether axillary dissection can be avoided when the sentinel node contains only micrometastasis. 2) Intraoperative Radiotherapy with Electrons (ELIOT). The rationale for the use of partial breast radiation therapy in place of whole-breast irradiation is based on the finding that approximately 85% of local breast relapses occur in the same quadrant of the primary tumor. Residual microscopic disease are usually located in close proximity to the primary tumor and is often the cause of local disease recurrence that in most cases occur in the same quadrant as the resection. Intraoperative Radiotherapy with Electrons (ELIOT) clinical trial started in 2000 randomizing patients to receive either 21 Gy intraoperatively in a single fraction or external beam radiotherapy with conventional fractionation. The trial enrollment has been completed in December 2007. Inclusion criteria were the following: age > 48 and < 75 years, tumor diameter < 2.5 cm, infiltrating histology, no mammographic evidence of multifocality. Of the 1306 patients that entered the trial 1187 were confirmed eligible. 655 in the external beam radiotherapy arm and 651 in the ELIOT arm. We are carefully monitoring all these patients in order to determine

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whether ELIOT can substitute traditional radiotherapy in women undergoing conservative treatment for early breast cancer. 3) ELIOT boost plus whole breast radiotherapy. It has been shown that a high dose of radiation to the tumor bed particularly in younger patients reduce local recurrence. This evidence coupled with the reported higher relapse rate in premenopausal women led to the design of a new program for premenopausal patients which started in June 2004. An anticipated electron intraoperative radiotherapy boost of 12 Gy is followed by a hypofractionated external beam radiotherapy course of 37.05 Gy delivered in 13 daily fractions of 2.85 Gy. More than 400 patients have been treated with this scheme, acute toxicity has been acceptable and comparable with external beam radiotherapy with conventional fractionation. Patient’s compliance is high. This treatment approach shortening the standard course of whole breast RT to less than 3 weeks avoid controversies regarding the sequence of local and systemic treatments. The procedure allow to deliver a high dose to the tumor bed and an adequate dose to the whole breast in a short overall treatment time with a potential gain in the radiobiological effect. Our institution is therefore currently treating premenopausal breast cancer patient with this innovative and very promising approach. 4) Breast Conservation for ispilateral breast tumor reappearance. Seeking the most effective treatment of early breast cancer with a conservative approach is perused also studying the feasibility of a second conservative treatment in women with local disease recurrence. With the increasing number of patients undergoing breast conserving surgery followed by radiotherapy (BCT) the treatment of ipsilateral breast recurrence (IBTR) is a challenging issue. In various series the 10-years local recurrence rate is reported in the range of 10-20%. Salvage mastectomy has been traditionally considered the standard local therapy after IBTR. Nonetheless patients

are frequently requesting a second chance to preserve the breast. Few experiences reported on further conservative surgery with or without irradiation mostly using brachytherapy. We are studying the feasibility of partial breast reirradiation by using intraoperative radiotherapy (ELIOT). Starting in 2000, we enrolled 37 patients with small recurrent breast carcinoma after initial BCT and RT, who underwent a further quadrantectomy followed by partial breast irradiation using ELIOT. 12 of the 37 were treated in 2007. The median interval from initial diagnosis and IBTR was 122 months (range 23-312); the histology was invasive in all cases. In 12 of the 37 patients, tumor recurred in the same region of the breast. All patients underwent a second excision of the tumor and were treated with ELIOT to the tumor bed and the surrounding tissue with a margin according to tumor presentation. Different applicators were used with diameters ranging from 3 to 6 cm and different energies from 4 to 10 MV according to the thickness of the gland. The dose delivered varies from 16 Gy to 21 Gy. The ﬁrst 12 patients were treated with lower doses ranging from 8 to 15 Gy. At a median follow up of 20 months (range11-63), 4 patients developed a second local relapse (10.8%) and in 2 cases a contralateral breast cancer (5.4%) was registered. Late side effects were available for 34 patients who completed at least 6 months of follow up. Fibrosis in the site of recurrence and reirradiation of grade 0/1 was reported in the majority of cases (21/37), while a grade 3 in 4 patients. The evaluation of cosmesis showed a poor result in 2 patients, fair in 13 and good/excellent results in the remaining cases. These preliminary experiences are encouraging. Reirradiation with ELIOT showed no signiﬁcant side effects, acceptable cosmesis and high level of satisfaction of patients and may be an acceptable alternative to the mastectomy in selected cases. 5) SNOLL. This is a technique that combines radioguided occult lesion localization (ROLL) with radio-guided sentinel node biopsy (SNB). It is used in patients with non-palpable lesions shown to be malignant by pre-operative biopsy or intraoperative histological analysis. The radio-tracer for ROLL and SNB are injected separately, but during surgery a gamma-probe is used to guide removal of both the occult lesion and the sentinel node. The advantage of SNOLL is that it allows complete treatment of non-palpable lesions (tumor removal, SNB and axillary dissection) in a single surgical session. 7) Intraoperative Avidination for Radionuclide Therapy (IART). This technique is being developed in conjunction with Nuclear Medicine. Avidin is injected intraoperatively

in and around the tumour bed after cancer removal (quadrantectomy) and the wound is closed as normal. Later, radiolabelled biotin is injected intravenously. The biotin binds to the avidin in the tumour bed, bringing with it radionuclide whose decay kills residual tumour cells. Preliminary studies indicate fast and stable uptake of radiolabelled biotin at the operated site, and the radiation dose released to the breast is over 5 Gy/GBq. These encouraging results have stimulated a feasibility study on IART. Medical Oncology Medical Oncology research is equally committed to scientific discovery and patient care. Our world-class clinical investigators work across disciplines, departments, and institutional boundaries to translate research findings into new diagnostics and therapeutics for patients. The cornerstone of translational research of medical oncology staff is collaboration: close interactions among basic scientists, computational biologists, chemists, clinical investigators, and others. The group also enjoys fruitful partnerships with pharmaceutical and biotechnology companies, which have the complementary resources needed to help transform promising compounds into drugs and biologics. A major departmental research theme is linking knowledge of the genes that cause cancer to the discovery and testing of new therapeutics, involving both small-molecule drugs and immune approaches (vaccines). Other key themes relate to developing personalized medicine strategies by using genetic, epidemiologic, and population-based studies to determine risk and ideal treatment for individual patients. The medical oncology team currently has nearly 50 open adult therapeutic clinical trials. It accrues several hundreds patients to therapeutic and non-therapeutic clinical protocols each year. Disease center members play a major role in the IEO research programs and in international cooperative group trials, such as the International Breast Cancer Study Group (IBCSG), the Breast Internation Group (BIG) and the Ludwig Institute for Cancer Research. Department investigators focus on testing new drugs in Phase I and II trials, particularly first-in-human studies that have the potential to move the boundaries of breast oncology care. Technologies being offered include the isolation, enumeration, and genotyping of circulating tumor cells, determination of plasma cytokine levels, and genotypic analysis of plasma-based tumor DNA. All these technologies are applied in clinical trials

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The milestones of our clinical research are here summarized: 1. Identification of biological features of disease in early breast cancer predictive of response to a target-oriented approach into the neoadjuvant and adjuvant setting. 2. Identification of mechanisms of resistance to trastuzumab in HER2 positive breast cancer disease and development of new strategies to target HER2 positive breast cancer.. 3. Role of aromatase inhibitors (AIs) and switching to AIs in the adjuvant setting for the treatment of early breast cancer. 4.Exploring the combination of endocrine therapy with biological agents targeting HER2, src or insulin growth factor receptor (IGFR) 5. Identification of prognostic and predictive molecular markers in metastatic setting of disease (circulating tumor cells) 6.Identification of new strategies (metronomic chemotherapy or combination of metronomics with biological agents) in the treatment of metastatic breast cancer. 7. Identification of surrogate markers predictive of response to antiangiogenic therapies with molecular profiling of patients included in trials with antivascular agents. 8. Identification and development of individualized treatments of specific breast cancer clinical presentations (pregnancy, very young patients, elderly patients). 9.Development of new drugs in the treatment of metastatic breast cancer 10.Selecting cancer vaccine targets for individual cancers. Analyzing the immunogenicity of T-cell and B-cell peptide epitopes and performing cytokine immune assessments to identify epitopes and cytokines that enhance immune responses 11.Identification of mechanisms of resistance to standard therapies by breast cancer stem cells exposure tests. Scientific production of the multidisciplinary team included several high impact peer reviewed manuscript. Most of them had a major clinical impact, changing practice in the neoadjuvant treatment of patients with breast cancer (Colleoni M et al. Breast Cancer Res Treat. 2009); in the adjuvant treatment of menopausal patients with endocrine responsive disease (BIG 1-98 Collaborative Group. N Engl J Med. 2009) and in the adjuvant treatment of patients with small tumors and HER2 positive breast cancer (Curigliano G et. J Clin Oncol. 2009). Other paper were focused specifically on the impact of oral chemotherapy with a low toxicity profile in the treatment of patients with metastatic breast cancer (Nolè F. et al. Cancer Chemother Pharmacol. 2009).

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“In the next 20 or 30 years, we will cure or significantly reduce the burden of several subtypes of breast cancer,” Aron Goldhirsch, Breast Cancer Program Director, sums up. “We’ll be able to identify the subtypes and treat them selectively, improving outcomes, and reducing toxic effects. It won’t be easy, but it will be an exciting time in clinical research.” Intraoperative Avidination for Radionuclide Therapy : IART Intraoperative Avidination for Radionuclide Therapy is a new approach of targeted radionuclide therapy of early breast cancer invented at IEO, which involves two steps: a)inoculation of avidin into residual glandular tissue immediately after primary surgery; b)intravenous injection of 90Y-DOTA-biotin the day after surgery. The 90Y-DOTA-biotin homes in specifically to the avidin in the breast tissue and provide a therapeutic dose of radiation to the breast. Our studies indicated fast and stable uptake of radiolabelled biotin at the operated site. Data from phase I-II study on 35 patients - who received IART as a boost to accelerated external beam radiotherapy - revealed that the procedure is safe since no side effects of avidin or radiolabelled biotin occurred. No haematological toxicity was observed. Local toxicity during EBRT was acceptable, with good cosmetic outcomes and quality of life. IART radiation dose to the heart from the irradiated breast area was negligible, also when left breast was treated, since the mean range of the beta particle of 90Y is 4 mm (Paganelli G. et al.Eur J Nucl Med Mol Imaging 2010;37:203-211). In 2009 in 15 patients who received IART we compared data derived from standard dosimetry with those obtained using voxel dosimetry. The Biological Effective Dose (BED) and Equivalent Uniform biological effective Dose (EUD) were evaluated in order to assess dose distribution heterogeneity. Absorbed doses from standard versus voxel dosimetry showed a difference of + 12±3%. The mean BED values and related standard deviation resulted to be 21.2±4.3 Gy in standard method versus 24.4±3.2 Gy in voxel dosimetry. The BED-volume histograms showed that 68±25% of voxels received a BED greater than 21 Gy. All voxels received a BED greater than 14 Gy. These results indicated that IART can deliver into tumor bed an anticipated boost comparable to a EBRT boost. The absorbed doses evaluated with standard method were systematically lower than those calculated with voxel method (differences up to 16%).The EUD/BEDvoxel ratio in the target was 0.9. This value, obtained thanks to the crossfire effect of 90Y, indicates a biological efficacy comparable to that of uniform dose distribution, thus the hypothesis of uniform activity distribution is not misleading.

A) Scheme of voxel dose calculation using a dedicated software (Matlab® support, Mathworks). Input data were obtained from SPECT images (activity distribution), biokinetic data (planar images) and S-voxel factors for 90Y and 4.4 mm voxels, speciﬁcally calculated for by Monte Carlo simulation (PENELOPE code).

B) BED-volume histograms showed that 73% of voxels received a BED greater than 21 Gy. All voxels received a BED greater than 14 Gy. BED distribution was evaluated considering radiobiological parameters derived from literature ( / =10, μrepair=0.5 h-1). Dose-volume and BED-volume histograms were generated, and the corresponding average BED (BEDvoxel) obtained. EUD was estimated considering the surviving fraction of the target ( =0.3 Gy-1).

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Brain Tumours

The prognosis of high-grade glioma is poor and, in most of cases, recurrence invariably occurs within a short period, despite aggressive therapies, such as surgery, radiotherapy and chemotherapy. Antibodies, particularly IgG, have been used frequently in tumour therapy, due to their ability to bind selectively to tumour-associated antigens. The extracellular matrix tenascin molecule represents the most exploited target for radioimmunotherapy in high-grade glioma, both in systemic and loco-regional approach, using a pretargeted method based on the avidin-biotin system [Paganelli G. et al. Neurol Res. 2006;28(5):518-22; Goldenberg D.M. et al. J Clin Oncol. 2006 10;24(5):823-34]. Our group previously assessed the safety profile and therapeutic efficacy of intravenous and loco-regional Pretargeted Antibody-Guided RadioImmunoTherapy (PAGRIT®) in several groups of patients affected by high-grade glioma. In both routes of administration, anti-tenascin monoclonal antibodies and radioactivity had been administered separately, according to the following scheme: on first day biotinylated BC4 MoAbs, on second day avidin, and on third day 90Y-biotin conjugate (intravenously or via a catheter placed into the surgical resection cavity) [Ferrari M. et al. J Nucl Med. 2006;47(1):105-12]. A new GMP-produced anti-tenascin monoclonal antibody ST2146, with the same specificity as old BC4 but lacking the non-functional light chain, was characterised. ST2146 showed a higher affinity and immunoreactivity, and appears to be a good alternative to BC4 for clinical use [Urbano N. et al. Eur J Nucl Med Mol Imaging 2007;34: 68-77]. We are presently working on possible improvements of such protocols, both in chemical synthesis of suitable reagents and in dosimetry planning. A new biotin-DOTA conjugate in which biotin was chemically linked to the DOTA chelator has been investigated by our group. This compound was chosen as the candidate for the development of a therapeutic radiopharmaceutical for further clinical investigations. The goal of the study was to

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optimize the radiometal chelation with 90Y or 177Lu and to study the stability of the label linked to this new biotin conjugate to support its use in pre-targeting therapy trials. We are studying these possible improvements, as in a retrospective evaluation of our experience in radioimmunotherapy in glioblastoma patients we found interesting results in overall survival. All the patients affected by progressive glioblastoma and were treated according to our 3-step RIT protocols. We observed 64% with progressive disease; 28% with a stabilization of disease and in 21 pts (8%) we observed a partial response. The GBM cumulative survival was 98,4% ± 0,9% at 6 months, 79,2% ± 2,1% at 12 months, 51,7% ± 2,6% at 18 months, and 30,7% ± 2,4% at 24 months. The median survival time from diagnosis was 19 months. These results could represent the basis for further prospective trials, which could also exploit new radioisotopes, such as Lu-177 or alpha emitters, and provide a way to assess timing and scheduling of radioimmunotherapy in the therapeutic algorithm of high-grade glioma patients. Moreover, these data suggest that the use of 90Y-biotin guided by anti-tenascin antibodies, interferes with the progression of high grade glioma, prolonging survival in a large series of patients. These results warrant phase III controlled studies although, due to limited budget it is unlikely that such a controlled study will start soon. Anti-cancer therapies with radiolabelled compounds require tumor dosimetry as key information. Standard methods for dosimetry are able to provide average doses in organs and tumours of spherical shape. More sophisticated techniques based on Monte Carlo simulations have been implemented in order to evaluate the dose conversion factors for 90Y, 177Lu, 131I - the most used radionuclides for radionuclide therapy in locoregional and systemic settings. Moreover, the dose distribution inside spheres and in surrounding volumes was investigated

to represent dose in tumours, radiosensitive bordering tissues and/or disease inﬁltrated tissues. The model studied offers suitable information for dosimetry in several radionuclide therapies. This represents an important tool towards the use of tailored treatments with improved safety and efﬁcacy. The comparison of the results can suggest the choice of a most appropriate radionuclide depending of the clinical situation [Botta F. et al. Eur J Nucl Med Mol Imaging 2008;35(Suppl 2):S201; Strigari L. et al. Eur J Nucl Med Mol Imaging 2008;35(2): P550. Our group is also active in the treatment of meningiomas. These are generally benign tumours and in most cases surgery is curative. However, high-grade histotypes or partially resected tumours frequently recur and therapeutic options are few. External beam radiation therapy (EBRT) is commonly used, although not always effective. We assessed peptide receptor radionuclide therapy (PRRT) with 90Y-DOTATOC in 29 patients with

somatostatin receptor-positive meningiomas recurring after standard treatments. The treatment was well tolerated by all patients. MRI 3 months after treatment completion showed disease stabilization in 19 of 29 patients (66%) and progressive disease in the remaining 10 (34%). Better results, as to morphological response and time to progression, were obtained in patients with grade I meningioma compared to those with grade II–III. We concluded that 90Y-DOTATOC can interfere with the growth of meningiomas and an adjuvant treatment, particularly in atypical and malignant histotypes, deserves further investigation [Bartolomei M. et al. Eur J Nucl Med Mol Imaging 2009; 36:1407-16].

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Gynaecological Tumours

The treatment of gynaecological tumours is based on the organ of origin and on the extent of disease. Every year in Italy, there are 4797 new ovarian cancers, 3418 new cervical cancers, and 7759 new endometrial cancers. The main research efforts are against ovarian cancer which represents the number 1 killer among gynaecological malignancies. The general aim of our research group is to improve treatment outcome and minimize related morbidities. Great attention is also given to new tools and new biomarkers for the early diagnosis of ovarian cancer. Ovarian cancer Basic research: The deepening of our understanding of normal biology has made it clear that stem cells (SCs) have a critical role not only in generating normal tissues, but also in developing tumours. Recent ﬁndings support the concept that cells with SCs properties are integral to the development and perpetuation of several forms of human cancer. Recently, a colture system has been described that allows for the in vitro enrichment and propagation of SCs (i.e. brain and breast SCs), based on their ability to survive in suspension and produce spherical colonies composed of both stem and progenitor cells. The ability of SCs from different tissues to survive in suspension suggests a common underlying mechanism. We applied this colture system to the study of ovarian SCs. Ovarian carcinoma (OC) is the most lethal gynaecological malignancy, most tumours being advanced at presentation. As yet, very little is known about the cell of origin of epithelial ovarian malignancy. Additionally, even though most epithelia are known to be organized according to a hierarchical system based on a dedicated adult stem cell population, the existence of ovary surface epithelial (OSE) SCs has not been demonstrated. We tested the hypothesis that normal and tumor OSCs do exist, developing a suspension colture system to isolate them, in human and mouse. After dissociation of human primary

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OCs (as well as of mouse OC), we isolated a population of cells that can be grown in suspension, forming floating spherical colonies, that we named “ovospheres”. We could demonstrate that cancer ovospheres: i) are clonal; ii) serially grow for up to 7 passages. Using the PKH26 staining, we could isolate cells with stem properties among those constituting the ovospheres. We also obtained human and mouse normal OSE ovospheres and were able to passage them a couple times. To finally prove the “staminality” of the cellular populations that we colture in vitro, we will perform in vivo transplantation experiments, by inoculating single cell suspensions of ovospheres into mice. Our final goal is to define the biological and molecular differences between normal and ovarian cancer SCs, with the aim of identifying more effective cancer diagnostic markers and anti-cancer therapeutic strategies exploiting the cancer stem cell as a target. Diagnosis and prevention: Transvaginal sonography is still the mainstay in the diagnosis of ovarian cancer. The group is involved in an international multicentric study in the discrimination of benign and malignant pelvic masses (IOTAII and III) using 3D sonographic evaluation. The most promising approach to ovarian cancer diagnosis is proteomics; new biomarkers for ovarian cancer early diagnosis (HE4 and proteomics) are being evaluated in a prospective ongoing study. At least 10% of ovarian cancer cases arise in individuals with genetic mutation of BRCA1 and BRCA2. This population represents an ideal model to study prevention strategies: a chemoprevention study with HPR has been started recently. Surgery: Surgery represents the cornerstone of treatment for patients with advanced ovarian cancer. We are evaluating the role of “maximal surgical effort” in patients with advanced ovarian cancer in a prospective study, with particular focus on stripping/full-thickness resection

of the diaphragm, spleenectomy, and rectal resection. We are also interested in exploring the possible role of surgery at relapse prior to second line chemotherapy. A randomized trial as been recently designed to assess whether surgery followed by chemotherapy may improve progression-free survival and overall survival compared to chemotherapy alone in patients with relapsing ovarian cancer. At present, we are conducting a prospective study in order to validate a score of predicting factors for complete resection in platinum sensitive recurrent ovarian cancer. Chemotherapy: Despite several trials conducted in the last 10 years investigating whether at the addition of more cytotoxic drugs may improve survival of patients with advanced ovarian cancer, standard treatment remains the combination of carboplatin and paclitaxel. Patients with newly diagnosed ovarian cancer have been enrolled in the EORTC phase III trail of Erlotinib (maintenance treatment: 150 mg/day orally up to 2 years) versus observation in patients with no evidence of disease progression after firstline platinum-based chemotherapy and in the MIMOSA trial, investigating the role of Abagovomab as maintenance therapy after completion of front line chemotherapy. We are waiting for the definitive results of these trials. Also we took part our participation in a double blind, randomized phase II study, in which we enrolled 10 patients, evaluating the efficacy and safety of Sorafenib compared to placebo in epithelial ovarian cancer patience who have achieved a complete clinical response after primary chemotherapy. Finally, during the last quarter of 2009, a phase III study of the Gynecologic Cancer Intergroup was approved to evaluate the efficacy and safety of pazopanib monotherapy versus placebo in ovarian cancer women who have not progressed after first-line chemotherapy. For patients with relapsed ovarian cancer, the results for a phase III trial were presented comparing the combination carboplatin/pegylated liposomal doxorubicin with the

standard carboplatin/paclitaxel in patients with platinumsensitive disease. The preliminary data demonstrated a significantly longer PFS (HR 0.82, 18% risk reduction) and a better toxicity profile in favour of the experimental arm carboplatin-PLD, which is becoming the new standard in this setting of patients. For platinum-sensitive recurrences, a phase II doubleblind placebo-controlled multi-centre randomized study of ZD4054 (an orally active specific endothelin receptor antagonist) plus carboplatin and paclitaxel or placebo plus carboplatin and paclitaxel is currently ongoing. The primary objective of this study is to evaluate the progression-free survival in patients treated with ZD4054 compared with the standard arm. For partially platinum-sensitive recurrences, it is widely debated whether re-introducing a platinum-based therapy is better than using a non-platinum treatment. The MITO-8 study, recently approved, which compares the use of pegylated liposomal doxorubicin versus the combination carboplatin-paclitaxel, should help to clarify this controversy. Finally, the enrollment of patients with advanced solid tumours in a dose-finding study of Caelyx and RAD001 (a mTOR-inhibitor) is almost completed, where we included 25 patients with ovarian and endometrial cancer recurrences. We are waiting for the definitive analyses of data concerning two phase III studies, recently completed. The first compared PATUPILONE (EPO906) versus pegylated liposomal doxorubicin (DOXIL/CAELYX) in taxane/platinum refractory/resistant patients with recurrent epithelial ovarian, primary fallopian or primary peritoneal cancer. The second evaluated the addiction of VOLOCIXIMAB (an anti-angiogenic integrin inhibitor) to pegylated doxorubicin in the same setting of patients. We are also awaiting the efficacy analysis of AVE0005 (VEGF Trap), administered intravenously every two weeks in patients with platinum-resistant and topotecan and/ or liposomal doxorubicin-resistant advanced ovarian cancer, as well as the definitive data on the combination

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of Caelyx-Velcade (a selective proteasome inhibitor) in patients with ovarian cancer failing platinum containing regimens. Endometrial cancer: Endometrial cancer is the most common gyneacologic tumours. It is more frequent in postmenopausal women, however, 25% of cases occur in premenopausal women. In this setting, the group is testing the therapeutic effect of a levonorgestrel medicated IUD (MIRENA) in patients affected by atypical endometrial hyperplasia or grade 1 endometrioid adenocarcionoma confined to the endometrium in a prospective study; the study has accrued more than 40 patients and a first data analysis has already been completed. Most postmenopausal cases are detected early resulting in a good prognosis for cure and survival with a five-year survival of more than 85%. Initial treatment consists of surgery, surgery and radiation, or hormonal therapy. For those women in whom endometrial cancer has progressed despite available treatment options, chemotherapy is the only option. Chemotherapy consists of taxanes, anthracyclines, and platinum agents. Response to chemotherapy has ranged from 20 to 35% for single agents and up to 70% for combination regimens with progression-free survival (PFS) ranging from 5 to 9 months duration. As a result, further studies are being undertaken to look at multi-drug regimens to determine what combination may improve response rates, increase PFS, and decrease the incidence of side effects (e.g., peripheral neuropathies). Therefore, treatments with advantages over current chemotherapy such as decreased toxicity and improved overall survival are needed. We have recently activated a phase II trial of orally administered deferolimus, a rapamycin-analog that inhibits mTOR, compared to progestin in patients with recurrent endometrial cancer. Cervical Cancer Prevention: The Division is evaluating new modalities of early

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diagnosis and prevention for cervical cancer. The use of HPV testing in the management of patients with abnormal cytology is the subject of three investigations: the role of HPV genotyping in the follow-up protocol of patients conservatively treated for CIN2-3; a comparison of the FDA approved HC2 viral test vs PCR-based methodologies; self sampling vs standard sampling from the physician or nurse. Ongoing studies include also a population-based project of HPV vaccination in 18 year old adolescents. Early cervical cancer: Fertility preserving surgery About 40% of patients with stage I cervical cancer are younger than 40 at time of diagnosis. To preserve the fertility potential in selected and motivated patients, a prospective study has been started in order to evaluate the role of conization and laparoscopic pelvic node dissection in the treatment of young patients with stage IB1 cervical cancer. A subgroup of patients not suitable for this study because of high risk factors will be treated with neo-djuvant chemotherapy followed by conization and laparoscopic lymphadenectomy. Goals of these two studies will be to provide good oncologic outcomes while preserving fertility potential. Advanced cervical cancer: In collaboration with the EORTC Gynecologic Group we are coordinating a randomized study comparing neo-adjuvant chemotherapy followed by radical surgery against the standard approach consisting of concomitant chemo-radiation in patients with FIGO stage IB2- IIA > 4 cm, IIB cervical cancer. Recurrent cervical cancer: Surgery We are investigating new techniques for viscera reconstruction after pelvic exenteration: complication rates and quality of life are the major endpoints. We are also evaluating the role of IORT (intra-operative radiotherapy) after exenteration for side pelvic recurrences.

Sentinel node identification in cervical and vulvar cancer. The presence of lymphnode metastases demands a more aggressive treatment in patients with stage IA2-IB1 cervical cancer. The possibility to identify lymphnode involvement without the need for extensive surgical procedures could allow the selection of patients not suitable for surgery but best treated with radiotherapy , avoiding the co-morbidity of the two treatment modalities. The identification of nodal metastasis by means of PET scan, MRI and lymphoscintigraphy has been carried out in a new trial in order to assess the positive predict value of such diagnostic modalities to avoid unnecessary surgical procedures. To this respect laparoscopy can allow the detection and removal of suspicious node(s) after injection in the cervix of technetium radio-colloid (the day before surgery) and patent blue violet (at the beginning of surgery); we are interested in defining which method is associated with the highest accuracy, in confirming the negative predictive value of sentinel node; and in evaluating different methods for the identification of micrometastases (such as standard staining on serial sections, immuno-histochemistry). The diagnostic modality (lymphoscintigraphy, FDG PET-CT and MRI) associated with the best positive predictive value will be also evaluated in patients with early stage cervical cancer. A prospective observational study has been conducted in patients with early stage squamous cell carcinoma (diameter < 4 cm) of the vulva without suspicious lymph nodes at palpation, planned for surgery (radical local excision or radical vulvectomy in combination with sentinel lymph node procedure). To check the efficacy of staging by the sentinel node procedure, stopping rules have been made. These rules include the occurrence of groin recurrences in those patients with histopathologically negative sentinel node(s). Furthermore a new trial regarding the efficacy of replacing complete inguino-femoral lymphadenectomy by adjuvant radiotherapy will be activated.

routine gynecologic oncology surgery using a computerenhanced robotic surgical system allowing minimally invasive surgery to be more precise and ﬁne-tuned so that more sophisticated procedures can be performed endoscopically. The da Vinci Computer-Enhanced Robotic Surgical System is operated by a surgeon from a remote workstation in the same room where the patient lies consists of a surgeon's viewing/control console with an integrated high-performance three-dimensional vision system and a patient sidecart with four robotic arms responding in real time to the surgeon's movements. Endo-wrist instruments precisely mimic the surgeon's hand and wrist movements and allow translation of the movement onto the operative site. This remarkable technology facilitates suturing and dissection. It also and makes skeletonization and ligation of vessels, dissection of pelvic spaces, and lymph node dissection feel more like conventional open surgery than the standard laparoscopic technique. Furthermore, the learning curve is shorter compared to that of conventional laparoscopy. Finally, the overall increase in the time needed to complete full procedures is spent in equipment set-up, such as positioning the surgical cart at the bedside, and not on the actual operation. In 2009 we performed about 140 surgical operations with the da Vinci System.

Minimally-Invasive Robotic Surgery - da Vinci System At present we are evaluating the feasibility of conducting

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Head & Neck Cancer Research Programme

Modus Operandi The Clinical Head & Neck Multidisciplinary Working Group meets each Wednesday, to discuss each patient referred to IEO with a cancer of the oral cavity, pharynx, larynx, salivary glands and thyroid for plannnig therapy, evaluating treatment outcome, and scheduling follow-up. The last Wednesday of each month the Head & Neck Task Force (including Surgeons, Medical Oncologists, Radiotherapists, Radiologists, Pathologists, Nurses, Speech Therapists, Dietologists, Basic Reseachers) discusses new protocols, reviews on-going trials, diagnostic and therapeutic guidelines. Every third month an invited Visiting Professor will attend the department, meet the staff and give a lecture. Published Clinical researches Compartmental surgery in tongue tumours The aim of curative surgical oncology is to remove the primary tumour with a wide margin of normal tissue. What constitutes a sufficiently wide margin particularly in oral cancer is fundamentally unclear. The currently accepted standard is to remove the primary lesion with a 1.5-2 cm circumferential macroscopic margin. In the last ten years, anatomical considerations in the approach to primary, advanced and untreated tumours of the tongue led us to develop and improve a new surgical approach to their demolition and reconstruction. From July 1999 to July 2009, 155 patients were treated in our Institute, while defining and refining the concept of compartmental tongue surgery (CTS) and its main components: 1) anatomical approach to the disease that requires removal of the primary lesion and all of the potential pathways of progression--muscular, lymphatic and vascular; 2) identification of a distinct territory at risk of metastatic representation of the disease: the parenchymal structures between the primary tumour and the cervical lymphatic chain that include the muscular (mylohyoid), neuro-vascular (lingual nerve and vein) and glandular (sublingual and submandibular) tissues; 3) preparation for a rational reconstruction in consideration of a functional defect resulting from this anatomical demolition. (Calabrese L et al Acta Otorhinolaryngol Ital. 2009).

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US-guided transcutaneous tru-cut biopsy of laryngo-hypopharyngeal lesions Aim of the study was to evaluate the feasibility and performance of ultrasound-guided transcutaneous tru-cut biopsy (USGTCB) in selected patients (with stenosis of airways or difﬁcult intubation or contraindication to general anaesthesia) with untreated or previously treated suspicious laryngo-hypopharyngeal masses. Biopsies were performed with a free-hand technique by a single radiologist. Thirty-six USGTCBs were scheduled in 34 patients (24 males, 10 females; age range 4795 years). Two USGTCBs were not performed, as lesions were not detectable: therefore, 16 USGTCBs were performed for an untreated mass suspicious for malignancy and 18 were performed for a mass suspicious for recurrence after radiotherapy alone, or associated with endoscopic laser surgery or chemotherapy. USGTCB diagnosed 25 squamous cell carcinomas (73.5%) and nine benign lesions (26.5%); no false positives and two false negatives were reported, both in patients previously treated with radiotherapy. The sensitivity, speciﬁcity, positive and negative predictive value of the technique was 92.5%, 100%, 100% and 77.7% respectively, with no major complications. Although biopsy under microlaryngoscopy remains the "gold-standard" technique, USGTCB is feasible, carries the advantages of avoiding general anaesthesia, is suitable for outpatients and is cost-effective. If applied to selected patients, it could be considered for the histological diagnosis of both primary and recurrent laryngo-hypopharyngeal masses. (Preda L et al Eur Radiol. 2009) Laser surgery for early glottic cancer: impact of margin status on local control and organ preservation. Aim of the study was to assess the impact of margin status on disease-free survival, overall survival, and organ preservation in early glottic cancer treated by endoscopic laser surgery. It was a prospective nonrandomized study. A total of 274 patients with untreated (possibly biopsied) cTis, cT1a/b, cT2, cN0 glottic cancer; adequate exposure of the glottic region; no contraindications to general anesthesia; and the ability to give informed consent. Patients with negative margins (>1 mm) were followed, patients with close margins (< or =1 mm) or 1 positive margin

(tumor on margin) had another operation, and patients with more than 1 positive margin had postoperative radiotherapy. Median follow-up was 58 months. Margins were negative in 180 patients, close in 40, and positive in 54. A second laser resection was performed in 36 of 94 patients with close or positive margins. Radiotherapy was administered to 36 patients. Patients with close or positive margins who did not undergo further treatment had a greater recurrence risk (hazard ratio, 2.53; 95% confidence interval, 0.97-6.59, P = .06) than did those with negative margins, mainly owing to relapses in 5 of the 8 protocol breakers with positive margins not treated further. Eight-year relapse-free survival was 88.2%, 5-year overall survival was 90.9%, and the larynx was preserved in 97.1%. Laser removal of early glottic cancer is oncologically adequate with margins greater than 1 mm from the tumor edge. Positive margins require further treatment; close margins may require further treatment depending on tumor characteristics. (Ansarin M et al. Arch Otolaryngol Head Neck Surg. 2009). Role of prophylactic central neck dissection in cN0 papillary thyroid cancer Prophylactic central neck dissection in papillary thyroid cancer is controversial. In this retrospective cohort study, the aim was to assess possible advantages of prophylactic central neck dissection with total thyroidectomy in cN0 papillary thyroid cancer. A total of 244 consecutive patients with papillary thyroid cancer, without clinical and ultrasound nodal metastases (cN0), were evaluated out of 1373 patients operated for a thyroid disease at the IEO, Milan, Italy from 1994 to 2006. Of these 244 patients, 126 (Group A) underwent thyroidectomy with central neck dissection, while 118 (Group B) underwent thyroidectomy alone. Demographic, clinical and pathological features were analysed. Overall recurrence rate was 6.3% (8/126) in Group A and 7.7% (9/118) in Group B, with a mean follow-up of 47 (Group A) and 64 (Group B) months. In Group A patients, 47% were pN1a and all patients with recurrence had nodal involvement (p = 0.002). Survival rate did not differ in the two groups. Nine patients were lost to follow-up. Group A patients were older and their tumours were larger in size; according to the pT distribution, a higher extra-capsular invasion rate was observed. The two groups were equivalent as far as concerns histological high risk variants and multifocality. Nodal metastases correlated with stage: pT1-2 vs. pT3-T4a, p = 0.0036. A lower risk of nodal metastases was related to thyroiditis (p = 0.0034). In conclusion, central neck metastases were predictive of recurrence without influencing prognosis. From data obtained, possible greatest efficacy of central neck dissection in pT3-4 papillary thyroid cancer without thyroiditis is suggested. (Costa S et al Acta Otorhinolaryngol Ital. 200)9. On going clinical researches Locally advanced Head and Neck Squamous Cell Carcinoma: finding new prognostic and predictive factors.

Squamous Cell Carcinoma of the Head and Neck (SCCHN) is the sixth most common cancer worldwide, with an estimated annual incidence of approximately 600,000 cases. Patients with locally advanced disease presentation may or may not be amenable to surgery at the time of diagnosis and until now, we have few parameters to design a specific treatment strategy for each patient. What we know is that, compared to radiation, concomitant chemoradiation could offer an 8% improvement of the 5-y Overall Survival (OS), for patients who are candidates for surgical treatment and a 5% improvement for those who are not. Despite great progress in the overall treatment approach and in understanding the mechanisms that lead to head and neck squamous cell carcinoma, many patients (almost 50%) with locally advanced tumor develop a recurrence within three years from treatment, and about 10% of them experience distant metastases. Only 30% of stage III and 15% of stage IV patients are alive and disease-free at 5 years. In order to better select patients who could benefit from non surgical therapies, prognostic and predictive factors for response to chemotherapeutic or biological target drugs and to radiotherapy are therefore required. Actually, only tumor extension (evaluated by TNM parameters) is used in clinical practice to define the therapeutic approach. Recent papers suggest that some defined pathological tumor characteristics, like related-HPV tumor, could be considered as prognostic and predictive factor of response to therapies. In the same way, functional imaging modalities as FDG PET and functional MRI (diffusion weighted MRI -DWI-MRI and dynamic contrast enhanced MRI – DCE-MRI) could improve the correct staging and predict the response to treatments. The aim of our study is to prospectively collect baseline data from both pathological specimens, FDG PET and functional MRI exams in order to correlate these parameters with response to non surgical treatment and clinical outcome (in terms of Overall Survival and Disease Free Survival) in patients with locally advanced head and neck squamous cell carcinoma. This project is organized in tasks: Task 1: The aim of this task is to evaluate, on the initial biopsy specimen, the expression as well as the prognostic and predictive value of some biomarkers potentially involved in treatment failure: ERCC1, CD105, p16INK4,p53 VEGFR, PTEN. All these biomarkers will be evaluated with immuno-histochemical analysis. Task 2-3: The aim of these tasks is to evaluate the predictive and prognostic value of radiological and nuclear medicine parameters. A baseline functional MRI (DCE- and DWI-MRI) and an FDG-PET will be performed before starting the radio-chemotherapy treatment regimens and objective parameters will be collected. Data from baseline pathological specimens, functional MRI and FDG-PET exams will be analyzed and correlated with the response to treatment and patient clinical outcome.

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Lung Cancer

Lung cancer is one of the leading causes of morbidity and death all over the world and a significant burden on health care resources of most countries. It has been calculated that this tumor has accounted for 219,440 new cases (15% males and 14% females with cancer) and 159,390 deaths (30% males and 26% females deceased for cancer) in the United States during 2009. In Italy, the cumulative risk (0-74 years) of developing lung cancer is 67.7‰ in males (1 case for every 15 men) and 14.1‰ in females (1 case for every 71 women), while the death risk is 56.4‰ in males and 10.4‰ in females (data available from http://www.registri-tumori.it/ incidenza1998-2002/gruppi.html). The scientific activities carried out at the European Institute of Oncology on the theme of lung cancer during 2009 have focused on the following areas of interest. Early detection. The high lethality rate of lung cancer is related not only to its intrinsic biological aggressiveness, which makes its detection too late in most cases with locally advanced or widely disseminated disease at the time of the first diagnosis, but also to the lack of an effective early detection as increased chances of cure may be obtained by lowering tumor stage at diagnosis and targeting high-risk population because of tobacco consumption. In this frame of mind, the diagnostic anticipation COSMOS project (Continuous Observation of Smoking Subjects), which was activated at the European Institute of Oncology in 2004, permitted the introduction of CT scan for screening and validation of noninvasive guidelines for the diagnostic work-up of screening detected undetermined lung nodules. The results have showed that 0.7-1% of screened subjects presented an asymptomatic lung cancer, most of which were early-stage and resectable tumors at the time of the diagnosis, with stage-I disease in more than 70% (89% respectability), and 84% overall survival rate at the 4-year follow-up. A low recall rate was shown in the screening population with high overall compliance. An estimated 49% reduction of cancer mortality was calculated in the screened population in comparison with US smoker population according to published age and sex specific mortality rates. Also,

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a risk model has recently been developed for the COSMOS population, which was able to identify different subgroups of individuals at different lung cancer risk rate according to four clinical variables in addition to the characteristics of baseline low-dose computed tomography (LDCT) scans. These results will be useful to optimize the interval time between consecutive LDCT examinations according to the individual risk of developing lung cancer. Chemoprevention studies. As screening LDCT discovers small, undetermined peripheral nodules, which may be pre- or early invasive neoplasia, we nested into our screening program a randomized trial on budesonide (an inhaled corticosteroid) in current or former smokers with lung nodules to evaluate the role of this chemopreventive agent in the reduction of peripheral lesions. The primary endpoint was a per-subject response after 1-year treatment on lung nodules, whereas secondary endpoints were the decrease in size or number of the lung lesions, the modulation of tumor markers in sputum and plasma, the drug toxicity and the effect on pulmonary function. A total of 202 subjects received 800μg budesonide twice daily or placebo. No difference was observed in the complete or partial response after one year in the treatment arm versus the placebo arm according to per-subject analysis, however the per-lesion analysis revealed a significant effect of budesonide on regression of existing target nodules (p=0.02), the appearance of new lesions was similar in both groups and thus the significance was lost in the analysis of all lesions. The evaluation by nodule type revealed a trend toward regression of nonsolid and partially solid lesions after budesonide treatment. Budesonide was well tolerated with no unexpected side effects identified. Over 80% of cases took at least 50% of the dose (no difference between the two arms) and no significant toxicity was registered in the subject cohort. We concluded that inhaled budesonide did not significantly reduce peripheral lung nodules in individuals at high-risk for lung cancer, but the changes occurring in nonsolid nodules, which are considered precursors of adenocarcinoma, are interesting and deserve further investigations.

Surgical treatment. An issue of the surgical research activity has dealt with the introduction of robotic surgery for the treatment of lung cancer and mediastinal masses. We analyzed the feasibility and safety of robotic approach for the treatment of early stage lung cancer with standard lobectomy and described the technique of robotic assisted lobectomy (RAL) and mediastinal lymph node dissection (MLD). During a 21-month period, 54 patients underwent robotic-assisted lobectomy for early stage lung cancer at our Institution. The approach included three ports and one utility incision. Dissection and isolation of the hilar structures was performed using the four arms Da Vinci System®. Vascular and bronchial resections were done with the use of standard endoscopic staplers. Standard MLD was performed after completion of the lobectomy. The 54 patients were individually matched for age (±5 years), sex, stage, nodal status and forced expiratory ventilation in 1 sec with patients who underwent open lobectomy in the same Institution during the same frame of time, and were divided into three series based on the learning curve according to duration of surgery. In 7 patients (13%) conversion from robotic to open surgery was necessary. The number of overall postoperative complications (20%) and the mean number of lymph nodes removed (18.1±7.9 in open versus 16.8±7.5 in RAL) were similar in both groups. The median time for RAL decreased by 52 minutes between the first and the last two series of interventions. The median length of post-operative stay was significantly shorter after RAL than open intervention (4.5 days robotic in the third series vs. 6 days open). We conclude that robotic approach for lobectomy with mediastinal lymph node dissection is a feasible and safe procedure. It is an acceptable treatment for early stage lung cancer with equal results to open surgery during the early postoperative course. Further investigations are warranted to elucidate the benefit in terms of postoperative pain, respiratory function and quality of life. Genetic investigations. The COSMOS screening program also represents an important resource for genetic investigations, as it deals mostly with early stage tumor patients who are most likely to harbor early genetic defects. By simply counting the number of chromosomes in the tumor cells, it has been possible to ascertain that abnormal genomic contents (aneuploidy) were present in most tumors, despite their early stage. In other words, completely asymptomatic screening-detected tumors exhibited genomic abnormalities that are typical of full-blown malignant tumors. The tumors with a normal genomic content, albeit made up the minority of the cases under evaluation, were assessed for testing the hypothesis that defects affecting subtle DNA regions were anyhow present but not detectable through conventional cytogenetics. These samples have thus undergone further analysis by studying 50000 DNA sequence

variations (polymorphisms) spanning the entire genome and allowing the copy number evaluation at each genomic region represented by each single polymorphism. In this way, it has been possible to identify genomic regions with copy number variations (different from 2 as the normal copy number), which could harbor genes involved in the tumor development acting as players of early transforming events. In particular, our study evidenced 2 common amplified regions located on chromosomes 5 and 8, whose detailed analysis is ongoing. Another aspect of the genetic investigations on lung cancer has dealt with the study of tumor stem cells, using a murine experimental model for isolating stem cells from normal lung tissue as a reference for the identification of tumor counterparts. We have set up an in vitro culturing system, which is able to isolate mouse stem cells as spherical structures, the so-called pneumospheres. Cells constituting these structures can propagate themselves through sequential generations, showing their self-renewal capacity as one would expected by cells with stem properties. Also, pneumospheres-forming cells, when suitably stimulated, can differentiate into different cell types thereby showing their potentialities to give rise to different pulmonary tissues. Finally, cells isolated through the pneumosphere assay show their multi-potentiality, as they can originate, after proper stimulation, cells of tissues other than lung, such as adipose and bone tissues. These results strongly indicate that our system is able to isolate lung normal stem cells. On the basis of these preliminary results, we chose to apply a similar strategy to a mouse lung tumor model obtained by activating a k-Ras mutated allele and to use the pneumosphere assay to isolate lung tumor stem cells. This part of the project is currently ongoing. Next steps will be the translation of the results obtained in this in vivo system to the human counterpart, both normal and tumorous, in order to evidence specific markers able to differentiate normal from tumor lung stem cells. The final goals will be to employ these markers as putative diagnostic indicators (only lung stem cells express these markers, hence their presence should indicate the presence of disease) and targeted therapy (by targeting cells expressing these markers, it should be possible to directly affect the disease). Pathological investigations. A multi-institutional Italian project has been activated in 2009 under the guidance of the Diagnostic Histopathology Unit of the European Institute of Oncology, the so-called I.N.S.P.E.C.TOR (Italian Network for the Study of Pulmonary Endocrine Cell TumORs) project that has been funded by an external company. This is an observational, retrospective, study involving 8 different Italian large hospitals with great expertise in lung cancer pathology, which will raise surgically excised pulmonary neuroendocrine tumors dating back to 1987 and up to the end of December 2007. Main end-points of the project, which has been approved by the Ethic Committee

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of the European Institute of Oncology, will be to construct a vast collection of surgically excised neuroendocrine tumors (the expected number is between 1500-2000) favoring diagnostic, prognostic and targeted therapy studies, to validate the current WHO classification criteria and possibly identify additional diagnostic tools that may be clinically relevant, and to participate in international projects on pulmonary neuroendocrine tumors with a proactive role. Furthermore, another multi-institutional Italian project has been put forward in 2009 by the Diagnostic Histopathology Unit for investigating the issue of the epithelial-mesenchymal transition in pulmonary sarcomatoid carcinomas, which are uncommon variants of non-small cell lung carcinomas. Lung carcinoid tumors. Carcinoids make up a particular and uncommon type of lung cancer, in which surgical resection is currently thought to be the standard for treatment. When surgery is not feasible, common strategies for their cure are missing, since chemotherapy often is not effective because of the low proliferation rate of such tumors. As drug efficacy and treatment strategy cannot be easily demonstrated by large-scale clinical trials, we devised a genetic approach in order to identify genes that could play a role in carcinoid tumorigenesis and, more importantly, become effective targets for therapy. Therefore we have carried out gene expression microarray analyses on a cohort of 6 typical and 7 atypical carcinoids (4 with and 3 without lymph node metastases) matched with 8 normal lung tissue samples (4 for typical and 4 for atypical carcinoids). The analysis of the gene expression profiles led us to identify 254 genes differently expressed in atypical vs typical tumors. Among them, 54 genes are of particular interest since involved in resistance/sensitivity to standard chemotherapy regimens or representing putative targets for new drugs. We are currently validating these data by mean of real-time PCR in a different set of samples. Moreover, we found out a pool of three genes, which specifically identify one of the two subtypes exclusively. Taking advantage of specific antibodies, we are now testing these new markers by immunohistochemistry in a new cohort of patients. If confirmed, these results will help both the clinicians, to choose the best treatment for the patient, and the pathologists, who can benefit of a new tool of diagnostic markers. Pharmacogenetics/pharmacogenomics studies. Translational pharmacogenetics is able to tailor chemotherapy for lung cancer, because it examines the genetic bases of variation in drug metabolism, targeting and transport, and provide the molecular bases for rationally choosing most effective drugs. In fact, most anti-neoplastic agents undergo enzymatic bio-transformation within tumor cells, becoming either pharmacologically active or inactive. Enzymatic activities involved in these processes show a significant inter-individual

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variability, leading to remarkable changes in drug efficacy and tolerability. Therefore, this approach is a promising one to discover the mechanisms of drug resistance/sensitivity/tolerability and optimize the clinical outcome of patients treated with cytotoxic agents. In collaboration with researchers of the Institute of Pharmacology of Pisa University, we quantified by real-time PCR the profile of genes involved in platin and gemcitabine metabolism in patients with non-small cell lung carcinoma to assess the predictive value of this assay in terms of response to drug administration. The first step was to investigate the influence of tumor subtyping and sampling site (primary tumor vs limph node metastasis) on the expression of genes involved in gemcitabine and cisplatin activity in non-small-cell lung cancer patients, including excision repair cross-complementing 1 (ERCC1), human equilibrative nucleoside transporter-1 (hENT1), deoxycytidine-kinase (dCK), 5’-nucleotidase (5’-NT), cytidinedeaminase (CDA), and ribonucleotide-reductase subunits (RR1 and RR2). No differences were observed between adenocarcinoma and squamous cell carcinoma, as well as between primary tumor and lymph node metastasis. These data suggest a similar genetic susceptibility to the gemcitabine-cisplatin regimen for both tumor types and support the use of lymph node or primary tumor for profiling. In the second step we are now trying to find out a correlation between gene expression and resistance/sensitivity to chemotherapy. We have already treated 68 patients with the standard regimen with platin and gemcitabine and measured the clinical response by CT-scan according to RECIST criteria. The statistical analysis is still ongoing. Clinical trials. Two clinical trials are currently ongoing at the European Institute of Oncology to evaluate the efficacy of immunotherapy as adjuvant treatment in non-small cell lung carcinoma patients after complete surgical resection. Briefly, a double-blind, randomized, placebo-controlled phase III study (MAGRIT) with recombinant MAGE-A3 protein plus an immunological adjuvant system is underway in patients with resected, MAGE-A3-positive non-small cell lung cancer. MAGEA3 is a tumor specific antigen that can be used to stimulate the immune system of the patients against cancer cells expressing this protein. A second closely correlated trial (ONCO RD-001) is currently open and is aimed at studying whether the expression of MAGEA3 and other proteins in tumor tissues and corresponding serum samples may change according to anti-cancer treatments.

(Neuro) Endocrine Tumours

(Neuro)endocrine tumours (NETs) arise from dispersed neuroendocrine cells, distributed almost ubiquitously in the body. The majority arise from the digestive system and are able to produce a number of specific hormones, such as gastrin, insulin, serotonin, somatostatin, glucagon, pancreatic polypeptide, VIP in gastro-enteropancreatic (GEP) tract, catecolamines in adrenal medulla, ACTH, GH, prolactin, FSH, LH or TSH in anterior pituitary, PTH in parathyroid gland tumours. Conversely, the majority of these cells are also able to produce chromogranins and synaptophysin, which are considered aspecific markers. Therefore, from a clinical point of view, NETs can be either functioning or non-functioning. Since 2000, the WHO classification of endocrine tumours has clearly defined the neuroendocrine phenotype. NETs tend to be slow growing (although aggressive forms exist) and are often diagnosed when they have already metastasised, thus when a radical treatment is no longer possible. Treatment of NETs is typically multidisciplinary and should be individualised according to the tumour type, burden, and to symptoms. Therapeutic tools in NETs include surgery, interventional radiology and medical treatments such as somatostatin analogues, interferon, chemotherapy, new targeted drugs and peptide receptor radionuclide therapy (PRRT) with radiolabelled somatostatin analogues. Surgery is crucial in many phases, from the eradication of the primary to the debulking of metastatic lesions, in view of other therapies. It is also used to control hormonal symptoms. Interventional radiology techniques (TACE, RFA, HIFU, radioembolization) in NETs are important, due to the common spread to the liver with hypervascular metastases. Medical therapy is used for treating symptoms and/or reducing tumour growth. Traditional chemotherapy is not commonly applied in well-differentiated NETs, since most are slow growing. However, schemes based on platinum derivatives and etoposide are considered in poorly differentiated and/or rapidly progressive NETs, but the choice of the regimen is based on the site of the primary and the histopathological differentiation and proliferation index. NETs usually over-express somatostatin receptors on their cell surface, thus enabling the therapeutic use of somatostatin

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analogues. One of the basic tools for NETs, in fact, is somatostatin analogue biotherapy, which is able to reduce signs and symptoms of hormone hypersecretion, to improve quality of life and to slow tumour growth. Interferons, and particularly -interferon, have been used in NETs, with similar therapeutic effects. . Presently, the combined use of -interferon and somatostatin analogues as first-line therapy is not justified by data in literature, while it could be indicated after progression to a single agent. [Fazio N. Ann Oncol 2007; 18(1):13-9] Nowadays, new molecular drugs, targeting small cellular proteins or messengers involved in proliferation, are being experimented in phase II studies. The most efficient and studied are vascular endothelial growth factor (VEGF) and mTOR inhibitors. Radiolabelled somatostatin analogues have been experimented in NETs for more than a decade. Several clinical trials have indicated that PRRT with somatostatin analogues 90Y-DOTATOC and 177Lu-DOTATATE is an efficient tool in the management of NETs. Present knowledge and our experience at the European Institute of Oncology (IEO) indicate that it is possible to deliver high activities, and therefore high absorbed doses, to tumours expressing sst2 receptors, with achievement of partial and complete objective therapeutic responses in up to 30% of patients. Side effects, involving the kidney and the bone marrow, are mild if adequate renal protection is used [Bodei L. et al. Eur J Nucl Med Molec Imaging 2004;7:1038-1046]. Nowadays, tumour candidates for PRRT with radiolabelled somatostatin analogues are basically sst2 expressing NETs, mainly of the GEP and bronchial tract, but also pheochromocytomas, paragangliomas, medullary thyroid carcinomas, and, at least theoretically, any other tumour histotype known and documented as over-expressing sst2, as seen at diagnostic OctreoScan scintigraphy [Bodei L. et al. J Endocrinol Invest 2009;32(4):360-9]. Patient are selected on the basis of a somatostatin receptor scintigraphy or, more recently, a receptor PET with 68Gaoctreotide. During 2008, the new automated synthesis module for the radiolabelling of 68Ga radiopharmaceutical has been installed in out Institute, along with a GMP 68Ge/68Ga 1.1 GBq generator with high elution yields. Optimization of the synthesis and quality control of 68Ga-DOTATOC has been carried out

and new compounds (such as receptor peptides) are under investigation for 68Ga radiolabelling. In order to improve the predictivity of dosimetry as regards renal toxicity, specific studies have been focused on radiobiological models have been applied to PRRT by means of the Biological Effective Dose concept. An important correlation has been found between BED values and renal toxicity, with results overlapping the curves derived for external beam radiation therapy [Wessels BW, et al. J Nucl Med. 2008;49(11):1884-99]. In particular, a threshold BED of 33 Gy for renal damage and a BED50 (associated to 50% probability of manifesting toxicity) of 44 Gy were found. The analysis of clinical results confirmed that multiple cycle approach in PRRT allows to lower the side effects on kidney, as indicated by radiobiology models [Cremonesi M. et al. Q J Nucl Med Mol Imaging. 2010;54(1):37-5]. Finally, the influence of risk factors (such as hypertension, diabetes, etc.) has been assessed as a lowered tolerability of the renal parenchyma and recovery of renal parameters [Bodei L, et al. Eur J Nucl Med Mol Imaging. 2008;35:1847-56]. Following an internal guideline, at IEO, patients are discussed weekly by the multidisciplinary team on NETs and, according to radiobiological principles, the indication for PRRT is preferably given in the minimum disease as possible, within a short timing after radiological and/or surgical debulking when feasible [Oyen W.J. et al. Ann Oncol. 2007;18(11):1782-92] Recently, in order to further increase the objective response to PRRT, particularly in aggressive, metabolically active tumours, a combined treatment of 177Lu-DOTATATE with the radiosensitizer capecitabine, has been proposed and tested on a small patients’ population. As GEP NETs may also express cholecystokinin 2, bombesin, neuropeptide Y, or vasoactive intestinal peptide receptors, even simultaneously, the potential availability and biological stability of radio-analogues of these peptides will improve in the future the multireceptor targeting of the neuroendocrine cell.

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Prostate Cancer Research Programme

SURGERY See also the dedicated chapter on robotic surgery RADIOTHERAPY (See also the chapter on the Division of Radiotherapy) During the past decade, radiotherapy has confirmed its role as a viable modality in the management of localized prostate cancer. Modern radiotherapy techniques in use at the Division include high-precision external beam radiotherapy (ERT) in the form of 3D-conformal dynamic arc radiotherapy (3D-CRT), Intensity Modulated Radiotherapy (IMRT), Image-Guided Radiotherapy (IGRT) and brachytherapy with permanent radioactive seed implants. High precision approach is expected to reduce significantly radiotherapy side effects, in particular late proctitis. In prostate cancer, minimizing radiation-induced urinary and rectal complications is of a great value. They represent a dose-limiting factor for dose-escalation and, when present, significantly affect the patient’s quality of life even long time after treatment. Interestingly, recent in vivo and clinical data suggest that prostate cancer may benefit from hypofractionation (higher dose/fraction, shorter overall treatment time) due to the lower / ratio than that of the rectum and other late responding tissue ( / for prostate cancer is 1.5 Gy and for rectum > 4 Gy). The / ratio describes the sensibility of the tissue to the fraction size. The low / ratio typical to the prostate tumor cells indicates high sensibility to the fraction size. This means that increasing the fraction size over the conventional one (>2 Gy/fraction) should increase the tumoricidal effect of radiotherapy. Such increase in the dose fraction should have minor effect on the normal tissue due to its higher / ratio (as mentioned earlier, 4 Gy and 1.5 Gy for rectum and prostate cancer, respectively). Some studies have been launched to investigate the clinical benefit of hypofractionation in the prostate cancer, and majority of them use daily imageguided radiotherapy (IGRT) systems. Apart from hypothetical radiobiological advantages, hypofractionated schedules would significantly increase convenience to patients, patient compliance and quality of life. Moreover, shorter radiotherapy schedules can allow to reduce the waiting lists in busy radiotherapy

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centers and to increase the number of treated patients. While the role of brachytherapy has been confirmed in the management of patients with favourable prognostic factors, several questions still remain unanswered regarding its efficacy for patients belonging to the intermediate prognostic group. In patients with intermediate or unfavourable prognostic factors, recent findings from dose-escalation studies, based on 3D-CRT, indicate that prostate cancer can be a relatively radioresistant disease. In order to achieve optimal results in terms of local tumour control, therefore, doses above 78 Gy ought to be administered. In that patient population the combination of external beam radiotherapy with High Dose Rate brachytherapy will be tested. In conclusion, the main research activities in prostate cancer include: • Image-guided radiotherapy (IGRT): daily prostate localization with use of one out of three IGRT systems installed in the Division of Radiotherapy (b-mode acquisition and targeting ultrasound system, X-ray 6D system, cone beam computer tomography system) allowing for the verification of set-up errors and organ motion (fig. 1-3). • Hypofractionated RT with scheme of 2.7 Gy/day up to 70.2 Gy in 5 weeks (radiobiologically equivalent to the conventional scheme of 84 Gy in 42 fractions over 8 weeks). In all patients IGRT is applied. Various dosimetric and clinical aspects are being investigated including quality of life evaluation. • Intensity Modulated Radiotherapy using dynamic RapidArc modality associated with Image-Guided Radiotherapy (IGRT) verification and hypofraction for post-prostatectomy irradiation. • Set-up verification with electronic portal imaging device (EPID): Institutional EPID algorithm has been established (daily EPID for the first 5 fractions) – applied for the patients treated out of the IGRT/hypofractionated protocol. • Institutional dose volume histograms (DVH) constraints have been established based on the follow-up data and review of the literature. Several planning studies on the dose distribution and the optimisation of the treatment plans have been

published and the other are ongoing. • Intraoperative radiotherapy IORT (anticipated boost) in intermediate and high risk prostate cancer. • Postoperative radiotherapy vs. salvage radiotherapy: indications, the role of combined RT and endocrine therapy. • Stereotactic radiotherapy for oligometastatic prostate cancer patients. • Artificial neural network use to predict acute rectal toxicity after external beam radiotherapy for prostate cancer (collaboration with and Politecnico of Milan). • Endocrine therapy: optimisation of the combined treatment: radiotherapy and endocrine manipulation, prevention of side effects in the patients treated with bicalutamide (ongoing phase III study), observational national study on endocrine therapy for prostate cancer. Medical Oncology The course of prostate cancer from diagnosis to death is categorized as a series of clinical states defined by extent of disease (localized disease, rising PSA after local therapy, advanced disease with absence or presence of detectable metastasis) and by response to androgen deprivation. Our scientific activity on prostate cancer addresses the issues commonly encountered by practising oncologist. The research themes varied including: -Development of surrogate markers that may have utility in predicting prognosis and monitoring the antitumor effects of treatment in castration-resistant prostate cancer. -Clinical trials addressing new drugs in patients with castration-resistant prostate cancer -Clinical trials including biological agents targeting different critical points of the signaling cascade or proteins of the mitotic machine. Despite its limitations, PSA is the best tumor marker of prostate cancer currently available in clinical practice. We are developing an alternative biomarker strategy, testing the prognostic and predictive value of Circulating Tumor Cells (CTCs) in prostate cancer, using the CellSearch System, an immunomagnetic method for detecting CTCs (see specific chapter) automated, standardized and approved by the Food and Drug Administration for routine clinical use in metastatic breast cancer and recently also in Castration-Resistant Prostate Cancer (CPRC). Two different populations are under investigation in our projects: • Patients with Castration-Resistant Prostate Cancer (CRPC) who are starting first or second line systemic treatment for advanced disease. In this population we are exploring the prevalence of detectable CTCs, the correlation between the presence of CTCs and known independent prognostic factors such as haemoglobin, alkaline phosphatase and PSA values

and their predictive and prognostic impact. Beside, a relevant aim of this study is to determine the predictive role of changes in CTCs number during chemotherapy courses as an early surrogate marker of response. Thirty five patients of the 54 planned were included since the end of 2008. Preliminary analysis showed 71% of prevalence of intact CTCs. The baseline value of CTCs showed a positive correlation with PSA levels (p-value=0.003), serum alkaline phosphatase (p-value <0.001) and a negative correlation with Hb value (p-value 0.039). A value of CTCs >20/7.5 ml of blood, after 6 weeks from the beginning of the therapy, correlates with a significant decrease in progression free survival (p-value=0.025). These data strengthen the trial hypothesis about the prognostic and predictive value of CTCs but we need a larger sample population and the enrolment is ongoing to reach the target population number. • The second group we are going to study, includes patients with clinically localized prostate cancer eligible for radical prostatectomy or curative radiation therapy. Ninety patients, with biopsy proven prostate cancer, localized disease (clinical stage T1-3NxM0, any Gleason score) suitable for curative local treatment (radical prostatectomy or curative irradiation-external or brachytherapy) will be enrolled In this study. The aims of this project are to investigate the prognostic value of CTCs before and after curative treatments, the correlation between CTCs and other known prognostic factors and biomarkers. We also will evaluate the correlation between CTCs levels and acetylation status of primary human prostate tumor cells. Advanced disease is associated with prostate cancer-specific mortality and morbidity due to disease. First line therapy of metastatic prostate cancer is androgen blockade with bilateral orchiectomy, diethylstilbestrol or luteinizing hormone releasing hormone (LHRH) agonists given with or without an anti-androgen (e.g. flutamide, bicalutamide, or nilutamide) which provides effective palliation, temporary tumor control or regression, and biochemical responses (e.g. decline in serum prostate specific antigen PSA in 80- 85% of patients). Despite the established efficacy of hormonal therapy as first-line treatment of metastatic prostate cancer, virtually all patients will eventually develop disease progression that is resistant to hormonal treatments. Docetaxel and prednisone demonstrated an improvement in survival of approximately 2 months compared to mitoxantrone and prednisone, and provided symptom palliation in less than half of patients in metastatic Castration-Resistant Prostate Cancer. To improve the Docetaxel results, different drugs are under investigation in this setting of patients. • Dasatinib, a potent orally active tyrosine kinase inhibitor, is

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under investigation in a, randomized, double-blind phase III trial. This trial comparesDocetaxel/prednisone and Dasatinib with Docetaxel/prednisone and placebo in castration-resistant prostate cancer. It has been approved by our Ethic Committee and will start early in 2009. Dasatinib in a potent, orally active tyrosine kinase inhibitor and has been shown to inhibit at least 5 protein kinases families: SRC family kinases, BCR-ABL, KIT, EPHA2 and PDGF receptor, all of which have been implicated in oncogenesis as well as the expression and persistence of malignant phenotypes. Besides, complex interactions of the Androgen Receptors with other signalling pathways play a role in hormonal resistence of prostate cancer. Src family kinases, PDGFR- , c-kit and other receptor tyrosine kinases found to be constitutively activated, overexpressed or activated by androgen receptors in prostate cancer. Primary end-point of the study is overall survival. • MDV3100 is under investigation in a randomized, double bind, placebo controlled efficacy and safety study in patients with progressive castration resistant prostate cancer who have previously treated with docetaxel-based chemotherapy. It has been approved by our Ethic Committee in 2010. MDV 3100 is a novel small molecule androgen receptor antagonist selected for its activity against prostate cancer cells with overexpressed androgen receptor. MD3100 also inhibits androgen receptor function by blocking nuclear translocation of the androgen receptor and DNA binding; it has no known agonist activity when the androgen receptor is overexpressed. Under investigation is another patient population, represented by men with non-metastatic castration-resistant prostate cancer with biochemical failure to hormonal manipulations (PAS failure). There is no standard therapy for men with a PSA-only recurrence before instrumental evidence of metastastic disease and numerous therapies are currently being used, including watchful waiting. • A multicentric phase III, randomized, placebo controlled, double-blind study has been approved at the end of 2008, to access the efficacy and safety of ZD4054 a specific blockade of the endothelin A (ETA) receptor, which may lead to the inhibition of multiple processes in cancer such as tumour cell proliferation, tumour cell survival, tumour angiogenesis and the pathophysiology of bone metastases. This clinical trial will test if the Endothelin A Receptor Antagonist ZD4054 can improve progression-free survival and overall survival against a background of existing prostate cancer treatments. All patients participating in this clinical trial indeed will receive existing prostate cancer treatments in addition to trial therapy. In addition half the patients will also receive ZD4054, and half the patients will receive placebo.

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1 Image guided radiotheraphy systems based on the ultrasound prostate localisation

2. Stereoscopic X-ray imaging

3 On-board images

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High Intensity Focused Ultrasound (HIFU)

Introduction High intensity focused ultrasound (HIFU) is a highly precise medical procedure using focused ultrasound energy for burning and destroy the tumor tissue at depth within the body, selectively and without harming overlying and adjacent structures within the path of the beam. The possibility that focused ultrasound therapy might be developed as a result of controlling local heating phenomena was introduced by Lynn et al in the 1940s, but the technique was not developed at that time because of inadequate targeting methods. The advent of more sophisticated imaging has led to a resurgence of interest in HIFU. Unlike radiofrequency or cryoablation, which are also used to ablate tumors, HIFU is completely non-invasive and can be used to reach tumoral areas that are deep within the body, if there is an acoustic window for allowing the transmission of ultrasound energy. Preliminary reports underline a reduced toxicity with HIFU ablation compared with other ablation techniques because of the non-invasive nature of the procedure. First devices never used widely in clinical practice were trans-rectal probes, which have been used predominantly to treat the prostate cancer. Extracorporeal devices are signiﬁcantly larger and can be used to treat a variety of problems, most commonly intra-abdominal solid tumors. As a result, US-guided HIFU equipment

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Focused transducer

these extracorporeal devices use transducers with a longer focal length and use both ultrasound or MRI for targeting the organ. Background The JC HIFU system (installed since 2007 at IEO), is an extracorporeal device using a 20-cm-diameter plane lead-zirconate-titanate disc transducers with an aluminium alloy lens with focal length of 16 cm, driven at 0.8 MHz. It operates at relatively high intensities up to 20,000 W/ cm2. The imaging probe is situated in the centre of the high-intensity focused ultrasound transducer allowing for real-time US image monitoring during treatment. Another available extracorporeal device uses an MRI-compatible 10-cm-diameter focused transducer with an 8-cm radius of curvature, operating at 1.5 MHz, and only licensed for use with uterine ﬁbroids. Guidance and monitoring of acoustic therapy is most important to ensure that the desired region is treated and to minimize damage to adjacent structures. Real-time imaging, such as ultrasound, ensures that HIFU beam targeting is maintained within the correct area throughout the procedure. Both MRI and Ultrasound methods have their advantages and disadvantages. MRI has the advantage of providing temperature data within seconds after HIFU exposure (sonication). However, MRI guidance is expensive, labour-intensive and of lower spatial resolution in some cases, although it is superior to sonography in obese

patients. Sonographic guidance provides the benefit of using the same form of energy that is being used for therapy. The significance of this is that the acoustic window can be verified with sonography. Successful treatment of a small liver metastasis from colon cancer close to inferior vena cava. (a). US (ultrasound) image obtained before HIFU treatment (b). US image obtained immediately at the end of HIFU ablation shows hyperechoic region in the treated area.

Imaging methods to assess HIFU treatment are similar to those used to assess the response to other methods of ablation such as radiofrequency ablation and include contrast enhanced CT and MRI. In addition, the use of micro-bubbles ultrasound contrast-enhanced is also being examined as a method for evaluating efficacy of HIFU traetment. These methods all examine the change in vascularity of the treated volume. PET scan is another diagnostic tool currently used for oncologic applications; it is used for assessing changes in metabolic activity after HIFU treatment. Diagnostic ultrasound has an excellent safety profile, with no clinically significant deleterious biologic effects having been reported using current diagnostic equipment. However, at high energy, ultrasound can result in tissue heating and necrosis, cell apoptosis and cell lysis. Several mechanisms are involved in the tissue damage induced by HIFU. Localized heat generation due to absorption of the acoustic energy is the main biological effect in tissues where it rapidly raises temperatures from 55°C to 100°C, causing coagulation necrosis within a few seconds. The precise and well delimited US focusing (e.g., 1 mm diameter and 9 mm length), minimizes the potential thermal damage to the tissue located along the acoustic pathway, because the energy is much lower outside the focal region. Mechanical phenomena, in addition to thermal effects, are associated only at high energies. PET-CT image scan before (a) and after (b) HIFU treatment of bone metastasis from multiple myeloma.

Cavitation is the most important mechanical phenomenum; it can be deﬁned as the creation or motion of gas bubbles

within an acoustic field due to alternating compression and expansion of tissue as ultrasound waves propagate through it. Thermal effects, acoustic cavitation and vessel damage can occur simultaneously within the targeted tissue in HIFU ablation. Therefore, the coagulation necrosis induced by HIFU can be considered as the result of biological effects from a combination of heat, cavitation and vascular destruction on tissue. Combination between imaging and technologies for local therapy has made ablative procedures more reliable and practical, allowing for safe and feasible application of HIFU treatments in clinical practice. USguided extracorporeal high-intensity focused ultrasound has been recently used to treat patients with various kinds of malignancies, including liver, breast, kidney, bone, and soft tissue both in Asia and Europe. Magnetic Resonance guided Focused Ultrasound is currently used in North America, Europe and Asia for treating uterine fibroids. Investigation and applications of HIFU are growing rapidly worldwide. Although major clinical application for HIFU is in treatment of benign and malignant solid tumors, several other potential therapeutic applications of HIFU are being investigated, including thrombolysis, arterial occlusion for the treatment of tumors and bleeding, haemostasis of bleeding vessels and organs. In China, the first HIFU procedure was performed for male patient with tibia osteosarcoma on December 1997. From December 1997 to December 2007, more than 10,000 patients with primary and metastatic liver cancer, malignant bone tumor, breast cancer, sarcoma, kidney cancer, pancreatic cancer, bone metastatic tumors, uterine fibroids, breast fibroadenoma, and hepatic hemangioma, have been treated with US-guided HIFU ablation in more than 40 HIFU centres. The group from Chongqing University has reported to date the largest series of HIFU clinical applications, in which 1038 patients have been treated with US guided HIFU in ten centers for the purpose of either cure or palliation. Liver Several studies on HIFU treatment of HCC and secondary liver metastases in human clinical trials have been already published. Wu et al. used an extracorporeal HIFU device to treat 68 patients with liver malignancies showing complete tumor ablation in 30 cases in which surgical excision followed HIFU treatment. HIFU has also been used for palliation in patients with advanced-stage liver cancer by Li CX et al. observing after treatment 87% of symptomatic improvement. In Oxford, UK, in 2005 a total of 22 patients with liver metastases were treated with HIFU revealing a favourable adverse event profile when compared to open or minimally invasive techniques. The only Randomized Clinical Trial is from Wu et al. that randomized patients between transarterial chemoembolization (TACE) alone and HIFU after TACE. The median survival times were 11.3 months

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in the combined HIFU–TACE group and 4 months in the TACE-only group (p = 0.0042). To date, both animal and human subject investigations show significant promise in the treatment of hepatic malignancies with HIFU. Authors from Chongqing University (China) observed that ablation with HIFU can safely achieve large areas of coagulation necrosis in tumors close to major blood vessels, without damage to vascular integrity. Hence this therapy can be applied to treat many patients who can not undergo conventional treatment techniques due to either absolute and relative contraindication or tumor location. Kidney Treatment of renal tumors has been evaluated in 2006 (Roberts WW) in animal models. HIFU ablation of renal tumors in humans is still within early clinical trials. A clinical feasibility study using HIFU to ablate renal tumors has been performed on eight patients who showed histologic evidence of ablation in the treated areas after excision. Pancreas HIFU for palliative treatment of pancreatic cancer may be useful in patients who develop symptoms that would benefit from local tumor control. Results from an open-label study in China in 251 patients with advanced pancreatic cancer (TNM stages II–IV) suggested that HIFU treatment can reduce the size of pancreatic tumors without causing pancreatitis and thus prolonging survival. All treated patients showed a survival benefit with a symptomatic relief observed in 84% of patients with visceral pain due to pancreatic cancer after treatment with HIFU. Bone The HIFU technology has been used also for the treatment of bone and soft tissue malignancies with good functional outcome as a limb-preserving alternative to conventional therapy. In 2004, Wu et al described the use of HIFU system for treating 153 cases of bony malignancy, and 77 cases of soft tissue sarcoma. The interesting aspect observed was the spontaneous regression of lung metastases after HIFU ablation of a primary bone tumor, in the absence of any other systemic therapy. Such findings have not previously been reported and may imply a beneficial immunological process such as specific antitumor immune sensitization. In contrast to other ablative techniques, such as radiofrequency ablation and cryoablation, primary malignancies have been treated more frequently than secondary tumors. In summary, the intention of treatment has been both curative and palliative, depending on clinical circumstances. Hence HIFU has been proposed as a limb-sparing alternative to amputation, and it is implied that it should be used according traditional surgical principles, alongside neoadjuvant and adjuvant chemotherapy where appropriate.

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Uterine fibroids Clinical trials correlated both treatment volume and symptomatic response to the ablation of uterine fibroids. Stewart et al. reported a series of 109 premenopausal women who underwent MR-guided HIFU ablation of symptomatic uterine leiomyomas. A validated questionnaire, revealed symptom reduction at 6 months in 71%, and 51% at 12 months. The authors concluded that this treatment results in short-term symptom reduction for women with symptomatic uterine leiomyomas with an excellent safety profile. Ieo experience At IEO first study on coagulative lesion formation by high intensity focused ultrasound on ox liver in vitro tests started in 2007; it included a sequence of experiments under different exposure conditions followed by tissue sectioning. We confirmed previously published data about the development of HIFU-induced tissutal lesions, by changing different parameters as acoustic propagation, absorptive heating, power and intensity of sonication in bovine liver as a labora-

tory model. At the end of “in vitro Phase” we concluded that HIFU is a feasible and a safe technique to perform a precise tissue ablation at different power and intensity of sonication. As a comprehensive cancer center from November 2007 and December 2009 at EIO, 71 patients (age range 16-77 years) have been treated with ultrasound-guided HIFU, within a feasibility study phase. All patients included in this study were deemed not candidate for surgery, nor suitable for local ablative techniques such as radiofrequency ablation (RFA), transcatheter arterial chemoembolization or embolization (TAE), or were unwilling to have any of those treatments. Among the 71 patients, eight with HCC, 12 with liver metastases from colorectal cancer (CRC), two with chest wall metastasis from CRC, one with liver and lung metastasis from CRC, two with soft tissue metastasis from CRC, one

with liver and lung metastases from CRC, two patient with liver metastasis from breast cancer, two patients with 2 liver metastases from neuroendocrine tumor (NET), five patients with bone metastases from breast cancer, one with pubic bone metastasis from bladder cancer, one with osteosarcoma, one patient with leg metastasis from lung cancer, one with iliac metastasis from multiple myeloma, one with abdominal liposarcoma, three patients with pancreatic tail NET. With the aim of palliation we have treated twelve patients with pancreatic cancers, eleven at the advanced stage with no indication for surgical resection; one patient had local relapse after surgery. In these patients we observed disappearance of visceral pain. All of them had chemotherapy and radiotherapy with progressive local disease at the time of HIFU treatment. Patients were evaluated with conventional tests before treatment. MDCT or/and PET-CT or MRI were performed as baseline-imaging. Specific bowel preparations were required for patients with pancreatic cancers and for patients with abdominal tumors close to the bowel. The patients were asked for fasting overnight the day before HIFU treatment. The day of procedure skin surface overlaying the lesion was shaved and purified in every patient, for avoiding the presence of hairs within the acoustic pathway. In order to treat the symptoms and obtain a dimensional reduction we performed HIFU ablation on 16 cases of uterus fibromyoma since march 2009, all in Day Surgery. We obtained symptoms disappearance and uterine myoma ablation in all ladies which were not observed any side effects. Control MRI showed lesion shrinkage in 11 patients

had one session of treatment, fifteen patients had a second session after a mean period of 6 months from first treatment. PET-CT or/and MDCT showed complete response in about 80 % liver metastases; bone lesions were palliated in symptoms and PET-CT detection; MDCT and PET-CT showed lesions of chest wall, leg muscle, abdominal wall and lung completely ablated; the huge liposarcoma has been almost completely ablated at MRI; pancreatic lesions were palliated in symptoms and at PET-CT or MRI. MRI at 1, 3 and 6 month a complete ablation of all the uterine fibromyoma without side effects. Local oedema was observed in 4 patients. No other complications were observed. All the patients returned

Pancreatic cancer : PET-CT images. (a). Pre-treatment PET-CT image showing positive of the tumour. (b). One month posttreatment, PET-CT image showing negative of the tumour.

home 1-3 days after treatment.

after 6 months of follow-up. For this study, HIFU therapy has always been performed under general anaesthesia, necessary for preventing the patient from experiencing any pain or discomfort and to ensure immobilization during treatment. As feasibility study IR team considered procedure timing as fundamental. Room time, including preparation time and treatment time, ranged from 1 hours 50 minutes to 7 hours 5 minutes. The sonication time, deﬁned as the exposure time, which was related to the tumor size, tumor site and blood supply, ranged from 1 minute 20 seconds to 172 minutes. Fifty-six patients

HIFU of Uterine myoma. a) MRI imaging; b) intraoperative UScontrast media showing the vascular feeding; c) US planning before HIFU; b) US-contrast media administered after HIFU procedure showing complete ablation of the myoma.

Early Breast Cancer Over the past decades, there has been a radical change in the surgical treatment for breast cancer. moving towards a more conservative approach, achieving the same clinical efficacy and maintaining the integrity of the woman’s body. The IEO Breast Division is highly involved in clinical research to improve the effectiveness of breast cancer treatments reducing invasiveness and potential side effects of surgical treatments. Many papers has been published on the results on HIFU treatment for breast cancer. Although the results are very encouraging and significant, the small number of cases and the non-homogeneous of enrolling criteria and post histopathological and/or instrumental HIFU evaluation, leave large space for scientific speculation. Along this perspective, from the beginning of 2008, we started a feasibility study on breast cancer treatment by mean HIFU approach, paying particular attention to establish very restrictive criteria for patient recruitment. The study consists on HIFU treatment of selected breast cancer and surgical resection at least one week after treatment, in order

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to evaluate histological findings such as residual viable tumour, coagulative necrosis extension and so on. All patients enrolled for HIFU treatment must meet all of the following inclusion criteria: • Female subject, age > 18 years; • Diagnosis of breast cancer, confirmed by breast corebiopsy; • Single tumour • Clinical stage cT <1,5cm; cN=any; cM=any • Lesion boundaries visualized with US imaging • Lesion circumscribed at least more than 5mm from skin or rib cage • Lesion circumscribed at least more than 20mm from nipple areola complex • passed the HIFU Simulation test • patient able to read, understand and sign the study specific informed consent form; The exclusion criteria were: • Presence of skin ulceration and/or scar • Presence of widespread pathological microcalcifications Before HIFU treatment, all patients were subjected to a careful clinical assessment including anamnesis and pre-surgical routine tests. In addition, each patient was re-evaluated by a radiologist with High-Frequen-

bubble of air within the acoustic pathway. The day before surgery, the patient underwent radiological investigations (breast US and digital mammography) for verifying the changes in the tumour area, caused by thermal damage. Surgical specimen was extensively examined histologically in order to verify the thermal damage and conﬁrm the absolute integrity of the surrounding parenchyma. Only 16 out of 58 patients (age range 41-74 years), initially included according to clinical reports at the ﬁrst medical visit, were selected and then treated by mean of USgHIFU. Tumour lesions ranged from 5 mm to 15 mm. All patients, included in the study, were deemed candidate for breast conservative surgery that was performed after 1-6 weeks Cosmetic result 25 days after HIFU; before surgery

from HIFU treatment. Great importance was given to the time of the procedure. Room time ranged from 3 hours 55 minutes to 1 hour 20 minutes. The overall treatment time, deﬁned as the time from the beginning to the end of HIFU energy emission, ranged from 1 hours 55 minutes to 8 minutes. Sonication time, deﬁned as the exposure time to HIFU energy, ranged from 15 minute 4 seconds to 38 seconds. Intraoperative image: fascia of muscle pectoralis major has been also involved by HIFU, due to the deep position of treated cancer

cy linear array US probe and digital mammography. All HIFU treatments were performed in a single session under local anaesthesia plus MAC (Monitored Anaesthesia Care) in order to improve the room time and preserving patient comfort and collaboration, although at the beginning of experience, general anaesthesia was used in order to avoid any possible external effects disturbing the treatment as pain, discomfort and involuntary movements. The day of procedure, skin surface overlying the lesion was degassed and degreased with 95° alcohol to remove micro-

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No side effects were observed after the HIFU treatment. Only soft local mammary oedema was noted overlying the

From Left to Right (H&E staining): 1-2: coagulation necrosis (x100), ( x20); 3: Thrombosis e microhemorrhages

lesion. Patients return home the same day of treatment or the day after if the treatment was completed in late afternoon, without any medication. Between 1 and 6 weeks from the HIFU treatment, patients underwent conserving surgery associated to sentinel lymphnode biopsy or complete axillary dissection, if the sentinel lymph-node was metastatic. Macroscopic observation of surgical specimen showed the ablated tumour gray-white or gray-yellow in colour and felt ﬁrm on palpation. There was no evidence of haemorrhage in the central region of the treated lesion. Microscopic examination showed tumour cells with typical characteristics of coagulation necrosis induced by thermal energy.

not allow for dye staining in those cases which had shown receptor expression positivity in the pre HIFU treatment immunohistochemical evaluation. Another method to evaluate the effect of the HIFU was tested: 2,3,5-triphenyltetrazoliumchlorid staining (TTC) was used to estimate the capability of treated cells to absorb a viable dyeing at the gross examination. All lesions evaluated with this kind of method showed to be not stained.

Surgical specimen: hemorrhagic ring surrounding the tumour

Haematoxylin & Eosin stain showed that cellular structure looked normal in some cancer cells, without any signs of tumour cell breakdown. The tissues maintained their cytologic staining characteristics giving an appearance similar to viable cells. By using conventional histological techniques, it was difﬁcult to identify whether the treated tumor cells were dead or alive, in this region, under light microscope. Very interesting was the indirect detection of thermal damage in the immuno-histochemical evaluation of oestrogen and progesterone receptors (ER and PR) expression. Protein denaturation indeed, caused by the high temperature, did

This preliminary research, although limited to a small number of patients, but quantitatively comparable to those presents in literature, showed the USgHIFU treatment appears to be feasible and safe to destroy primary early breast cancer, without side effects. Hence, histopathology evaluation performed on surgical specimen after HIFU treatment, conﬁrmed the correct ultrasound guided targeting and release of thermal energy only within the tumour. Based on these premises, for the next year the Breast Division research activity will be directed towards different targets. Technology upgrade was the other main issue within our research activity: the introduction of a new Real-time Variable-Band Linear Array - 13-4 MHz US probe and a new dedicated HIFU transducer have been developed in order to improve the US guidance/monitoring and reduce treatment time, respectively. Ongoing Research The principal purpose arising from our 2009 experience is addressed to conﬁrm the kind of patients would be the best

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suitable for this non-invasive treatment. Over this main issue, ongoing research is focused to continue the upgrade of HIFU technology in order to optimizing the technique and reducing the treatment time. From the anaesthesiology point of view, our results indiImmuno-histochemical evaluation of oestrogen receptors (ER) expression: Left: before HIFU; right after HIFU

cated that local anaesthesia together with MAC is feasible for HIFU procedure and our encouraging results may change our patient’s management in the future. Regarding the histopathology point of view, speciﬁc methods will be improved for assessing cell viability after HIFU through the development of viable colour methods and cell cultures. As a “so called” non-invasive technology, HIFU approach may play a key role in the future management of solid malignancies. The preliminary research activity and application of this new technology in patient care started more than one year ago and was limited to a small number of patients with short-term follow-up, in focusing the feasibility and safety of this new technique. Based on the preliminary results, US imaging-guided focused ultrasound treatment appears to be feasible and safe to ablate primary and metastatic liver tumors, primary and metastatic bone tumors, pancreatic cancers and soft tissue tumours, without signiﬁcant side effects, and can have a role in “multi-modal” management of patients with tumour disease. The research activity is deeply integrated with the daily clinical work and the principal purpose arising from our studies are addressed to follow up all patients to ascertain which patient group would be the best suited to this non-invasive treatment. Over this main issue, ongoing research is focused also on implementation of HIFU technology in order to optimize the patient’s approach technique.

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Robotic Surgery Research Programme

Robot-assisted surgery is the latest evolution of minimally invasive surgery; it has been evolving from simple adjustable arms to support cameras in laparoscopic surgery through to the more sophisticated four-armed machines now available. The da Vinci™ operating robot is a telemanipulation system consisting of a surgical arm cart, a master console and a conventional monitor cart. It acts as remote extensions completely governed by the surgeon and thus are best described as master-slave manipulators. At IEO two of da Vinci™ operating robots (Surgical Intuitive, Inc., Mountain View, CA) are active, the first one since October 2006. To maximize utilization and reduce maintenance costs, they are jointly used by the departments of Urology, Gynecology, AbdominalPelvic, Thoracic, General-Laparoscopic and Head and Neck Surgery. The main technological advantages of this system are realistic 3-D imaging, motion-scaling and tremor filtration, facilitating more precise and accurate endoscopic surgery. It makes difficult and previously inaccessible body areas easier for surgeons to access and may lead to decreased morbidity for patients. Various surgical procedures have proved feasible and safe when performed with the da Vinci™ robot. The advantage of the system is best seen in tiny areas difficult of access and when dissecting delicate, vulnerable anatomical structures, like mesorectum, prostate, uterus, pulmonary lobes or larynx. In the light of our present experience, we regard prostatectomy, hysterectomy, pulmonary resection, adrenalectomy, left colectomy, gastrectomy and total mesorectal excision deep in the pelvis as appropriate for a robotic approach, whereas splenectomy, pancreasectomy and liver resection need further evaluation. The steric vision and the intuitive use of the instruments are of great assistance, although the lack of robotic stapler devices and flexible instruments sometimes hamper full robotic performance. The robotic approach is significantly more expensive than conventional minimally invasive surgery. This extra cost is due to longer operating times, as well as the high cost of the robot itself and higher costs for the robotic

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instruments, which are re-usable ten times only. The time delay may be explained by the learning curve. However, with increasing experience on the part of the entire team (surgeons, scrub nurses, and theatre attendants), the setup time has been markedly reduced in our Institution, and no longer involves any time loss. Cost-effectiveness is a major issue; 2 recent studies comparing robotic procedures with conventional operations showed that although the absolute cost for robotic operations was higher, the major part of the increased cost was attributed to the initial cost of purchasing the robot and yearly maintenance. Both factors are expected to decrease as robotic systems gain more widespread acceptance. Decreasing operative time and hospital stay will also contribute to the cost-effectiveness of robotic surgery. Other drawbacks to robotic surgery include the bulkiness of the robotic equipment currently in use. Lack of tactile and force feedback to the surgeon is another major problem, for which haptics (ie, systems that recreate the “feel” of tissues through force feedback) offers a promising, although as yet unrealized, solution. The patients’ satisfaction following a robotic approach is high. Ready acceptance of the robotic approach may result from the satisfactory cosmetic and symptomatic results, but also from the patients’ impression that they had taken part in the dawn of a new surgical era. In the single Divisions’ Chapters, a complete list of all ongoing studies is presented . Perspectives Robotic surgery was originally developed to render possible a kind of telesurgery bridging thousands of kilometers or even continents. Although the feasibility of this aspect was proven and gained some media attention, it is not the future of robotic surgery. More probable opportunities for robots are image fusion and surgical training. At this stage the superposition of different radiologic imaging systems permits more precise and detailed surgical planning. The da Vinci™ system will implement this technique in the operating room itself by ﬂashing a patient’s

scan images into the virtual three-dimensional view on the console. This will enable the surgeon to more easily detect and identify hidden anatomical structures, and in this way robotic surgery will help to make minimally invasive surgery safer. Another great potential for the da Vinci™ robot probably lies in its impact on surgical training. It will be possible to carry out a particular patient’s complete surgical procedure using his CT scans and robotic virtual-reality training programs. Thus, similar to a pilot on a ﬂight simulator, surgeons in training will perform new operations only after performing them successfully in virtual reality.

development of intracorporeal robotics, the feasibility of which has been demonstrated. At this moment there are no reports to clearly demonstrate the superiority of robotics over conventional laparoscopic surgery. Further research and more prospective randomized trials are needed to better deﬁne the optimal application of this new technology in gastrointestinal oncologic surgery. The challenge for today’s robotic surgeons is to advance the system through clinical research in such a way that it becomes suitable and indispensable for future routine applications.

Conclusions With the da Vinci surgical robot surgery regains two fundamental tools of surgical procedures: intuitive control over the surgical instruments and steric perception of the operative ﬁeld. Only several centers are currently using surgical robots and publishing data. In gastrointestinal surgery, robotic surgery is applied to a wide range of procedures, but is still in its infancy. Most studies reported that robotic gastrointestinal surgery is feasible and safe, provides improved dexterity, better visualization, reduced fatigue and high levels of precision when compared to conventional laparoscopic surgery. In a relatively short time, robotic procedures spanning the whole spectrum of surgery have been successfully executed. Initial results show that mortality, morbidity, and hospital stay compare favorably to conventional laparoscopic operations. However, only a limited number of randomized, prospective studies that compare outcomes of robotic techniques with conventional methods exist. While current robotic systems have considerable advantages over conventional laparoscopic techniques, they are not without limitations. Robotics main drawbacks in surgical practice are the absence of force feedback and extremely high costs. Miniaturization of robotic components and systems is feasible and necessary to allow minimally invasive techniques to reach full potential. The ultimate extrapolation of this progress is the

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Interventional Radiology

Clinical needs for local tumor control, by means of minimally invasive approach more than classical invasive, risky and expensive surgical ones, represents the main reason for the very fast developing of new hyper-technological branches in Oncology. Due to the high specialization required for most of the interventional procedures in Oncology, the appellative of “Interventional Oncology” has been proposed very recently to define this field. Common elements of different approaches in this branch is image guidance for very precise treatments and the low invasiveness compared with more classical options. For academic purposes Interventional Radiology may be divided in two different main fields: Vascular treatments and Percutaneous procedures. The use of blood flow for delivering drugs and/or particles directly and selectively to the tumor is the aim of all Vascular treatments in Interventional Radiology and they are mainly oriented for liver tumor control. Percutaneous procedures include all image-guided tumor ablative techniques which required percutaneous insertion of dedicated devices (such as needle for radiofrequency). Moreover, a totally new locoregional therapy for tumor ablation has been very recently added to the IR armamentarium: High Intensity Focused Ultrasound (HIFU-see dedicated chapter).

Vascular Treatments Introducing the multimodal approach for hepatic lesions Hepatic lesions represent a main challenge in clinical oncology, both for primary and metastatic tumours. This is mainly due to the crucial role this organ represents for the prognosis. In hepatocellular carcinoma, the simultaneous presence of cirrhosis and tumour lesions in the majority of patients, limits the indication for an aggressive local approach, because of the high risk of post-treatment liver failure. Moreover the high rate of new nodule development after any local treatment (>80% after 4 years), plays a key role in the decision making on treatment strategy.

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Regarding liver metastases, local treatments such as surgery for liver metastases (mainly from colorectal cancer) after systemic chemotherapy seems to induce better results than chemotherapy alone. Minimally invasive local techniques are therefore being continuously developed in order to supplement the surgical resection. Super-selective bland arterial embolization with microparticles. Hepatocellular Carcinoma: The intra-arterial treatment using transarterial chemoembolization (TACE) and transarterial embolization (TAE) is considered a palliative therapy for multifocal HCC not suitable for surgical resection or percutaneous ablation therapies, but the efficacy of intra-arterial treatment in patient with single HCC is not well defined (7,8,9). However, it is not clear whether embolization alone gives the same survival advantage (5) nor whether specific patient characteristics affect outcome or any particular technique in performing transarterial therapy is better than any other. The aim of this study, is to assess feasibility and local response in patients affected by unresectable hepatocellular carcinoma, treated with TAE by using very small sized fine-calibrated microparticles (40 and 100 microns). Between October 2007 and september 2009 63 HCC patients underwent 100 TAE, and a total of 87 target lesions were embolized with a super-selective technique. Embolization was preformed by using very small size finecalibrated micro-particles of 40 μ and/or 100 μ in diameter. These represent a new spherical embolic agent designed with a hydrogel core of polymethyl-methacrylate (PMMA) combined with a coating shell of Polyzene-F i.e. Embozene ™ In all cases liver perfusion scintigraphy was performed by injecting Tc-99 labelled albumin macro-aggregates (MAA) via the main hepatic artery, just before treatment. That way a reasonable mimicking diffusion imaging of what embozene were supposed to do was obtained: in four patients a 3-5 % amount of pulmonary shunting was observed and embolization has been performed by upsiz-

A)HCC relapse after left lobectomy B)CT 1 month after TAE C)CT scan 6 months after TAE D) CT scan 15 months after TAE

ing microparticles to 100 m, in order to reduce the risk of pulmonary embolism. Twenty-four hours after treatment, upper abdomen MDCT was performed in order to assess the early local result after embolization. Median follow-up for all 63 patients was 26 months (range, 1-38 months). Of the 63 patients, 4 have died: one died within 24 hours after intervention and three patient died for liver progressive disease. Up to now 29 out of 63 patients have at least one year follow up with an overall survival rate of 96%. Local outcomes on target lesions at 1 year follow-up in these 29 patients are: 6 complete response (CR ), 5 partial response (PR ), 7 stable disease (SD ), and 9 progressive disease (PD ) for new lesions (i.e. new nodules) while 2 PD on treated lesions. Bland embolization is an effective therapeutic modality in the treatment of unresectable HCC. In our initial experience with Embozene ™ 40 and 100 μm we observed a very good handiness in embolization, obtaining good results in lesion and disease control, at a median follow-up of 12 month. Microparticles 40 and 100 μm in diameter are very effective in local response, with good clinical outcomes, but patients must be selected very carefully in order to reduce major complications Metastases: The aim of this new ﬁeld is to assess the feasibility and efﬁcacy in metastatic lesion control with a new kind of coated small particles: 40 and 100 microns EMBOZENE® Between October 2007 and september 2009 42 patients with liver metastatic disease underwent TAE: 10 CRC, 17 NET, 5 breast cancer, and 10 from other malignancies. Technique used was the same as in primary liver cancer for vascular access and microspheres injection. The intent was not curative only in patients with NET mets because of radionuclide therapy. The real purpose in fact was to obtain a “debulking” effect. At the moment response assessment is feasible in 38 patients following RECIST criteria: 12 PD, 15 SD, 8 PR and 3 CR with a mean follow-up of 13 month. 3 pt were lost at follow-up. Yttrium-90 internal radiation therapy for hepatic malignancy Surgical resection is the only potentially curative strategy in the treatment of patients with hepatic malignancy. Unfortunately, due to advanced stage, underlying liver

A) liver NET metastases before treatment B) 3 months after TAE with 40 microns

disease, or medical co-morbidities, most patients are inoperable at this time of presentation. As a result, various loco-regional therapies have emerged for otherwise unresectable hepatic tumors. Radioembolization (i.e. intraarterial administration of 90Y microspheres-SIR-Spheres®) is a therapeutic option that allows preferential delivery of radiations into the tumor without signiﬁcant liver toxicity. SIR-Spheres® (SIRTEX Medical, Australia) is a resin-based microsphere, of 20-40 μm-diameter, and a typical dose of 3-8 milion per injection. 90Y is a pure -emitting radioisotope, it has a high average energy (0.936 MeV), limited tissue penetration (mean 2.5 mm; max 11 mm), and short half-life (64 h), making it an ideal transarterial liver-directed agent. 90Y is selectively injected into the whole liver parenchyma, or in one liver lobe, via the main lobar hepatic arterial branches. Prior to radioembolization, detailed imaging (CT, PET/CT)

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A) Basal CT-PET performed before SIR-Spheres® therapy in patient with metastases from breast cancer shows some lesions into the hepatic right lobe; B) CT-PET 2 years after treatment conﬁrmed no FDG uptake. CT-PET is completely negative.

and therapy planning must be completed. All patients that are considered for 90Y internal radiation therapy must undergo a preliminary hepatic angiography in order to assess local vascular anatomy. At the end of angiography, simulation of treatment is performed by injecting 99mTcmacroaggregated albumin, in all hepatic arterial branches, and nuclear medicine scan is carried out (MAA scintigraphy) . This work-up aims to determine the presence of hepato-pulmonary shunting, and the amount of the dose to administer to the patient. IEO experience. From October 2005 up to now, 29 patients with liver metastases (from colorectal cancer in 11 pts, breast cancer in 10 pts, uterine cancer in 3 pts, other can-

cers in 5 pts) have been treated with SIR-Spheres® in our Institute. Two patients screened for SIRT, were excluded at preparation time because of mismatch between MAA scintigraphy and CT/PET scan. Once vascular hepatic anatomy was assessed, for each patient a dosimetric evaluation, based on 99mTc-MAA images (WB and SPECT, 30 min p.i.) and CT scans, has been performed in order to estimate the activity to be safely injected. Endovascular embolization of extrahepatic arteries (eg. GDA, right GA, and other extra-hepatic arteries originating from liver arteries) were performed with endovascular metallic coils in order to obtain just one vessel feeding the whole liver parenchyma. Then Sir-Spheres ® were gently administered via the

Multiple liver metastases from larynx cancer. A) white arrows show one of the huge liver lesions at pre radioembolization CT. B) CT after 11 months show the dramatic shrinkage of the same lesion (yellow arrows)

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hepatic arteries. In most cases, patients well tolerated the procedure and required only brief observation. Side effects were usually limited to nausea, vomiting, abdominal pain and transient elevations in liver enzymes. In 3 pts a reversible gastroduodenal ulceration was documented by endoscopy. Concerning the efficacy profile, revaluation CT and PET/CT (6, 12 and 18 weeks after treatment) documented objective response in 83% of patients (fig. 2b), with complete response in 3 of them. At 24 months FU only one patients is still with complete response (see the picture). Relapse of disease was observed in 10 patients, while in 5 patients an extraheaptic progression at disease was registered. In the remaining patients a partial response to treatment or stable disease was evaluated via CT or PET scan. In our experience we observed that the optimal dose to normal liver is up to 40-50 Gy, but the optimal response to the treatment has been achieved in those lesions with high T/NT ratio at MAA scintigraphy. A regionally administration of microparticles is preferable, that means to deliver microparticles just one liver lobe per time, instead whole liver parenchyma, in order to be more safe and to achieve a major radiation dose. SIRT can be considered as a 2nd or 3rd line therapy for liver secondary lesions, but it must be considered only in a multi-disciplinary and multitreatment behaviour. Percutaneous Procedures - Percutaneous thermal ablation Hepatocellular carcinoma and colorectal liver metastases are the two most common malignant liver tumors. While surgical resection remains the gold standard of therapy, only a few patients are suitable candidates for curative surgical resection, because of the presence of liver malig-

nancy in unresectable locations, the number and anatomic distribution of tumor lesions, or the presence of extrahepatic disease, poor liver function or medical comorbidities. From experiences reported in literature, despite the limitations of the data, reasonable safety of the procedure has been established, with mortality and morbidity rates in the largest series of 0.2% and 1.7% respectively. The major complications described are peritoneal bleeding, hepatic abscesses, intestinal perforation, large biloma, acute cholecystitis. Tumor seeding is probably over-emphatisized and can be avoid by a hot withdrawal of the electrode. Candidate patients with HCC and cirrhosis have to be in Child’s class A or B; tumour has to be single or, if multiple, with no more than three nodules, without vein thrombosis or extra-hepatic spread. The dimensions of the nodules should preferably be below 3–3.5 cm. By using combination therapy (hepatic artery occlusion and RFA; RFA and Pringle manoeuvre during surgery) larger tumours can be treated. RFA is considered a safe bridge to liver transplantation in several recent papers. Pretreatment work-up includes ultrasound (US) examination, unenhanced and contrast enhanced CT or magnetic resonance (MR); serum tumor markers, namely alfaphetoprotein (AFP) and carcinoembryonary antigen (CEA) dosage. Recently FDG-PET examination has been introduced in staging liver metastases. For HCC greater than 2 cm, two positive imaging techniques or one positive imaging technique and AF values higher than 400 ng/ml sufﬁce for diagnosis and no biopsy is required. Tumors close to the gall bladder or subcapsular adjacent to hollow organs should be preferably treated by a laparoscopic or open approach; RFA of a lesion close to the hilum plate puts the patient at high risk of biliary injury. In our institute

Liver metastases from CRC in segment 7 (arrow): A) CT before treatment; B) post-treatment CT shows a low-density area (=necrosis) where thermal ablation has been performed

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Percutaneous Thermal Ablation Of Hepatic Lesions survival rates: 93 pts Interventional Radiology Unit

Tumour Histology Mean F.U. (range) n°

2y 3y

HCC

42 (3-94) mo.

100 80 71

METS

46 (2-98) mo.

100 76 45

Table data about our own experience about Percutaneous thermal ablation of hepatic lesions

to RFA. Nine-teen patients (9 males; mean age 66.5 years) with liver lesions (8 metastases from CRC, 2 HCC, 2 CCC, 3 breast mets and 4 mets from different origin) underwent the combined approach (TAE and RFA) in the same session. TAE was performed with 40 μm Embozene™ injected within segmental or sub-segmental arteries supplying the lesion, until cessation of blood ﬂow. RFA was then performed under US/CT guidance with 3.5 to 5 cm LeeVeen needle, according to the lesion size (mean diameter 38.2 mm; range 25-70 mm). Technical success was 100%. We obtained an area of coagulation necrosis larger than

CHARACTERISTICS OF THE LESIONS we treated 108 patients, 76 with liver metastases (follow up 2 – 98 months, median 46) and 32 with HCC (follow up 2-94 months, median 42); RF was performed with ultrasound and/or CT guidance under conscious sedation and requires 3 days of hospitalization. In our experience we had a very low percentage of complications, less than 1%, and no mortality. To assess the results a contrastenhanced CT is performed after one month: coagulation necrosis appears as a non-enhancing low density area in both arterial and portal phases; on MR T2 hypointense images and loss of enhancement on gadolinium enhanced MR correspond to complete necrosis. FDG-PET allows for detection of pathological increase of glucose metabolism, as it happens in solid tumor. If a residual non-ablated tumor or tumor re-growth is detected a re-treatment with RF can be performed. Liver bland embolization associated with radiofrequency thermal ablation Tumor size, site and morphology may affect the local results of liver radiofrequency ablation (RFA). We are evaluating feasibility, safety and local effectiveness of a single session combined approach for treating difﬁcult liver lesions with bland embolization (TAE) immediately prior

Hystology

FU Lenght (months)

Tumor Size (mm)

Size of inducted

CRC CRC HCC CRC

28 14 16 13

30 25 30 40

60 55 79 80

CCC

CRC HCC CCC BREAST CRC

44 29 11 11 11

35 35 70 27 32

68 50 85 86 93

CRC

LUNG CRC BREAST BREAST UTERUS

7 6 4 4 2

34 39 19 30 37

79 62 45 83 92

UTERUS

PD (New hepatic mets)

OVARIC

108

a) residual liver metastases after chemotherapy treated with percutaneous RFA b) result after 1 year

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EASL CR CR CR CR PD (New hepatic mets) CR CR CR CR CR PD (local relapse) CR CR CR CR CR

A-B) Cholangiocarcinoma of segment IV at CT and PET-CT examination C) 24 hours after TAE+RFA D) result at CT scan 1 month after E) PET-CT scan 2 month after

needle size (mean diameter 73 mm; range 44-108 mm). All lesions were completely ablated with a safe margin; 3 recurrences occurred: only in one case in the same site of treated lesion (see table). Follow up range is from 1 to 28 month: 3 PD and 15 SD with EASL criteria evaluation Combined approach for treating difﬁcult liver lesions seems to be feasible and safe. TAE with micro-particles performed immediately before RFA allows for a larger area of coagulation necrosis, improving the volume of ablation. Even if long-term clinical results are needed, this technique could be a useful therapeutic option for patient with large or difﬁcult liver lesions appropriate for localized treatment. Radiofrequency thermal ablation in RCC With the increasing number of patients with renal cell carcinoma detected at an early stage, nephron-sparing approaches are becoming more popular. More recently as an alternative to partial nephrectomy radiofrequency thermal ablation (RFA) has been introduced to treat small renal cell carcinoma, with the rationale of in situ tumor destruction to further reduce morbidity and invasiveness. This technique, destroying tumor tissue by heating, may be an appropriate alternative in patients with RCC detected

in a solitary kidney or with marginal renal function in whom surgical resection would likely result in the need of dialysis. RFA should be proposed to patients with a disease limited to the kidney (ie T1N0M0), dimension of the lesion less or equal to 4 cm and with a reasonable life expectancy (more than 12 months), in whom the risk/beneﬁt ratio is believed to be favorable. Whenever possible biopsy should be performed before ablation, to be sure that the lesion ablated is RCC and not a benign lesion and to determine RCC subtype for systemic chemiotherapy in case of future metastases. Up to now we treated 13 patients with RCC (9 male): dimension lesion were from 11 to 30 mm. We had only 3 minor complications: one subcapsular hematoma that didn’t require any treatment and solved spontaneously, one skin-burn at insertional site and a little subcapsular urinoma. We obtained a completely necrosis of the tumor in 12/13 cases (100% survival rate after 11.15 months – median 5 months). Radiofrequency thermal ablation of lung tumors Percutaneous ablation with use of radiofrequency (RFA) is a minimally invasive technique for local treatment and

A) Small RCC (arrowhead) very close o the ureter (yellow arrow) at the pre-RFA CT; B) CT at one month after treatment shows complete necrosis (white arrow).

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its use is well established in patients with malignant liver tumors, primary or metastatic, who are not candidates for surgical resection. It induces temperature changes by using high-frequency alternating current applied via electrodes placed within the tissue capable to create large areas of necrosis. Several studies demonstrated that it is an effective and safe technique for treatment tumors with mortality and morbidity rates in the largest series of 0.2% and 1.7% respectively. Recently RFA has been proposed to treat pulmonary malignancy. In fact this technique can be used in patient that are not suitable for surgical resection for poor pulmonary reserve, comorbid cardiopulmonary disease, number and anatomic distribution of tumor lesions or in patients that refuse to undergo surgery. In patients with metastatic pulmonary disease, especially from colorectal carcinoma, surgical resection can achieve 5-yeatrs survival rates of 20-40% but it is indicate in a relatively small percentage of carefully selected patients. If surgery is not indicated these patients would normally receive palliative systemic chemotherapy: RFA can be proposed to obtain a better long-term survival. Also in primary lung tumors RFA can be proposed in patients with unresectable lesions. In fact inpatients with recurrent lung carcinoma or with coexistent chronic obstructive broncho-pneumopathy or other associated disease the only chance is chemotherapy or radiation therapy alone or associated. In these cases percutaneous RFA represents an effective and safe minimally invasive treatment. Are candidate to RFA patients with histological or clinical proof of lung malignancy, maximum diameter of the tumor equal or less than 4 cm. Also for primary tumors haemostasis disorders, location of the lesion adjacent to major pulmonary vessels and to major bronchi are exclusion criteria.. The procedure is performed under CT guidance, with the

patient under general anesthesia and requires 3 days of hospitalization; the RFA needle is positioned in the center of the lesion and than radiofrequency energy applied. The major complications described are pneumothorax, pleural effusion and haematoma of the chest wall. In our institute we treated 19 patients, 15 with colorectal carcinoma metastases (follow up 1 – 36 months, median 13) and 4 with primary lung tumors (follow up 8-31 months, median 20). In our experience we had no complications and no mortality. To assess the results a contrast-enhanced CT and a PET scan is performed after one, three and every six months; the main criteria for complete response are reduction in diameter of the lesion, reduction of density in ct exams and disappearance of 18FDG uptake in PET scan. If a residual non-ablated tumor or tumor re-growth is detected a re-treatment with RFA can be performed.

a) CT shows lung metastasis within the right inferior lobe; b) RFA needle within the nodule, during treatment; c) CT result one year after treatment: the lesion is replaced by scar tissue

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Tumour Imaging by Nuclear Medicine

Lymphoscintigraphy and sentinel node biopsy in oral/ oropharyngeal squamous cell carcinoma (OSCC) Preparation of guideline was carried out by a multidisciplinary surgical/nuclear medicine/pathology expert panel under the joint auspices of the European Association of Nuclear Medicine (EANM) Oncology Committee and the Sentinel European Node Trial (SENT) Committee. Involvement of the cervical lymph nodes is the most important prognostic factor for patients with oral/oropharyngeal squamous cell carcinoma (OSCC), and the decision of whether to electively treat patients with clinically negative necks remains a controversial topic. Sentinel node biopsy (SNB) provides a minimally invasive method for determining the disease status of the cervical node basin, without the need for a formal neck dissection. This technique potentially improves the accuracy of histologic nodal staging and avoids overtreating three-quarters of this patient population, minimizing associated morbidity. The technique has been validated for patients with OSCC, and larger-scale studies are in progress to determine its exact role in the management of this patient population. This document is designed to outline the current best practice guidelines for the provision of SNB in patients with earlystage OSCC, and to provide a framework for the currently evolving recommendations for its use. Eur J Nucl Med Mol Imaging. 2009 Nov;36(11):1915-36. Ann Surg Oncol. 2009 Nov;16(11):3190-210 Lymphoscintigraphy and sentinel node biopsy in melanoma EANM-EORTC general recommendations for sentinel node diagnostics in melanoma have been published. The accurate diagnosis of a sentinel node in melanoma includes a sequence of procedures from different medical specialities (nuclear medicine, surgery, oncology, and pathology). If speciﬁc recommendations given cannot be based on evidence from original, scientiﬁc studies, referral is given to "general consensus" and similar expressions. The recommendations are designed to assist in the practice of referral to, performance, interpretation and reporting of all steps of the sentinel node procedure in the

248 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

hope of setting state-of-the-art standards for good-quality evaluation of possible spread to the lymphatic system in intermediate-to-high risk melanoma without clinical signs of dissemination. Eur J Nucl Med Mol Imaging. 2009 Oct;36(10):1713-42. Cutaneous melanoma in the elderly The aim of this study was to analyze the difficulties in diagnosing and treating elderly patients with cutaneous melanoma. It focused on the main causes for late diagnosis and relatively poor prognosis in these patients. Early detection of melanoma is vital to reduce mortality in these patients and surgery is often curative. Adequate treatment of elderly patients with melanoma requires knowledge of the clinical features and histopathology of the disease, and the therapeutic options. We also examined the main surgical procedures for primary melanoma and regional lymph node staging, and the curative and palliative procedures indicated for those elderly patients with advanced disease. It is expected that several molecular genetic factors will soon provide further prognostic information of possible benefit for elderly patients with melanoma. Melanoma Res. 2009 Jun;19(3):125-34. Sentinel node biopsy in patients with previous breast aesthetic surgery. Sentinel lymph node biopsy (SLNB) is the standard method for axillary staging of early breast cancer. Recent studies have focused on questioning the initial contraindication to the technique. We studied 70 patients with previous breast aesthetic surgery submitted to SLNB. Fifty had a previous breast augmentation and 20 had breast reduction mammoplasty. All patients underwent lymphoscintigraphy with 99Tc according to our standard technique and sentinel node was identified in all cases. The sentinel node identification rate was 100%. SLN was positive in 23 patients (32%); there were 18 cases with macrometastasis and 7 cases with micrometastasis. After median follow-up of 19 months, no axillary recurrences have been observed. Our conclusion is that past history of breast augmentation

or reduction is not a contraindication to SLNB technique. Ann Surg Oncol. 2009 Apr;16(4):989-92. Peritoneal carcinomatosis The diagnosis of peritoneal carcinomatosis secondary to ovarian cancer is a real challenge in the cancer imaging field. In this retrospective study, we evaluate the accuracy of Single Detector Computed Tomography (SDCT), Multi Detector Computed Tomography (MDCT), and Positron Emission Tomography-Computed Tomography with F18fluorodeoxyglucose ([18F]FDG-PET/CT) in the diagnosis of peritoneal seeding and we evaluate the possible applications of MDCT to predict the complete surgical removal of the peritoneal deposits. A total of 228 scans (91 SDCT, 89 MDCT, and 48 [18F]FDG-PET/CT of patients with peritoneal carcinomatosis secondary to ovarian cancer proved at laparoscopy and confirmed by histopathology were retrospectively reviewed by two independent groups of Radiologists and Nuclear Medicine Physicians for the evaluation of ascites, peritoneal nodules, and omental cake signs. MDCT showed 81% of true positives, SDCT 72.5%, and [(18)F]FDG-PET/CT 77%. False negatives were 19% for MDCT, 27.5% for SDCT, and 23% for [(18)F]FDG-PET/ CT. From our results, we concluded that MDCT is the technique of choice in the diagnosis of peritoneal seeding, while [18F]FDG-PET/CT, though showing similar accuracy, remains the most accurate technique for monitoring therapeutic response and disease recurrence. MDCT could play an important role due to its ability to predict the possibility of complete surgical removal of disease thus influencing the treatment plan aimed to improve quality of life. Abdom Imaging 2009 Ongoing studies Role of FDG-PET/CT in locally advanced rectal cancer along neoadjuvant chemo-radiotherapy To assess the impact of FDG-PET/CT in locally advanced rectal cancer patients (LARC) for staging purpose and in predicting therapy response to neo-adjuvant chemoradiotherapy. We will enroll a series of consecutive patients with histologically rectal adenocarcinoma. At baseline all patients will be submitted to total body CT with contrastenhancement (ceCT) and/or magnetic resonance of the pelvis, FDG-PET/CT (FDG-PET/CT1), eco-endoscopy and colonoscopy. Along neo-adjuvant therapy patients will be submitted to FDG-PET/CT at 40 Gy (FDG-PET/CT2) and at the end of treatment (FDG-PET/CT3). Patients will receive capecitabine concomitant to radiotherapy on the pelvis (50.4 Gy/ 28 fractions), followed by two cycles of capecitabine until 2 weeks before surgery. FDG-PET/CT images will be evaluated visually by a binomial scale (positive/ negative) and semi-quantitatively by the measurement of

the maximal standardized uptake value (SUVmax) on the primary tumour. We will also calculate the percentage reduction of SUV max along the three PET scan (PET1-2, PET2-3, PET1-3). Surgical specimen of the radical surgery will be expressed by the TNM system and the regression index by Dworak. Role of FDG-PET/CT in NHL DLBC Aim of this prospective multicentric study is to analyze the predictive value of FDG-PET/CT along chemoterapeutic regimen (R-CHOP14) in diffuse large B cells non-Hodgkin lymphoma (DLBC). All patients will be submitted to baseline evaluation through FDG-PET/CT and contrast-enhancement CT (ceCT); the enrolment will be defined in relation to the presence of measurable disease at the baseline FDG-PET/CT. After the enrolment FDG-PET/CT and ceCT will be performed after two cycles of chemotherapy, after four cycles of chemotherapy and at the end of chemotherapy. The enrolment of the patients is ongoing. Role of PET in the selection of patients with breast cancer candidate to Sentinel Node Biopsy after neoadjuvant therapy The main objective of this study is to determine the role of [18F]-2-fluoro-2-deoxy-D-glucose – positron emission tomography (FDG-PET) in the selection of patients with breast cancer candidate to sentinel node biopsy after neoadjuvant therapy. The study foresees enrollment of patients with primary breast cancer clinically classified as cT2, cT3 or cT4a-c cN0-N2 or cN3 and M0 and with a baseline FDG-PET scan positive both in the site of primary tumor and axillary lymph nodes. According to local guidelines and/or protocols appropriate primary therapy and a second PET scan at the end of treatments are given to patients. In case of axillary FDG-PET uptake at the second PET scan, lymph node dissection (ALND) is performed. Should the second FDG-PET scan be negative for axilla involvement, sentinel node biopsy (SNB) will be performed in order to evaluate axillary lymph nodes status. Preliminary results on 44 patients demonstrate a relatively high positive predictive value of FDG-PET for detection of axillary lymph nodes metastases after neoadjuvant therapy. These data seem to suggest a role of FDG-PET in selecting patients who, after neoadjuvant therapy, are candidate to axillary lymph nodes dissection, avoiding SNB. However, this issue requires a confirmation on a larger series of patients and the enrollment is still ongoing.

249 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Radioimmunotherapy

The concept of targeting radionuclides to tumours using radiolabeled monoclonal antibodies (MoAbs) against tumour associated antigens (Radioimmunotherapy – RIT) was proposed more than a century ago. With the development of the hybridoma technology and the availability of MoAbs against tumour-associated antigens, this concept was investigated in animal models and in cancer patients. RIT combines the specificity of a monoclonal antibody in targeting the cancer antigen with the activity of a radioactive molecule. Despite more than 30 years since the introduction of radioimmunotherapy, this technology has attracted interest of oncologists only in the last few years. This interest can be attributed to the availability of two clinically approved RIT drugs and their remarkable therapeutic efficacy in the management of non-Hodgkin’s lymphoma (NHL). The success of RIT in B-cell NHL therapy is, at least in part, related to the characteristics of the disease, such as the radiosensitivity and the abundance of the target antigens on its surface, and the deliver of a cytotoxic radiation to the tumour by the presence of a suitable therapeutic radionuclide. So, the conjugation of a monoclonal antibody with a radioisotope represents a valid option to strengthen the efficacy of MoAbs, supplying doses of radioactivity to specific targets. Yttrium-90 (90Y) ibritumomab-tiuxetan (Zevalin®) is a beta-emitter, radio-conjugated, anti-CD20 antibody which includes ibritumomab, a murine parent of the humanized anti-CD20 MoAb Rituximab, conjugated by tiuxetan to 90Y. Zevalin® has been already demonstrated to be safe and effective, as a complementary approach, in the treatment of follicular and diffuse large B-cell NHLs, as well as in cases resistant or refractory to Rituximab. We evaluated the efficacy and safety of a single dose of Zevalin® in elderly patients in first relapsed or primary refractory diffuse large B cell Non-Hodgkin Lymphoma, ineligible for stem-cell transplantation, and demonstrated high response rates to Zevalin® in this setting of patients, in a prospective multicenter non randomized

250 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

phase II trial. The overall response rate (ORR) was more than 50% with complete response rates of more than 25%. When administered at the conventional prescribed activity of 14.8 MBq/kg (0.4 mCi/kg), the red marrow is the critical organ and toxicity is primarily haematological but reversible. The activity of 1.18 GBq (32 mCi) has been identified as the maximum tolerable, in spite of the single patient absorbed dose to the red marrow. In order to increase its efficacy, Zevalin® has been investigated at myeloablative activities: a high dose (HD) trial based on 90Y-ibritumomab-tiuxetan activities up to 55.5 MBq/kg, followed by autologous stem cell transplantation (ASCT) is ongoing in our Institution for the treatment of resistant/refractory follicular B-cell non Hodgkin’s Lymphoma. To date, 39 patients affected by resistant/refractory NHL have been enrolled in the trial. In this trial, the patients’ recruitment is based on patient specific dosimetry, in order to evaluate safety and toxicity and confirm the justification of such an aggressive therapy. Before RIT all patients underwent dosimetry, and therapy was administered only if a 20-Gy limit dose to normal organs (except red marrow) was guaranteed. The dosimetric analysis has highlighted the need of deepening the red marrow absorbed dose, as its evaluation by the standard method applied - based on the blood curve – might underestimate the dose in these patients- showing spine marrow uptake. Alternative methods based on imaging and bone aspirates have indicated higher values with a factor of 2–2.5 and have been applied to assess also safe time for transplantation. Studies are ongoing on this matter. Two of the patients monitored for this therapy showed atypical pharmacokinetics and biodistribution, with prevalent high liver uptake. In both patients, liver absorbed doses higher than 50 Gy would have resulted with administration of the prescribed activity; following these dosimetric results, both patients did not receive the planned therapy because the treatments would have been unjustified and with possible serious adverse effects. The importance of highlighting these clinical cases en-

ables to enrich an evidence-based discussion about drug use and development in clinical practice, especially for protocols based on the administration of high activities of radiopharmaceuticals. Another field of investigation has been the optimization of the radiolabelling and quality control of Zevalin® at high activities, to simplify the procedure and to decrease the finger radiation exposure of the operator. Studying the parameters of the reactions, it has been possible to reduce the steps of preparation and to improve the handling of high activity samples, maintaining the quality and the safety of the radiopharmaceutical. In another study, we considered marginal-zone lymphomas (MZL), a subgroup of disease in which this treatment could offer an alternative option when they relapse or do not respond to conventional treatment. Seventeen patients were enrolled in a prospective, single-arm, openlabel, pilot phase II trial of RIT with Zevalin®. Seven out of 17 patients had Helicobacter Pylori-negative gastric MALT (mucosa-associated lymphoid tissue) NHL. The remaining 10 patients had non gastric extranodal MZL. At time of treatment 10 out of 17 patients had disseminated disease (stage III/IV); bone marrow biopsy showed disease localization in 4 out of 17 patients. Median number of previous therapies was 2, including prior chemotherapy, Rituximab, and radiotherapy. Patients received standard activities of 0.4 mCi/Kg, after pre-loading with Rituximab. The therapy was well tolerated without acute and subacute side effects. G3 and G4 neutropenia and thrombocytopenia in 10 out 17 patients, with transfusional support in 2 patients and infections in 3 patients were observed. Twelve out 17 patients obtained a complete response (71%); partial response was observed in 2 out 17 patients (12%). Three patients (18%) obtained a stabilization of disease; one of them progressed after 4 months from RIT. Complete responses are still ongoing (35 months). The high observed overall response and complete response rates indicate that Zevalin® in MZL is efficient. If these preliminary results will be confirmed in larger number of patients, one single administration

of 90Y-Ibritumomab Tiuxetan delivered at conventional activity (0,4 mCi/kg) could be considered a possible alternative option in the treatment of such an indolent disease. During 2008, we participated in a phase I trial using an iodine-131 labelled antibody fragment raised against the EDB fragment of ﬁbronectin in patients with advanced solid and haematological malignancies. Biodistribution and dosimetric studies showed a rapid pharmacokinetics, especially as compared with other radiolabelled MoABs, indicating low risk of haematological toxicity For therapeutic activity of 3.7 GBq, median absorbed doses (Gy) were: 3.70 (kidneys), 3.0 (liver), 2.0 (lungs), 4.20 (testes), 0.85 (red marrow), 12.2 Gy (tumor lesions). The favourable dosimetry results indicated the feasibility of therapeutic protocols with further increased activities (> 3.7 GBq).

251 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Publications, Clinical Research Projects, Seminars and IEO Foundation

Full Papers 2009

AUTHORS

TITLE

JOURNAL

VOLUME

PAGE

I.F.

ABBATE F., BACIGALUPO L, LATRONICO A., TRENTIN C., PENCO S., MENNA S., VIALE G., CASSANO E., BELLOMI M.

Ultrasound-guided vacuum assisted breast biopsy in the assessment of C3 breast lesions by ultrasound-guided ﬁne needle aspiration cytology: results and costs in comparison with surgery.

BREAST

73 - 77

2,155

AGAZZI A., SAMMASSIMO S., LASZLO’ D., LIPTROTT S. J., CASCIO R., ALIETTI A., RABASCIO C., MANCUSO P., PRUNERI G., MARTINELLI G.

Is there a role for ‘modiﬁed VAD’ in the treatment of multiple myeloma?

ECANCERMEDICALSCIENCE

AGNELLI G, GUSSONI G, BIANCHINI C, VERSO M, MANDALA’ M, CAVANNA L, BARNI S, LABIANCA R, BUZZI F, SCAMBIA G, PASSALACQUA R, RICCI S, GASPARINI G, LORUSSO V, BONIZZONI E, TONATO M, ON BEHALF OF PROTECHT INVESTIGATORS, DE BRAUD F.

Nadroparin for the prevention of thromboembolic events in ambulatory patients with metastatic or locally advanced solid cancer receiving chemotherapy: a randomised, placebo-controlled, double-blind study.

LANCET ONCOL

943 - 949

13,283

ALKUREISHI L W, BURAK Z, ALVAREZ J A, BALLINGER J, BILDE A, BRITTEN A J, CALABRESE L., CHIESA C, CHITI A, DE BREE R, GRAY H W, HUNTER K, KOVACS A F, LASSMANN M, LEEMANS C R, MAMELLE G, MC GURK M, MORTENSEN J, POLI T, SHOAIB T, SLOAN P, SORENSEN J A, STOECKLI S J, THOMSEN J B, TRIFIRO’ G., WERNER J, ROSS G L, THE EUROPEAN ASSOCIATION OF NUCLEAR MEDICINE (EANM) ONCOLOGY COMMITTEE, EUROPEAN SENTINEL NODE BIOPSY TRIAL (SENT) COMMITTEE

Joint Practice Guidelines for Radionuclide Lymphoscintigraphy for Sentinel Node Localization in Oral/Oropharyngeal Squamous Cell Carcinoma.

ANN SURG ONCOL

3190 3210

3,898

ALKUREISHI L W T, BURAK Z, ALVAREZ J A, BALLINGER J, BILDE A, BRITTEN A J, CALABRESE L., CHIESA C, CHITI A, DE BREE R, GRAY H W, HUNTER K, KOVACS A F, LASSMANN M, LEEMANS C R, MAMELLE G, MCGURK M, MORTENSEN J, POLI T, SHOAIB T, SLOAN P, SORENSEN J A, STOECKLI S J, THOMSEN J B, TRIFIRO’ G., WERNER J, ROSS G L, THE EUROPEAN ASSOCIATION OF NUCLEAR MEDICINE (EANM) ONCOLOGY COMMITTEE, EUROPEAN SENTINEL NODE BIOPSY TRIAL (SENT) COMMITTEE

Joint Practice Guidelines for Radionuclide Lymphoscintigraphy for Sentinel Node Localization in Oral/Oropharyngeal Squamous Cell Carcinoma.

EUR J NUCL MED MOL I

1915 - 1936

4,532

ALTUCCI L, MINUCCI S.

Epigenetic therapies in haematological malignancies: searching for true targets.

EUR J CANCER

1137 - 1145

4,475

AMABILE G, D’ALISE AM, IOVINO M, JONES P, SANTAGUIDA S., MUSACCHIO A., TAYLOR S, CORTESE R

The Aurora B kinase activity is required for the maintenance of the differentiated state of murine myoblasts.

CELL DEATH DIFFER

321 - 330

7,548

AMATORI S, PAPALINI F, LAZZARINI R, DONATI B, BAGALONI I, RIPPO MR, PROCOPIO A, PELICCI P. G., CATALANO A, FANELLI M

Decitabine, differently from DNMT1 silencing, exerts its antiproliferative activity through p21 upregulation in malignant pleural mesothelioma (MPM) cells.

LUNG CANCER

184 - 190

2,97

ANDREONI B.

Il paziente con dolore addominale acuto

Diagnosi e trattamento delle emergenze medicochirurgiche - ANDREONI B., CHIARA O, COEN D., VESCONI S.

331 - 347

ANDREONI B., CROSTA C., SONZOGNI A. M., PIROLA ME, PAVAN A, BISANTI L, SENORE C, SASSATELLI R, SGUINZI R., BERTANI E., BIANCHI P.P., CHIAPPA A.

Comparison between endoscopic and surgical treatment of screen-detected nonscreen-detected colorectal cancers

ECANCERMEDICALSCIENCE

255 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

ANDREONI B., GOLDHIRSCH A., ORECCHIA R., VENTURINO M., SPIRITO R, TADINI L. G., CORBELLINI C., BERTANI E., VERONESI U.

Correlation between administered treatment and patient’s Living Will

ECANCERMEDICALSCIENCE

ANDREONI B., COEN D

Il paziente terminale a domicilio

ANDREONI B., DESTREBECQ G, SESANA G

158

Diagnosi e trattamento delle emergenze medicochirurgiche - ANDREONI B., CHIARA O, COEN D., VESCONI S.

387 - 394

La formazione degli operatori in area critica

Diagnosi e trattamento delle emergenze medicochirurgiche - ANDREONI B., CHIARA O, COEN D., VESCONI S.

007-11

ANDREONI B., BETTONCELLI G

La prevenzione primaria

TUMORI

S19 - S21

0,791

ANDREONI B., CROSTA C., CHIAPPA A., BIANCHI P. P., SONZOGNI A. M., MISITANO P., CORBELLINI C., SENORE C, VERONESI U.

Sentinel node in colon cancer

NOWOTWORY J ONC

120e 122e

ANELLI V, SANTORIELLO C, DISTEL M, KOSTER RW, CICCARELLI F., MIONE M

Global repression of cancer gene expression in a zebraﬁsh model of melanoma is linked to epigenetic regulation.

ZEBRAFISH

417 - 424

ANSARIN M., SANTORO L., CATTANEO A., MASSARO M. A., CALABRESE L., GIUGLIANO G., MAFFINI F. A., OSTUNI A, CHIESA F. G.

Laser surgery for early glottic cancer

ARCH OTOLARYNGOL

135

385 - 390

1,829

ANTONIOU AC, SINILNIKOVA OM, MCGUFFOG L, HEALEY S, NEVANLINNA H, SIMARD J, SPURDLE AB, BEESLEY J, CHEN X, THE KATHLEEN CUNINGHAM FOUNDATION CONSORTIUM FOR RESEARCH INTO FAMILIAL BREAST CANCER, NEUHAUSEN SL, DING YC, COUCH FJ, WANG X, FREDERICKSEN Z, PETERLONGO P, PEISSEL B, BONANNI B., VIEL A, BERNARD L, RADICE P, SZABO CI, FORETOVA L, ZIKAN M, CLAES K, GREENE MH, MAI PL, RENNERT G, LEJBKOWICZ F, ANDRULIS IL, OZCELIK H, GLENDON G, OCGN, GERDES AM, THOMASSEN M, SUNDE L, CALIGO MA, LAITMAN Y, KONTOROVICH T, COHEN S, KAUFMAN B, DAGAN E, BARUCH RG, FRIEDMAN E, HARBST K, BARBANY-BUSTINZA G, RANTALA J, EHRENCRONA H, KARLSSON P, DOMCHEK SM, NATHANSON KL, OSORIO A, BLANCO I, LASA A, BENITEZ J, HAMANN U, HOGERVORST FB, ROOKUS MA, COLLEEE JM, DEVILEE P, LIGTENBERG MJ, VAN DER LUIJT RB, AALFS CM, WAISFISZ Q, WIJNEN J, VAN ROOZENDAAL CE, HEBON, PEOCK S, COOK M, FROST D, OLIVER C, PLATTE R, EVANS DG, LALLOO F, EELES R, IZATT L, DAVIDSON R, CHU C, ECCLES D, COLE T, HODGSON S, EMBRACE, GODWIN AK, STOPPA - LYONNET D, BUECHER B, LEONE M, BRESSAC-DE-PAILLERETS B, REMENIERAS A, CARON O, LENOIR GM, SEVENET N, LONGY M, FERRER SF, PRIEUR F, GEMO, GOLDGAR D, MIRON A, JOHN EM, BUYS SS, DALY MB, HOPPER JL, TERRY MB, YASSIN Y, BREAST CANCER FAMILY REGISTRY, SINGER CF, GSCHWANTLER-KAULICH D, STAUDIGL C, HANSEN TV, BARLARKARDOTTIR RB, KIRCHHOFF T, PAL P, KOSARIN K, OFFIT K, PIEDMONTE M, RODRIGUEZ GC,, WAKELEY K, BOGGESS JF, BASIL J, SCHWARTZ PE, BLANK SV, TOLAND AE, MONTAGNA M, CASELLA C, IMYANITOV EN, ALLAVENA A, SCHMUTZLER RK, VERSMOLD B, ENGEL C, MEINDL A, DITSCH N, ARNOLD N, NIEDERACHER D, DEIBLER H, FIEBIG B, SUTTNER C, SCHONBUCHNER I, GADZICKI D, CALDES T, DE LA HOYA M, POOLEY KA, EASTON DF, GEORGIA CHENEVIX-TRENCH, ON BEHALF OF CIMBA

Common variants in LSP1, 2q35 and 8q24 and breast cancer risk for BRCA1 and BRCA2 mutation carriers

HUM MOL GENET

4442 4456

7,249

256 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

APOLONE G, CORLI O, NEGRI E, MANGANO S, MONTANARI M, GRECO M T, ON BEHALF OF THE WRITING PROTOCOL COMMITTEE AND THE CANCER PAIN OUTCOME RESEARCH STUDY GROUP (CPOR SG) INVESTIGATORS, SBANOTTO A.

Effects of Transdermal Buprenorphine on Patients-reported Outcomes in Cancer Patients Results From the Cancer Pain Outcome Research (CPOR) Study Group

CLIN J PAIN

671 - 682

2,889

APOLONE G, CORLI O, CARACENI A, NEGRI E, DEANDREA S, MONTANARI M, GRECO MT, ON BEHALF OF THE CANCER PAIN OUTCOME RESEARCH STUDY GROUP, SBANOTTO A.

Pattern and quality of care of cancer pain management. Results from the cancer pain outcome research study group

BRIT J CANCER

100

1566 1574

4,846

ARICO’ D., GRANA C. M., VANAZZI A., FERRARI M, MALLIA A. J., SANSOVINI M, MARTINELLI G., PAGANELLI G., CREMONESI M.

The role of dosimetry in the high activity 90Y-ibritumomab tiuxetan regimens: two cases of abnormal biodistribution.

CANCER BIOTHER RADIO

271 - 275

1,318

AZIM H., PECCATORI F. A., LIPTROTT S. J., CATANIA C. M., GOLDHIRSCH A.

Breast cancer and pregnancy: how safe is trastuzumab?

NAT REV CLIN ONCOL

367 - 370

AZIM H., BELLETTINI G., GELBER S, PECCATORI F. A.

Breast-feeding after breast cancer: if you wish, madam.

BREAST CANCER RES TR

114

007-12

5,684

AZIM H., SCARFONE G, PECCATORI F. A.

Carboplatin and weekly paclitaxel for the treatment of advanced non-small cell lung cancer (NSCLC) during pregnancy

J THORAC ONCOL

559 - 560

3,508

AZIM H., PRUNERI G., COCOROCCHIO E., CINIERI S., RAFANIELLO RAVIELE P., BASSI S., PREDA L., MARTINELLI G., PECCATORI F. A.

Rituximab in lymphocyte-predominant Hodgkin disease.

ONCOLOGY-BASEL

26 - 29

1,336

BAJETTA E, BOMBARDIERI E, SILVESTRINI R, CAPELLA C, CATENA L, CHITI A, CIRILLO F, COLAO A, DE ANGELIS C, DUCCESCHI M, MARCHIANO’ A, MARTINETTI A, MAZZAFERRO V, PAGANELLI G., PLATANIA M, SEREGNI E, SPREAFICO C, BAIO S. M., BHOORI S, BODEI L., CAMERINI T, CHINOL M., COPPA J, CREMONESI M., FAGGIANO A, LA ROSA S, LOMBARDI G, MILONE F, PUSCEDDU S, RAMUNDO V, SCHIAVO M, VITALI M

Basi scientiﬁche per la deﬁnizione di linee-guida in ambito clinico per i Tumori Neuroendocrini del tratto Gastro-EnteroPancreatico (GEP)

DIVULGAZIONE ON LINE

1 - 201

BALDUZZI A., MONTAGNA E., BAGNARDI V., TORRISI R., BERTOLINI F., MANCUSO P., SCARANO E., VIALE G., VERONESI P., CARDILLO A., ORLANDO L., GOLDHIRSCH A., COLLEONI M. A.

Infusional ﬂuorouracil, epirubicin, and cisplatin followed by weekly paclitaxel plus bevacizumab in locally advanced breast cancer with unfavorable prognostic features.

ANTI-CANCER DRUG

197 - 203

2,358

BALLABENI A., ZAMPONI R., CAPRARA G., MELIXETIAN M., BOSSI S., MASIERO L., HELIN K.

Human CDT1 associates with CDC7 and recruits CDC45 to chromatin during S phase.

J BIOL CHEM

284

3028 3036

5,520

BARKER J J, BARKER O, BOGGIO R, CHAUHAN V, CHENG R K Y, CORDEN V, COURTNEY S M, EDWARDS N, FALQUE V M, FUSAR F, GARDINER M, HAMELIN E M N, HESTERKAMP T, ICHIHARA O, JONES R S, MATHER O, MERCURIO C, MINUCCI S., MONTALBETTI C A G N, MULLER A, PATEL D, PHILLIPS B G, VARASI M, WHITTAKER M, WINKLER D, YARNOLD C J

Fragment-based identiﬁcation of Hsp90 inhibitors.

CHEMMEDCHEM

963 - 966

3,150

BARTOLOMEI M., BODEI L., DE CICCO C., GRANA C. M., CREMONESI M., BOTTERI E., BAIO S. M., ARICO’ D., SANSOVINI M, PAGANELLI G.

Peptide receptor radionuclide therapy with (90)Y-DOTATOC in recurrent meningioma.

EUR J NUCL MED MOL I

1407 - 1416

4,532

BAZZI C, RIZZA V, RAIMONDI S., CASELLATO D, NAPODANO P, D’AMICO G

In Crescentic IgA Nephropathy, Fractional Excretion of IgG in Combination with Nephron Loss Is the Best Predictor of Progression and Responsiveness to Immunosuppression.

CLIN J AM SOC NEPHRO

929 - 935

4,361

BEGNOZZI L, BENASSI M, BERTANELLI M, BONINI A, CIONINI L, CONTE L, FIORINO C, GABRIELE P, GARDANI G, GIANI A, MAGRI S, MORELLI M, MORRICA B, OLMI P, ORECCHIA R., PENDUZZU G, RAFFAELE L, ROSI A, TABOCCHINI MA, VALDAGNI R, VITI V

Quality assurance of 3D-CRT: indications and difﬁculties in their applications.

CRIT REV ONCOL HEMAT

24 - 38

4,589

BELLETTINI G, PECCATORI F. A.

Allattamento al seno dopo neoplasia mammaria

Fertilità e Oncologia Banna G.L., Catalano F., Condorelli R., Peccatori F.

121 - 123

BELLOMI M., RAMPINELLI C., DE FIORI E., PREDA L., VERONESI G.

Lung cancer screening update.

CANCER IMAGING

s122 - s125

257 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

BERMEJO R, CAPRA T, GONZALEZ-HUICE V, FACHINETTI D, COCITO A., NATOLI G., KATOU Y, MORI H, KUROKAWA K, SHIRAHIGE K, FOIANI M

Genome-Organizing Factors Top2 and Hmo1 Prevent Chromosome Fragility at Sites of S phase Transcription

CELL

138

870 - 884

31,253

BERTANI E., CHIAPPA A., TESTORI A., MAZZAROL G., BIFFI R., MARTELLA S., PACE U., SOTELDO J. A., DELLA VIGNA P., LEMBO R., ANDREONI B.

Desmoid tumors of the Anterior Abdominal Wall: results from a monocentric surgical experience and review of the literature

ANN SURG ONCOL

1642 1649

3,898

BERTOLINI F.

Biomarkers for angiogenesis and antiangiogenic drugs in clinical oncology

BREAST

4S8 - S50

2,155

BERTOLINI F., MANCUSO P., BRAIDOTTI P, SHAKED Y, KERBEL R S

The contribution of circulating endothelial cells to tumor angiogenesis.

Recent Advances in Angiogenesis and Antiangiogenesis

59 - 66

BERTOLINI F., MANCUSO P., BRAIDOTTI P, SHAKED Y, KERBEL RS

The multiple personality disorder phenotype(s) of circulating endothelial cells in cancer.

BBA-REV CANCER

1796

27 - 32

10,283

BIFFI R., ORSI F., POZZI S., PACE U., BONOMO G., MONFARDINI L., DELLA VIGNA P., ROTMENSZ N., RADICE D., ZAMPINO M. G., FAZIO N., DE BRAUD F., ANDREONI B., GOLDHIRSCH A.

Best choice of central venous insertion site for the prevention of catheter-related complications in adult patients who need cancer therapy: a randomized trial.

ANN ONCOL

935 - 940

4,935

BIFFI R., SABBATINI A.

Infezione della ferita chirurgica sede di biopsia per LNH inguinale: risultati di un trattamento multimodale.

ARCH MED THER

BIGUZZI E, MANCUSO P., FRANCHI F, CALLERI A., MANCUSO ME, SANTAGOSTINO E, BUCCIARELLI P, BERTOLINI F., MANNUCCI PM

Circulating endothelial cells (CECs) and progenitors (CEPs) in severe haemophiliacs with different clinical phenotype.

BRIT J HAEMATOL

144

803 - 805

4,478

BODEI L., FERONE D, GRANA C. M., CREMONESI M., SIGNORE A, DIERCKX R.A, PAGANELLI G.

Peptide receptor therapies in neuroendocrine tumors.

J ENDOCRINOL INVEST

360 - 369

1,888

BOKEMEYER C, BONDARENKO I, MAKHSON A, HARTMANN JT, APARICIO J, DE BRAUD F., DONEA S, LUDWIG H, SCHUCH G, STROH C, LOOS AH, ZUBEL A, KORALEWSKI P

Fluorouracil, leucovorin, and oxaliplatin with and without cetuximab in the ﬁrst-line treatment of metastatic colorectal cancer.

J CLIN ONCOL

663 - 671

17,157

BONANNI B., SERRANO D., GANDINI S., GUERRIERI GONZAGA A., JOHANSSON H. A., MACIS D., CAZZANIGA M., LUINI A., CASSANO E., OLDANI S., LIEN EA, PELOSI G., DECENSI A.

Randomized Biomarker Trial of Anastrozole or Low-Dose Tamoxifen or Their Combination in Subjects with Breast Intraepithelial Neoplasia.

CLIN CANCER RES

7053 7060

6,488

BONANNI B., LAZZERONI M.

Retinoids and breast cancer prevention

RECENT RESULTS CANCER RES

181

77 - 82

BONARDI M L, MARTANO L., GROPPI F, CHINOL M.

Rapid determination of (90)Sr impurities in freshly “generator eluted”(90)Y for radiopharmaceutical preparation.

APPL RADIAT ISOTOPES

1874 - 1877

1,114

BOOKMAN MA, BRADY MF, MCGUIRE WP, HARPER PG, ALBERTS DS, FRIEDLANDER M, COLOMBO N., FOWLER JM, ARGENTA PA, DE GEEST K, MUTCH DG, BURGER RA, SWART AM, TRIMBLE EL, ACCARIO-WINSLOW C, ROTH LM

Evaluation of new platinum-based treatment regimens in advanced-stage ovarian cancer: a Phase III Trial of the Gynecologic Cancer Intergroup

J CLIN ONCOL

1419 - 1425

17,157

BOTRUGNO O. A., SANTORO F., MINUCCI S.

Histone deacetylase inhibitors as a new weapon in the arsenal of differentiation therapies of cancer.

CANCER LETT

280

134 - 144

3,504

BOTTERI E., IODICE S., MAISONNEUVE P., ALFIERI M. F., BURZONI N. C., MANGHI L. E., MARTINETTI M., MONTANARI B. M., ALBERTAZZI E., BAZOLLI B., ROTMENSZ N.

Case mix at the European Institute of Oncology: ﬁrst report of the Tumour Registry, years 2000-2002

ECANCERMEDICALSCIENCE

149

BOTTERI E., ROTMENSZ N., SANGALLI C. A., TOESCA A., PERADZE N., DE OLIVEIRA FILHO H. R., SAGONA A., INTRA M., VERONESI P., GALIMBERTI V. E., LUINI A., VERONESI U., GENTILINI O. D.

Unavoidable mastectomy for ipsilateral breast tumour recurrence after conservative surgery: patient outcome.

ANN ONCOL

1008 - 1012

4,935

BOTTOMLEY A, COENS C, SUCIU S, SANTINAMI M, KRUIT W, TESTORI A., MARSDEN J, PUNT C, SALES F, GORE M, MACKIE R, KUSIC Z, DUMMER R, PATEL P, SCHADENDORF D, SPATZ A, KEILHOLZ U, EGGERMONT A

Adjuvant Therapy With Pegylated Interferon Alfa-2b Versus Observation in Resected Stage III Melanoma: A Phase III Randomized Controlled Trial of Health-Related Quality of Life and Symptoms by the European Organisation for Research and Treatment of Ca

J CLIN ONCOL

2916 2923

17,157

BOZZETTI F, ON BEHALF OF THE SCRINIO WORKING GROUP, BIFFI R., PAPIS D., ZAMPINO M. G.

Screening the nutritional status in oncology: a preliminary report on 1,000 outpatients.

SUPPORT CARE CANCER

279 - 284

2,422

258 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

BRISTOW RE, PEIRETTI M., GERARDI M, ZANAGNOLO V., UEDA S, DIAZ-MONTES T, GIUNTOLI RL II, MAGGIONI A.

Secondary cytoreductive surgery including rectosigmoid colectomy for recurrent ovarian cancer: Operative technique and clinical outcome

GYNECOL ONCOL

114

173 - 177

2,919

BRUNO S, ZUIN M, CROSIGNANI A, ROSSI S, ZADRA F, ROFFI L, BORZIO M, REDAELLI A, CHIESA A, SILINI EM, ALMASIO PL, MAISONNEUVE P.

Predicting Mortality Risk in Patients With Compensated HCV-Induced Cirrhosis: A Long-Term Prospective Study.

AM J GASTROENTEROL

104

1147 - 1158

6,444

BRUNO W, GHIORZO P, BATTISTUZZI L, ASCIERTO P A, BARILE M., GARGIULO S, GENSINI F, GLIORI S, GUIDA M, LOMBARDO M, MANOUKIAN S, MENIN C, NASTI S, ORIGONE P, PASINI B, PASTORINO L, PEISSEL B, PIZZICHETTA M A, QUEIROLO P, RODOLFO M, ROMANINI A, SCAINI M C, TESTORI A., TIBILETTI M G, TURCHETTI D, LEACHMAN S A, BIANCHI SCARRA’ G, ON BEHALF OF IMI THE ITALIAN MELANOMA INTERGROUP

Clinical genetic testing for familial melanoma in Italy: A cooperative study.

J AM ACAD DERMATOL

775 - 782

4,081

BUA D J, KUO A J, CHEUNG P, LONG LIU C, MIGLIORI V, ESPEJO A, CASADIO C., BASSI C., AMATI B., BEDFORD M T, GUCCIONE E, GOZANI O

Epigenome microarray platform for proteome-wide dissection of chromatinsignaling networks.

PLOS ONE

E6789

CAINI S, GANDINI S., SERA F, RAIMONDI S., FARGNOLI MC, BONIOL M, ARMSTRONG BK

Meta-analysis of risk factors for cutaneous melanoma according to anatomical site and clinico-pathological variant.

EUR J CANCER

3054 3063

4,475

CALABRESE L., GIUGLIANO G., BRUSCHINI R., ANSARIN M., NAVACH V., GROSSO E., GIBELLI B., OSTUNI A., CHIESA F.G.

Compartmental surgery in tongue tumours: description of a new surgical technique.

ACTA OTORHINOLAR ITAL

259-264

CALLERI A., BONO A., BAGNARDI V., QUARNA J., MANCUSO P., RABASCIO C., DELLAPASQUA S., CAMPAGNOLI E., SHAKED Y, GOLDHIRSCH A., COLLEONI M. A., BERTOLINI F.

Predictive potential of angiogenic growth factors and circulating endothelial cells in breast cancer patients receiving metronomic chemotherapy plus bevacizumab.

CLIN CANCER RES

7652 7657

6,488

CAMBRIA R., JERECZEK B. A., CATTANI F., GARIBALDI C., ZERINI D., FODOR C., SERAFINI F., PEDROLI G., ORECCHIA R.

Evaluation of late rectal toxicity after conformal radiotherapy for prostate cancer. A comparison between dose-volume constraints and NTCP use

STRAHLENTHER ONKOL

185

384 - 389

3,005

CAPRARA G., ZAMPONI R., MELIXETIAN M., HELIN K.

Isolation and characterization of DUSP11, a novel p53 target gene.

J CELL MOL MED

2158 2170

5,114

CAPURSO G, FAZIO N., FESTA S, PANZUTO F, DE BRAUD F., DELLE FAVE G.

Molecular target therapy for gastroenteropancreatic endocrine tumours: Biological rationale and clinical perspectives.

CRIT REV ONCOL HEMAT

110 - 124

4,589

CARDINALE D. M., COLOMBO A., CIPOLLA C.

Cardiotossicità da agenti antitumorali

PRATICA MEDICA & ASPETTI LEGALI

155 - 161

CARDINALE D. M.

Tossicità cardiaca da chemioterapici. Gestione clinica

IL CARDIOLOGO

18 - 24

CARDOSO F, BEDARD PL, WINER EP, PAGANI O, SENKUS-KONEFKA E, FALLOWFIELD LJ, KYRIAKIDES S, COSTA A, CUFER T, ALBAIN K S, ON BEHALF ON THE ESO MBC TASK FORCE, COLLEONI M. A.

International guidelines for management of metastatic breast cancer: combination vs sequential single-agent chemotherapy.

J NATL CANCER I

101

1174 - 1181

14,933

CARPI A., MENABO’ R, KALUDERCIC N, PELICCI P. G., DI LISA F, GIORGIO M.

The cardioprotective effects elicited by p66(Shc) ablation demonstrate the crucial role of mitochondrial ROS formation in ischemia/reperfusion injury.

BBA-BIOENERGETICS

1787

774 - 780

4,447

CASSANO E., BROLINI DELLE URBAN L. A., CIANFARANO A.

Diagnostica per immagini delle lesioni mammarie

CHIRURGIA DELLA MAMMELLA -RIETJENS M., URBAN C.A.

62 - 85

CASULA M, ALAIBAC M, PIZZICHETTA M.A., BONO R, ASCIERTO P A, STANGANELLI I, CANZANELLA S, PALOMBA G, ZATTRA E, THE ITALIAN MELANOMA INTERGROUP (IMI), PALMIERI G., TESTORI A.

Role of the EGF +61A>G polymorphism in melanoma pathogenesis: an experience on a large series of Italian cases and controls.

BMC CANCER

3,087

CASULA M, MUGGIANO A, COSSU A, BUDRONI M, CARACO’ C, ASCIERTO P A, PAGANI E, STANGANELLI I, CANZANELLA S, SINI MC, PALOMBA G, THE ITALIAN MELANOMA INTERGROUP (IMI), TESTORI A., PALMIERI G.

Role of key-regulator genes in melanoma susceptibility and pathogenesis among patients from South Italy.

BMC CANCER

352

3,087

259 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

CATALANO V, LABIANCA R, BERETTA G.D, GATTA G, DE BRAUD F., VAN CUTSEM E

Gastric cancer

CRIT REV ONCOL HEMAT

127 - 164

4,589

CATANIA C. M., ZAGONEL V, ITALIAN SOCIETY OF MEDICAL ONCOLOGY (AIOM) WORKING GROUP: “HUMANIZATION AND ORGANIZATION”

Care and self-care “god as a mirror of the self”.

J CANCER EDUC

80 - 82

0,513

CATANIA C. M., SOTIROPOULOS I, SILVA R, ONOFRI C, BREEN KC, SOUCA N, ALMEIDA OF

The amyloidogenic potential and behavioral correlates of stress.

MOL PSYCHIATR

95 - 105

12,537

CATUCCI I., VERDERIO P, PIZZAMIGLIO S, MANOUKIAN S, PEISSEL B, BARILE M., TIZZONI L, BERNARD L., RAVAGNANI F, GALASTRI L, PIEROTTI M A, RADICE P, PETERLONGO P

SNPs in ultraconserved elements and familial breast cancer risk

CARCINOGENESIS

544 - 545

4,93

CAZZANIGA M., GHEIT T, CASADIO C., KHAN N, MACIS D., VALENTI F., MILLER M. J., SYLLA B.S, AKIBA S, BONANNI B., DECENSI A., VERONESI U., TOMMASINO M

Analysis of the presence of cutaneous and mucosal papillomavirus types in ductal lavage ﬂuid, milk and colostrum to evaluate its role in breast carcinogenesis

BREAST CANCER RES TR

114

599 - 605

5,684

CAZZANIGA M., DECENSI A., BONANNI B., LUINI A., GENTILINI O. D.

Biomarkers for risk assessment and prevention of breast cancer.

CURR CANCER DRUG TAR

482 - 499

4,316

CAZZANIGA M., BONANNI B., GUERRIERI GONZAGA A., DECENSI A.

Is it time to test metformin in breast cancer clinical trials?

CANCER EPIDEM BIOMAR

701 - 705

4,77

CHAKERA A.H, HESSE B, BURAK Z, BALLINGER J R, BRITTEN A, CARACO’ C, COCHRAN A J, COOK M G, DRZEWIECKI K T, ESSNER R, EVEN SAPIR E, EGGERMONT A M M, GMEINER STOPAR T, INGVAR C, MIHM JR M C, McCARTHY S W, MOZZILLO N, NIEWEG O E, SCOLYER R A, STARZ H, THOMPSON J F, TRIFIRO’ G., VIALE G., VIDALSICART S, UREN R, WADDINGTON W, CHITI A, SPATZ A, TESTORI A.

EANM-EORTC general recommendations for sentinel node diagnostics in melanoma.

EUR J NUCL MED MOL I

1713 - 1742

4,532

CHIAPPA A., BERTANI E., MAKUUCHI M, ZBAR AP, CONTINO G., VIALE G., PRUNERI G., BELLOMI M., DELLA VIGNA P., ZAMPINO M. G., FAZIO N., TRAVAINI L. L., TRIFIRO’ G., CORBELLINI C., ANDREONI B.

Neoadjuvant chemotherapy followed by hepatectomy for primarily resectable colorectal cancer liver metastases

HEPATO-GASTROENTEROL

829 - 834

0,680

CHIAPPA A., MAKUUCHI M, LYGIDAKIS NJ, ZBAR AP, CHONG G, BERTANI E., SITZLER PJ, BIFFI R., PACE U., BIANCHI P. P., CONTINO G., MISITANO P., ORSI F., TRAVAINI L. L., TRIFIRO’ G., ZAMPINO M. G., FAZIO N., GOLDHIRSCH A., ANDREONI B.

The management of colorectal liver metastases: Expanding the role of hepatic resection in the age of multimodal therapy.

CRIT REV ONCOL HEMAT

65 - 75

4,589

CICALESE A., BONIZZI G., PASI C. E., FARETTA M. R., RONZONI S., GIULINI B., BRISKEN C, MINUCCI S., DI FIORE P. P.

The tumor suppressor p53 regulates polarity of self-renewing divisions in mammary stem cells.

CELL

138

1083 1095

31,253

CIOCCA M., PEDROLI G., ORECCHIA R., GUIDO A. N., CATTANI F., CAMBRIA R., VERONESI U.

Radiation survey around a Liac mobile electron linear accelerator for intraoperative radiation therapy.

J APPL CLIN MED PHYS

131 - 138

1,225

CIRO’ M., PROSPERINI E., QUARTO M, GRAZINI U., WALFRIDSSON J, MC BLANE J. F., NUCIFERO P, PACCHIANA G., CAPRA M., CHRISTENSEN J, HELIN K.

ATAD2 Is a Novel Cofactor for MYC, Overexpressed and Ampliﬁed in Aggressive Tumors.

CANCER RES

8491 8498

7,514

COATES A S, GOLDHIRSCH A., GELBER R.D., INGLE J N, THUERLIMANN B, SENN H.-J

Reply to Does the expert panel at the St Gallen meeting provide an unbiased opinion about the management of women with early breast cancer?

ANN ONCOL

1751 - 1752

4,935

CODACCI-PISANELLI G, SPINELLI GP, PECCATORI F. A.

Have we reached a plateau in adjuvant breast cancer therapy?

LANCET

374

1421

28,409

COELHO R.F, CHAUHAN S, PALMER K.J, ROCCO B. M., PATEL M B, PATEL V R

Robotic-assisted radical prostatectomy: a review of current outcomes.

BJU INT

104

1428 1435

2,704

COLLEONI M. A., BAGNARDI V., ROTMENSZ N., DELLAPASQUA S., VIALE G., PRUNERI G., VERONESI P., TORRISI R., LUINI A., INTRA M., GALIMBERTI V. E., MONTAGNA E., GOLDHIRSCH A.

A risk score to predict disease-free survival in patients not achieving a pathological complete remission after preoperative chemotherapy for breast cancer.

ANN ONCOL

1178 - 1184

4,935

260 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

COLLEONI M. A., BAGNARDI V., ROTMENSZ N., GELBER RD, VIALE G., PRUNERI G., VERONESI P., TORRISI R., CARDILLO A., MONTAGNA E., CAMPAGNOLI E., LUINI A., INTRA M., GALIMBERTI V. E., SCARANO E., PERUZZOTTI G., GOLDHIRSCH A.

Increasing steroid hormone receptors expression deﬁnes breast cancer subtypes non responsive to preoperative chemotherapy.

BREAST CANCER RES TR

116

359 - 369

5,684

COLLEONI M. A., VIALE G., GOLDHIRSCH A.

Lessons on responsiveness to adjuvant systemic therapies learned from the neoadjuvant setting

BREAST

S137 S149

2,155

COLLEONI M. A., SUN Z, MARTINELLI G., BASSER RL, COATES AS, GELBER RD, GREEN MD, PECCATORI F. A., CINIERI S., AEBI S, VIALE G., PRICE KN, GOLDHIRSCH A., FOR THE INTERNATIONAL BREAST CANCER STUDY GROUP, VERONESI U., LUINI A., ORECCHIA R., COCOROCCHIO E., RENNE G., MAZZAROL G., AGAZZI A., NOLE’ F., COSTA A., ZURRIDA S., VERONESI P., SACCHINI V., GALIMBERTI V. E., DE BRAUD F., PERUZZOTTI G., DIDIER F.

The effect of endocrine responsiveness on high-risk breast cancer treated with dose-intensive chemotherapy: results of International Breast Cancer Study Group Trial 15-95 after prolonged follow-up.

ANN ONCOL

1344 - 1351

4,935

COLOMBO N., CIVELLI M., CARDINALE D. M., LAMANTIA G., COLOMBO A., DE GIACOMI G. D., CIPOLLA C.

Long QT syndrome and torsade de pointes after anthracycline chemotherapy

ECANCERMEDICALSCIENCE

COLOMBO N., PEIRETTI M., CASTIGLIONE M, ESMO GUIDELINES WORKING GROUP

Non-epithelial ovarian cancer: ESMO clinical recommendations for diagnosis, treatment and follow-up.

ANN ONCOL

24 - 26

4,935

CONFALONIERI S, QUARTO M, GOISIS G., NUCIFORO P, DONZELLI M., JODICE G, PELOSI G., VIALE G., PECE S., DI FIORE P. P.

Alterations of ubiquitin ligases in human cancer and their association with the natural history of the tumor.

ONCOGENE

2959 2968

7,216

COSTA S., GIUGLIANO G., SANTORO L., YWATA DE CARVALHO A., MASSARO M. A., GIBELLI B., DE FIORI E., GROSSO E., ANSARIN M., CALABRESE L.

Role of prophylactic central neck dissection in cN0 papillary thyroid cancer.

ACTA OTO-LARYNGOL

61 - 69

0,868

CRIVELLARI D, COLLEONI M. A., DELLAPASQUA S., GOLDHIRSCH A.

Re: Metronomic chemotherapy in elderly patients: do risks exceed beneﬁts in some patients?

TUMORI

130 - 131

0,791

CURIGLIANO G., SPITALERI G., MAGNI E., LORIZZO K., DE COBELLI O., LOCATELLI M. A., FUMAGALLI L., ADAMOLI L., COSSU ROCCA M., VERRI E., DE PAS T., JERECZEK B. A., MARTINELLI G., GOLDHIRSCH A., NOLE’ F.

Cisplatin, Etoposide and Continuous Infusion Bleomycin in Patients with Testicular Germ Cell Tumors: Efﬁcacy and Toxicity Data from a Retrospective Study.

J CHEMOTHERAPY

687 - 692

0,843

CURIGLIANO G., VIALE G., BAGNARDI V., FUMAGALLI L., LOCATELLI M. A., ROTMENSZ N., GHISINI R., COLLEONI M. A., MUNZONE E., VERONESI P., ZURRIDA S., NOLE’ F., GOLDHIRSCH A.

Clinical Relevance of HER2 Overexpression/ Ampliﬁcation in Patients With Small Tumor Size and Node-Negative Breast Cancer.

J CLIN ONCOL

5693 5699

17,157

CURIGLIANO G., SPITALERI G., DE COBELLI O., SCARDINO E., SBANOTTO A., DE BRAUD F.

Health-related quality of life in patients with hormone refractory prostate cancer receiving geﬁtinib.

UROL INT

196 - 202

0,891

CURIGLIANO G., LOCATELLI M. A., FUMAGALLI L., GOLDHIRSCH A.

Immunizing against breast cancer: A new swing for an old sword

BREAST

S51 - S54

2,155

CURIGLIANO G., SPITALERI G., DE BRAUD F., CARDINALE D. M., CIPOLLA C., CIVELLI M., COLOMBO N., COLOMBO A., LOCATELLI M. A., GOLDHIRSCH A.

QTc prolongation assessment in anticancer drug development: Clinical and methodological issues

ECANCERMEDICALSCIENCE

130

DANEMAN R, RESCIGNO M.

The gut immune barrier and the blood-brain barrier: are they so different?

IMMUNITY

722 - 735

20,579

DE CENSI A., ROBERTSON C, GUERRIERI GONZAGA A., SERRANO D., CAZZANIGA M., MORA S., GULISANO M, JOHANSSON H. A., GALIMBERTI V. E., CASSANO E., MORONI S. M., FORMELLI F, LIEN E.A, PELOSI G., JOHNSON K.A, BONANNI B.

Randomized Double-Blind 2 x 2 Trial of Low-Dose Tamoxifen and Fenretinide for Breast Cancer Prevention in High-Risk Premenopausal Women.

J CLIN ONCOL

3749 3756

17,157

DE CICCO L., VAVASSORI A., CATTANI F., JERECZEK B. A., ORECCHIA R.

Salvage high dose rate brachytherapy after primary external beam irradiation in localized prostate cancer: a case report.

TUMORI

553 - 556

0,791

DE GRASSI A., CICCARELLI F.

Tandem repeats modify the structure of human genes hosted in segmental duplications.

GENOME BIOL

r137

6,153

261 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

DE PAS T., SPITALERI G., PELOSI G., DE CARLIS L, LORIZZO K., LOCATELLI M. A., CURIGLIANO G., TOFFALORIO F., CATANIA C. M., DE BRAUD F.

Erlotinib combined with cyclosporine in a liver-transplant recipient with epidermal growth factor receptor-mutated non-small cell lung cancer.

J THORAC ONCOL

138 - 139

3,508

DE PAS T., TOFFALORIO F., CATANIA C. M., NOBERASCO C., SPITALERI G., SPAGGIARI L., DE BRAUD F.

Gemcitabine and pemetrexed combination: the key role of the sequence of drugs administration.

ANN ONCOL

1747 - 1748

4,935

DE ROBERTO G., RAVIZZA D., FIORI G., TROVATO C. M., MAFFINI F. A., TAMAYO D., CROSTA C.

A massive gastric xanthomatosis.

ENDOSCOPY

E54 - E55

6,091

DE SANTA F., NARANG V, YAP ZH, TUSI BK, BURGOLD F. T., AUSTENAA L. M., BUCCI G., CAGANOVA M, NOTARBARTOLO S., CASOLA S, TESTA G., SUNG WK, WEI CL, NATOLI G.

Jmjd3 contributes to the control of gene expression in LPS-activated macrophages.

EMBO J

3341 3352

8,295

DE VECCHI G., VERDERIO P., PIZZAMIGLIO S., MANOUKIAN S., BARILE M., FORTUZZI S., RAVAGNANI F., PIEROTTI M. A., RADICE P., PETERLONGO P.

Evidences for association of the CASP8-652 6n del promoter polymorphism with age at diagnosis in familial breast cancer cases

BREAST CANCER RES TR

113

607 - 608

5,684

DE WULF P. A., MONTANI F., VISINTIN R.

Protein phosphatases take the mitotic stage.

CURR OPIN CELL BIOL

806 - 815

12,543

DELLA VIGNA P., MONFARDINI L., BONOMO G., CURIGLIANO G., AGAZZI A., BELLOMI M., ORSI F.

Coagulation Disorders in Patients with Cancer: Nontunneled Central Venous Catheter Placement with US Guidance--A Single-Institution Retrospective Analysis.

RADIOLOGY

253

249 - 252

5,996

DELLAPASQUA S.

Investigations after adjuvant therapy.

CANCER TREAT RES

151

331 - 352

DEMOE J, SANTAGUIDA S., DAUM J R, MUSACCHIO A., GORBSKY G J

A high throughput, whole cell screen for small molecule inhibitors of the mitotic spindle checkpoint identiﬁes OM137, a novel Aurora kinase inhibitor.

CANCER RES

1509 - 1516

7,514

DESMEDT C, GIOBBIE-HURDER A, NEVEN P, PARIDAENS R, CHRISTIAENS M.R, SMEETS A, LALLEMAND F, HAIBE-KAINS B, VIALE G., GELBER R.D, PICCART M, SOTIRIOU C

The Gene expression Grade Index: a potential predictor of relapse for endocrinetreated breast cancer patients in the BIG 1-98 trial.

BMC GENOMICS

3,926

DI LISA F., KALUDERCIC N., CARPI A., MENABO’ R., GIORGIO M.

Mitochondria and vascular pathology.

PHARMACOL REP

123 - 130

2,167

DI LISA F., KALUDERCIC N., CARPI A., MENABO’ R., GIORGIO M.

Mitochondrial pathways for ROS formation and myocardial injury: the relevance of p66(Shc) and monoamine oxidase.

BASIC RES CARDIOL

104

131 - 139

5,407

DIDIER F., RADICE D., GANDINI S., BEDOLIS R., ROTMENSZ N., MALDIFASSI A., SANTILLO B., LUINI A., GALIMBERTI V. E., SCAFFIDI E., LUPO F. N., MARTELLA S., PETIT J. Y.

Does nipple preservation in mastectomy improve satisfaction with cosmetic results, psychological adjustment, body image and sexuality?

BREAST CANCER RES TR

118

623 - 633

5,684

D’ONOFRIO A., CERRAI P.

A bi-parametric model for the tumour angiogenesis and antiangiogenesis therapy

MATH COMPUT MODEL

1156 - 1163

1,032

D’ONOFRIO A., GANDOLFI A

A family of models of angiogenesis and anti-angiogenesis anti-cancer therapy

MATH MED BIOL

63 - 95

1,289

D’ONOFRIO A.

Fractal growth of tumors and other cellular populations: linking the mechanistic to the phenomenological modeling and vice versa

CHAOS SOLITON FRACT

875 - 880

2,980

D’ONOFRIO A., MANFREDI P

Information-related changes in contact patterns may trigger oscillations in the endemic prevalence of infectious diseases.

J THEOR BIOL

256

473 - 478

2,454

D’ONOFRIO A., LEDZEWICS U, MAURER H, SCHATTLER H

On Optimal Delivery of Combination Therapy for Tumors.

MATH BIOSCI

222

13 - 26

1,148

D’ONOFRIO A., GANDOLFI A, ROCCA A

The dynamics of tumour-vasculature interaction suggest low-dose, time-dense anti-angiogenic schedulings

CELL PROLIFERAT

317 - 329

2,423

DOWSETT M, PROCTER M, MC CASKILL-STEVENS W, DE AZAMBUJA E, DAFNI U, RUESCHOFF J, JORDAN B, DOLCI S, ABRAMOVITZ M, STOSS O, VIALE G., GELBER RD, PICCART-GEBHART M, LEYLAND-JONES B

Disease-Free Survival According to Degree of HER2 Ampliﬁcation for Patients Treated With Adjuvant Chemotherapy With or Without 1 Year of Trastuzumab: The HERA Trial.

J CLIN ONCOL

2962 2969

17,157

262 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

DUELL E.J, MAISONNEUVE P., BAGHURST P A, BUENO DE MESQUITA HB, GHADIRIAN P, MILLER A B, ZATONSKI W, VRIELING A, BOFFETTA P, BOYLE P

Menstrual and reproductive factors and pancreatic cancer in the SEARCH program of the IARC.

CANCER CAUSE CONTROL

1757 - 1762

3,690

EGGERMONT A M M, TESTORI A., MARSDEN J, HERSEY P, QUIRT I, PETRELLA T, GOGAS H, MACKIE R M, HAUSCHILD A

Utility of adjuvant systemic therapy in melanoma.

ANN ONCOL

VI30 VI34

4,935

FABBRI C, SARTI G, CREMONESI M., FERRARI M, DI DIA A, AGOSTINI M, BOTTA F., PAGANELLI G.

Quantitative analysis of 90Y Bremsstrahlung SPECT-CT images for application to 3D patient-speciﬁc dosimetry.

CANCER BIOTHER RADIO

145 - 154

1,318

FAZIO N., LUCA F., MONFARDINI L., PELOSI G., BODEI L., LORIZZO K., DI MEGLIO G., GIBELLI B., RAVIZZA D., BONOMO G., GRANA C. M., BAIO S. M., SQUADRONI M., PAGANELLI G., DE BRAUD F.

Right pelvic mass in a patient with a radically resected carcinoid of the appendix

GUT

1200, 1259

9,766

FELICE B, CATTOGLIO C, CITTARO D, TESTA A, MICCIO A, FERRARI G, LUZI L

Transcription factor binding sites are genetic determinants of retroviral integration in the human genome.

PLOS ONE

E4571

FERRARI A., IVALDI G. B., LEONARDI M. C., RONDI E., ORECCHIA R.

Prone breast radiotherapy in a patient with early stage breast cancer and a large pendulous breast

TUMORI

394 - 397

0,791

FERRAZZI E, ZUPI E, LEONE FP, SAVELLI L, OMODEI U, MOSCARINI M, BARBIERI M, CAMMARERI G, CAPOBIANCO G, CICINELLI E, COCCIA ME, DONARINI G, FIORE S, LITTA P, SIDERI M. G., SOLIMA E, SPAZZINI D, TESTA A C, VIGNALI M

How often are endometrial polyps malignant in asymptomatic postmenopausal women? A multicenter study.

AM J OBSTET GYNECOL

200

235.e1 235.e6

3,453

FORMELLI F, MENEGHINI E, CAVADINI E, CAMERINI T, DI MAURO MG, DE PALO G, VERONESI U., BERRINO F, MICHELI A

Plasma retinol and prognosis of postmenopausal breast cancer patients.

CANCER EPIDEM BIOMAR

42 - 48

4,770

FOSSATI P., RICARDI U, ORECCHIA R.

Pediatric medulloblastoma: Toxicity of current treatment and potential role of protontherapy.

CANCER TREAT REV

79 - 96

4,729

FRANCHI D., BOVERI S., MAGGIONI A., CASADIO C., RIZZO S., SIDERI M. G.

OP24.04: Correlation of transvaginal ultrasound ﬁndings and surgical specimen pathology in early stage cervical cancer: a prospective, single institution observational study.

ULTRASOUND OBST GYN

140

2,690

FRITAH S, COL E, BOYAULT C, GOVIN J, SADOUL K, CHIOCCA S., CHRISTIANS E, KHOCHBIN S, JOLLY C, VOURC’H C

Heat-shock factor 1 controls genome-wide acetylation in heat-shocked cells.

MOL BIOL CELL

4976 4984

5,558

FROIO E., D’ADDA T, FELLEGARA G., MARTELLA E., CARUANA P., PRUNERI G., PESCI A, RINDI G.

Uterine carcinosarcoma metastatic to the lung as large-cell neuroendocrine carcinoma with synchronous sarcoid granulomatosis.

LUNG CANCER

371 - 377

2,970

FRULLONI L, GABBRIELLI A, PEZZILLI R, ZERBI A, CAVESTRO GM, MAROTTA F, FALCONI M, GAIA E, UOMO G, MARINGHINI A, MUTIGNANI M, MAISONNEUVE P., DI CARLO V, CAVALLINI G

PanCroInfAISP Study Group. Chronic pancreatitis: Report from a multicenter Italian survey (PanCroInfAISP) on 893 patients.

DIGEST LIVER DIS

311 - 317

2,577

FUSI A, COLLETTE S, BUSSE A, SUCIU S, RIETZ A, SANTINAMI M, KRUIT WH, TESTORI A., PUNT CJ, DALGLEISH AG, SPATZ A, EGGERMONT AM, KEILHOLZ U

Circulating melanoma cells and distant metastasis-free survival in stage III melanoma patients with or without adjuvant interferon treatment (EORTC 18991 side study).

EUR J CANCER

3189 3197

4,475

GADDUCCI A, FUSO L, COSIO S, LANDONI F., MAGGINO T, PEROTTO S, SARTORI E, TESTA A, GALLETTO L, ZOLA P

Are surveillance procedures of clinical beneﬁt for patients treated for ovarian cancer? A retrospective Italian multicentric study.

INT J GYNECOL CANCER

367 - 374

1,932

GALETTA D., PETRELLA F., BORRI A., GASPARRI R., SPAGGIARI L.

Vascular catheter mimicking persistent left superior vena cava.

THORAC CARDIOV SURG

185 - 186

0,770

GALIMBERTI V. E., CIOCCA M., LEONARDI M. C., ZANAGNOLO V., BARATELLA P., SARGENTI M., SAHIUM R. C., LAZZARI R., GENTILINI O. D., PECCATORI F. A., VERONESI U., ORECCHIA R.

Is electron beam intraoperative radiotherapy (ELIOT) safe in pregnant women with early breast cancer? In vivo dosimetry to assess fetal dose.

ANN SURG ONCOL

100 - 105

3,898

GALLETTI M., RICCARDO S, PARISI F, LORA C, SAQCENA M.K., RIVAS L., WONG B., SERRA A., SERRAS F., GRIFONI D., PELICCI P. G., JIANG J., BELLOSTA P.

Identiﬁcation of domains responsible for ubiquitin-dependent degradation of dMyc by glycogen synthase kinase 3beta and casein kinase 1 kinases.

MOL CELL BIOL

3424 3434

5,942

263 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

GANDINI S., RAIMONDI S., GNAGNARELLA P., DORE’ JF, MAISONNEUVE P., TESTORI A.

Vitamin D and skin cancer: a meta-analysis.

EUR J CANCER

634 - 641

4,475

GANDINI S., DE FRANCESCO L., JOHANSSON H. A., BONANNI B., TESTORI A.

Why vitamin D for cancer patients?

ECANCERMEDICALSCIENCE

160

GARCIA ETIENNE C. A., BARILE M., GENTILINI O. D., BOTTERI E., ROTMENSZ N., SAGONA A., FARANTE G., GALIMBERTI V. E., LUINI A., VERONESI P., BONANNI B.

Breast-Conserving Surgery in BRCA1/2 Mutation Carriers: Are We Approaching an Answer?

ANN SURG ONCOL

3380 3387

3,898

GARGIULO G., MINUCCI S.

Epigenomic proﬁling of cancer cells.

INT J BIOCHEM CELL B

127 - 135

4,178

GARGIULO G., LEVY S, BUCCI G., ROMANENGHI M., FORNASARI L, BEESON KY, GOLDBERG SM, CESARONI M., BALLARINI M., SANTORO F., BEZMAN N, FRIGE’ G., GREGORY PD, HOLMES MC, STRAUSBERG RL, PELICCI P. G., URNOV FD, MINUCCI S.

NA-Seq: a discovery tool for the analysis of chromatin structure and dynamics during differentiation.

DEV CELL

466 - 481

12,882

GENTILINI O. D., ZURRIDA S.

Awareness is the key to conquering male breast cancer

SOUTH MED J

102

736 - 737

1,058

GENTILINI O. D., BOTTERI E., ROTMENSZ N., DA LIMA L., CALISKAN M., GARCIA ETIENNE C. A., SOSNOVSKIKH I., INTRA M., MAZZAROL G., MUSMECI S., VERONESI P., GALIMBERTI V. E., LUINI A., VIALE G., GOLDHIRSCH A., VERONESI U.

Conservative surgery in patients with multifocal/multicentric breast cancer

BREAST CANCER RES TR

113

577 - 583

5,684

GENTILINI O. D., BOTTERI E.

Re: The survival impact of the choice of surgical procedure after ipsilateral breast cancer recurrence.

AM J SURG

198

461 - 462

2,605

GERBINO E., TAPINASSI C., MALAZZI O., MICUCCI C., CALASANZ MJ, BELTRAN HEREDIA JM, GASPARINI P., ODERO MD, PELICCI P. G., BELLONI E. C.

A novel t(7;13)(p12;q33 approximately q34) in AML-M2.

CANCER GENET CYTOGEN

195

198-200

1,482

GIANNI L, GELBER S, RAVAIOLI A, PRICE K N, PANZIN I, FANTINI M., CASTIGLIONE-GERTSCH M, PAGANI O, SIMONCINI E, GELBER R D, COATES A S, GOLDHIRSCH A.

Second non-breast primary cancer following adjuvant therapy for early breast cancer: a report from the International Breast Cancer Study Group.

EUR J CANCER

561 - 571

4,475

GIANOTTI L, BRAGA M, BIFFI R., BOZZETTI F, MARIANI L., FOR THE GLUTAMITALY RESEARCH GROUP OF THE ITALIAN SOCIETY OF PARENTERAL AND ENTERAL NUTRITION

Perioperative Intravenous Glutamine Supplemetation in Major Abdominal Surgery for Cancer: A Randomized Multicenter Trial.

ANN SURG

250

684 - 690

8,460

GIOBBIE-HURDER A, PRICE K.N, GELBER R. D, INTERNATIONAL BREAST CANCER STUDY GROUP AND BIG 1-98 COLLABORATIVE GROUP, COLLEONI M. A., VIALE G., VERONESI P., PERUZZOTTI G., CORSETTO L. A., GHISINI R., RENNE G., LUINI A., ORLANDO L., TORRISI R., ROCCA A., DE PAS T., MUNZONE E., GALIMBERTI V. E., ZURRIDA S., INTRA M., NOLE’ F., ORECCHIA R., MARTINELLI G., DE BRAUD F.

Design, conduct, and analyses of Breast International Group (BIG) 1-98: a randomized, double-blind, phase-III study comparing letrozole and tamoxifen as adjuvant endocrine therapy for postmenopausal women with receptor-positive, early breast cancer.

CLIN TRIALS

272 - 287

1,694

GIORDANO G, FERRUCCI P. F., NICCI C, GRAFONE T, TAMBARO R, PAPINI S, FARINA G, PIANO S, ZAPPACOSTA B, STORTI S

Medium adsorbance fraction of reticulocyte and myeloperoxidase index may individuate a patient subset with a low risk of chemotherapy-related neutropenia.

ONCOL REP

193 - 198

1,524

GOLDHIRSCH A., INGLE J.N., GELBER R.D., COATES A.S., THURLIMANN B, SENN H.-J., PANEL MEMBERS, COLLEONI M. A., VIALE G.

Thresholds for therapies: highlights of the St Gallen International Expert Consensus on the Primary Therapy of Early Breast Cancer 2009

ANN ONCOL

1319 - 1329

4,935

GUERRIERI GONZAGA A., BOTTERI E., ROTMENSZ N., BASSI F. D., INTRA M., SERRANO D., RENNE G., LUINI A., CAZZANIGA M., GOLDHIRSCH A., COLLEONI M. A., VIALE G., IVALDI G. B., BAGNARDI V., LAZZERONI M., DECENSI A., VERONESI U., BONANNI B.

Ductal intraepithelial neoplasia: postsurgical outcome for 1,267 women cared for in one single institution over 10 years.

ONCOLOGIST

201 - 212

6,630

HILGER M, BONALDI T., GNAD F, MANN M

Systems-wide analysis of a phosphatase knock down by quantitative proteomics and phosphoproteomics.

MOL CELL PROTEOMICS

1908 1920

8,834

264 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

HULLEMAN E., QUARTO M, VERNELL R, MASSERDOTTI G, COLLI E., KROS J.M, LEVI D, GAETANI P, TUNICI P, FINOCCHIARO G., RODRIGUEZ Y BAENA R, CAPRA M., HELIN K.

A role for the transcription factor HEY1 in glioblastoma

J CELL MOL MED

136 - 146

5,114

IGIDBASHIAN S., MAGGIONI A., CASADIO C., BOVERI S., CRISTOFORONI P., SIDERI M. G.

Sentinel Pap smears in 261 invasive cervical cancer patients in Italy.

VACCINE

a34 - a38

3,298

ILIEV I. D., SPADONI I., MILETI E., MATTEOLI G., SONZOGNI A. M., SAMPIETRO G.M, FOSCHI D., CAPRIOLI F, VIALE G., RESCIGNO M.

Human intestinal epithelial cells promote the differentiation of tolerogenic dendritic cells.

GUT

1481 - 1489

9,766

ILIEV I. D., MILETI E., MATTEOLI G., CHIEPPA M., RESCIGNO M.

Intestinal epithelial cells promote colitisprotective regulatory T-cell differentiation through dendritic cell conditioning.

MUCOSAL IMMUNOL

340 - 350

JERECZEK B. A., FARISELLI L, BELTRAMO G, CATALANO G., SERAFINI F., GARIBALDI C., CAMBRIA R., BRAIT L, POSSANZINI M., BIANCHI L. C., VAVASSORI A., ZERINI D., ORSI F., DE COBELLI O., ORECCHIA R.

Linac-based or robotic image-guided stereotactic radiotherapy for isolated lymph node recurrent prostate cancer.

RADIOTHER ONCOL

14 - 17

3,990

JERECZEK B. A., ZERINI D., VAVASSORI A., FODOR C., SANTORO L., MINISSALE A. F., CAMBRIA R., CATTANI F., GARIBALDI C., SERAFINI F., MATEI D. V., DE COBELLI O., ORECCHIA R.

Sooner or later? Outcome analysis of 431 prostate cancer patients treated with postoperative or salvage radiotherapy

INT J RADIAT ONCOL

115 - 125

4,639

JERECZEK B. A., CURIGLIANO G., ORECCHIA R.

Systemic therapies for non-metastatic prostate cancer: review of the literature.

ONKOLOGIE

359 - 363

1,545

LA PIETRA L.

Garantire un’ assistenza più sicura: il contributo del medico manage r- le dieci sﬁde che il medico manager è chiamato ad affrontare per il miglioramento della sicurezza dei sistemi sanitari

MEDICI MANAGER

20-27

LA PIETRA L., DERIU P. L.

Ieo quality model

MANAGEMENT DELLA SANITA’

44 - 46

LA PIETRA L

La sanità online: punti di luce da sfruttare

ABC SALUTE

006-12

LA PIETRA L.

Medici manager: i valori che uniscono, le sﬁde che ci attendono

MEDICI MANAGER

30 - 33

LA PIETRA L.

Perdita di documentazione sanitaria: come procedere?

MANAGEMENT DELLA SANITA’

60 - 61

LA ROSA S, KLERSY C, UCCELLA S, DAINESE L, ALBARELLO L, SONZOGNI A. M., DOGLIONI C, CAPELLA C, SOLCIA E

Improved histologic and clinicopathologic criteria for prognostic evaluation of pancreatic endocrine tumors.

HUM PATHOL

30 - 40

3,322

LAM TK, CROSS AJ, CONSONNI D, RANDI G, BAGNARDI V., BERTAZZI PA, CAPORASO NE, SINHA R, SUBAR AF, LANDI MT

Intakes of Red Meat, Processed Meat, and Meat Mutagens Increase Lung Cancer Risk.

CANCER RES

932 - 939

7,514

LANKISCH PG, DOOBE C, FINGER T, LUBBERS H, MAHLKE R, BRINKMANN G, KLOPPEL G, MAISONNEUVE P., LOWENFELS AB

Hyperamylasaemia and/or hyperlipasaemia: Incidence and underlying causes in hospitalized patients with non-pancreatic diseases.

SCAND J GASTROENTERO

237 - 241

1,980

LANKISCH PG, BREUER N, BRUNS A, WEBERDANY B, LOWENFELS AB, MAISONNEUVE P.

Natural History of Acute Pancreatitis: A Long-Term Population-Based Study.

AM J GASTROENTEROL

104

2797 2805

6,444

LANKISCH PG, WEBER-DANY B, MAISONNEUVE P., LOWENFELS AB

Re: Skin signs in acute pancreatitis: frequency and implications for prognosis.

J INTERN MED

266

220

5,412

LANKISCH PG, WEBER-DANY B, MAISONNEUVE P., LOWENFELS AB

Skin signs in acute pancreatitis: frequency and implications for prognosis.

J INTERN MED

265

299 - 301

5,412

LANKISCH PG, KARIMI M, BRUNS A, MAISONNEUVE P., LOWENFELS AB

Temporal Trends in Incidence and Severity of Acute Pancreatitis in Lüneburg County, Germany: A Population-Based Study.

PANCREATOLOGY

420 - 426

3,043

LANKISCH PG, WEBER-DANY B, HEBEL K, MAISONNEUVE P., LOWENFELS AB

The Harmless Acute Pancreatitis Score: A Clinical Algorithm for Rapid Initial Stratiﬁcation of Non-Severe Disease

CLIN GASTROENTEROL H

702 - 705

6,068

LEMOLI RM, SALVESTRINI V, BIANCHI E, BERTOLINI F., FOGLI M, AMABILE M, TAFURI A, SALATI S, ZINI R, TESTONI N, RABASCIO C., ROSSI L, MARTIN PADURA I, CASTAGNETTI F, MARIGHETTI P., MARTINELLI G, BACCARANI M, FERRARI S, MANFREDINI R

Molecular and functional analysis of the stem cell compartment of chronic myelogenous leukemia reveals the presence of a CD34- cell population with intrinsic resistance to imatinib.

BLOOD

114

5191 5200

10,432

265 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

LEO F., RADICE D., DIDIER F., CARVALHO BRAGA DOS SANTOS F., SPACCA E., MADINI M., ESPOSITO C., AGNELLO M., MENEGHIN S., SPAGGIARI L.

Does a personalized approach improve patient satisfaction in thoracic oncology?

AM J MANAG CARE

361 - 367

2,220

LEO F., GALETTA D., BORRI A., SPAGGIARI L.

Segmentectomy for carcinoid arising from an accessory cardiac bronchus.

EUR J CARDIO-THORAC

537

2,181

LIPTROTT S. J., PECCATORI F. A., COCQUIO A., MARTINELLI G.

Communication skills and raising awareness in clinical practice: an Italian experience

ECANCERMEDICALSCIENCE

LISSONI A.A, COLOMBO N., PELLEGRINO A, PARMA G. M., ZOLA P, KATSAROS D, CHIARI S, BUDA A, LANDONI F., PEIRETTI M., DELL’ANNA T, FRUSCIO R, SIGNORELLI M, GRASSI R, FLORIANI I, FOSSATI R, TORRI V, RULLI E

A phase II, randomized trial of neo-adjuvant chemotherapy comparing a three-drug combination of paclitaxel, ifosfamide, and cisplatin (TIP) versus paclitaxel and cisplatin (TP) followed by radical surgery in patients with locally advanced squamous ce

ANN ONCOL

660 - 665

4,935

LOPERA SIERRA M., AGAZZI A., BODEI L., PACIFICI M., ARICO’ D., DE CICCO C., QUARNA J., SANSOVINI M, DE SIMONE M, PAGANELLI G.

Lymphocytic toxicity in patients after peptide-receptor radionuclide therapy (PRRT) with 177Lu-DOTATATE and 90Y-DOTATOC.

CANCER BIOTHER RADIO

659 - 665

1,318

LORIZZO K., FAZIO N., RADICE D., BOSELLI S., ARIU L., ZAMPINO M. G., NOLE’ F., MAGNI E., ARDITO R., MINCHELLA I., ROCCA A., DI MEGLIO G., SQUADRONI M., DE BRAUD F.

Simpliﬁed FOLFIRI in pre-treated patients with metastatic gastric cancer.

CANCER CHEMOTH PHARM

301 - 306

2,740

LOWENFELS AB, MAISONNEUVE P.

Historical perspectives in surgery: The case of the clever clinician with an eponymous injury

MED GEN SURG

LOWENFELS AB, MAISONNEUVE P.

Pancreas: A new model to predict mortality in acute pancreatitis.

NAT REV GASTROENTEROL HEPATOL

203 - 204

LOWENFELS AB, MAISONNEUVE P., SULLIVAN T

The changing character of acute pancreatitis: epidemiology, etiology, and prognosis.

CURR GASTROENTEROL REP

97 - 103

LOYOLA A, TAGAMI H, BONALDI T., ROCHE D, QUIVY J.P, IMHOF A, NAKATANI Y, DENT S.Y.R., ALMOUZNI G

The HP1alpha-CAF1-SetDB1-containing complex provides H3K9me1 for Suv39-mediated K9me3 in pericentric heterochromatin.

EMBO REP

769 - 775

7,099

LUCA F., CENCIARELLI S., VALVO M., POZZI S., LO FASO F., RAVIZZA D., ZAMPINO M. G., SONZOGNI A. M., BIFFI R.

Full robotic left colon and rectal cancer resection: technique and early outcome.

ANN SURG ONCOL

1274 - 1278

3,898

LUINI A., ROSOSCHANSKY J., GATTI G., ZURRIDA S., CALDARELLA P., VIALE G., DOS SANTOS G. R., FRASSON A. L.

The surgical margin status after breastconserving surgery: discussion of an open issue.

BREAST CANCER RES TR

113

397 - 402

5,684

MACIS D., CAZZANIGA M., DE CENSI A., BONANNI B.

Role of traditional and new biomarkers in breast carcinogenesis

ECANCERMEDICALSCIENCE

157

MADDALUNO L., VERBRUGGE S.E, MARTINOLI C., MATTEOLI G., CHIAVELLI A., ZENG Y, WILLIAMS E.D., RESCIGNO M., CAVALLARO U

The adhesion molecule L1 regulates transendothelial migration and trafﬁcking of dendritic cells.

J EXP MED

206

623 - 635

15,219

MAFFINI F. A., BALDINI F., BASSI F. D., LUINI A., VIALE G.

Systemic therapy as a ﬁrst choice treatment for idiopathic granulomatous mastitis.

J CUTAN PATHOL

689 - 691

1,561

MAGGIONI A., ROVIGLIONE G., LANDONI F., ZANAGNOLO V., PEIRETTI M., COLOMBO N., BOCCIOLONE L., BIFFI R., MINIG L. A., MORROW CP

Pelvic exenteration: Ten-year experience at the European Institute of Oncology in Milan.

GYNECOL ONCOL

114

64 - 68

2,919

MAGGIONI A., MINIG L. A., ZANAGNOLO V., PEIRETTI M., SANGUINETI F., BOCCIOLONE L., COLOMBO N., LANDONI F., ROVIGLIONE G., VELEZ J.I

Robotic approach for cervical cancer: Comparison with laparotomy A case control study.

GYNECOL ONCOL

115

60 - 64

2,919

MAGNIFICO A, ALBANO L, CAMPANER S., DELIA D, CASTIGLIONI F, GASPARINI P, SOZZI G, FONTANELLA E, MENARD S, TAGLIABUE E

Tumor-initiating cells of HER2-positive carcinoma cell lines express the highest oncoprotein levels and are sensitive to trastuzumab.

CLIN CANCER RES

2010 2021

6,488

MAISONNEUVE P., IODICE S., LOHR JM, LOWENFELS AB

Re: ABO Blood Group and the Risk of Pancreatic Cancer.

J NATL CANCER I

101

1156 - 1157

14,933

MALLIA A. J., TRAVAINI L. L., TRIFIRO’ G., PAGANELLI G.

Detection of a cardiac mass by [18F]FDGPET/CT: a rare case.

ECANCERMEDICALSCIENCE

152

MANCUSO P., CALLERI A., BERTOLINI F., TACCHETTI C, HEYMACH J V, SHALINSKY D R

Quantiﬁcation of circulating endothelial cells by ﬂow cytometry.

CLIN CANCER RES

3640

6,488

266 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

MANCUSO P., ANTONIOTTI P., QUARNA J., CALLERI A., RABASCIO C., TACCHETTI C., BRAIDOTTI P, WU HK, ZURITA AJ, SARONNI L. M., CHENG JB, SHALINSKY DR, HEYMACH JV, BERTOLINI F.

Validation of a standardized method for enumerating circulating endothelial cells and progenitors: ﬂow cytometry and molecular and ultrastructural analyses.

CLIN CANCER RES

267 - 273

6,488

MANNARINI L., KRATOCHVIL V., CALABRESE L., GOMES SILVA L, MORBINI P, BETKA J, BENAZZO M

Human Papilloma Virus (HPV) in head and neck region: review of literature.

ACTA OTO-LARYNGOL

119 - 126

0,868

MARINELLI A., BOSSI D., PELICCI P. G., MINUCCI S.

Redundant function of retinoic acid receptor isoforms in leukemogenesis unravels a prominent function of genome topology and architecture in the selection of mutagenic events in cancer.

LEUKEMIA

417 - 419

8,634

MARTELLA S., AGAZZI A., PETIT J. Y., BARBOSA F. S., RIETJENS M., LETKUS J., PALERMO BRENELLI F., CALDARELLA P., CHIFU C., LUINI A., CALISKAN M.

Severe skin necrosis after breast reconstruction with a transverse rectus abdominis musculocutaneous ﬂap in methylenetetrahydrofolate reductase deﬁciency

J PLAST RECONSTR AES

e356 e359

1,235

MARTIN PADURA I., BERTOLINI F.

Circulating endothelial cells as biomarkers for angiogenesis in tumor progression

FRONT BIOSCI

304 - 318

3,308

MARTINELLI G., GIGLI F., CALABRESE L., FERRUCCI P. F., ZUCCA E, CROSTA C., PRUNERI G., PREDA L., PIPERNO G., GOSPODAROWICZ M, CAVALLI F, MORENO GOMEZ H

Early stage gastric diffuse large B-cell lymphomas: results of a randomised trial comparing chemotherapy alone versus chemotherapy + involved ﬁeld radiotherapy.

LEUKEMIA LYMPHOMA

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Primary follicular and marginal-zone lymphoma of the breast: clinical features, prognostic factors and outcome: a study by the International Extranodal Lymphoma Study Group.

ANN ONCOL

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The tumor suppressor PRDM5 regulates Wnt signaling at early stages of zebraﬁsh development.

PLOS ONE

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NEK11 regulates CDC25A degradation and the IR-induced G2/M checkpoint.

NAT CELL BIOL

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Eps8 regulates axonal ﬁlopodia in hippocampal neurons in response to brainderived neurotrophic factor (BDNF).

PLOS BIOL

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MILETI E., MATTEOLI G., ILIEV I. D., RESCIGNO M.

Comparison of the immunomodulatory properties of three probiotic strains of Lactobacilli using complex culture systems: prediction for in vivo efﬁcacy.

PLOS ONE

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Early oral versus “traditional” postoperative feeding in gynecologic oncology patients undergoing intestinal resection: a randomized controlled trial.

ANN SURG ONCOL

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MINIG L. A., BIFFI R., ZANAGNOLO V., ATTANASIO A., BELTRAMI C., BOCCIOLONE L., BOTTERI E., COLOMBO N., IODICE S., LANDONI F., PEIRETTI M., ROVIGLIONE G., MAGGIONI A.

Reduction of Postoperative Complication Rate with the Use of Early Oral Feeding in Gynecologic Oncologic Patients Undergoing a Major Surgery: A Randomized Controlled Trial.

ANN SURG ONCOL

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MONICA V, CEPPI P, RIGHI L, TAVAGLIONE V, VOLANTE M, PELOSI G., SCAGLIOTTI GV, PAPOTTI M

Desmocollin-3: a new marker of squamous differentiation in undifferentiated large-cell carcinoma of the lung.

MODERN PATHOL

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Central nervous system metastases in a cohort of metastatic breast cancer patients treated with trastuzumab.

CANCER CHEMOTH PHARM

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MONTAGNA E., VIALE G., ROTMENSZ N., MAISONNEUVE P., GALIMBERTI V. E., LUINI A., INTRA M., VERONESI P., MAZZAROL G., PRUNERI G., RENNE G., TORRISI R., CARDILLO A., CANCELLO G., GOLDHIRSCH A., COLLEONI M. A.

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Daily clinical practice of fresh tumour tissue freezing and gene expression proﬁling; logistics pilot study preceding the MINDACT trial.

EUR J CANCER

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The 70-gene prognosis-signature predicts disease outcome in breast cancer patients with 1-3 positive lymph nodes in an independent validation study.

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MOURIDSEN H, GIOBBE-HURDER A, GOLDHIRSCH A., THURLIMANN B, PARIDAENS R, SMITH I, MAURIAC L, FORBES JF, PRICE KN, REGAN MM, GELBER RD, COATES AS, BIG 1-98 COLLABORATIVE GROUP, COLLEONI M. A., VIALE G., VERONESI P., PERUZZOTTI G., CORSETTO L. A., GHISINI R., RENNE G., LUINI A., ORLANDO L., TORRISI R., ROCCA A., DE PAS T., MUNZONE E., GALIMBERTI V. E., ZURRIDA S., INTRA M., NOLE’ F., ORECCHIA R., MARTINELLI G., DE BRAUD F.

Letrozole Therapy Alone or in Sequence with Tamoxifen in Women with Breast Cancer

NEW ENGL J MED

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PROFESSIONE & CLINICAL GOVERNANCE

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CURR OPIN CELL BIOL

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Safety, tolerability and biological effects of long-term metronomic administration of non-cytotoxic anti-angiogenic agents

ONCOLOGY-BASEL

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Evaluation of a candidate breast cancer associated SNP in ERCC4 as a risk modiﬁer in BRCA1 and BRCA2 mutation carriers. Results from the Consortium of Investigators of Modiﬁers of BRCA1/BRCA2 (CIMBA).

BRIT J CANCER

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Is adjuvant chemotherapy of beneﬁt for postmenopausal women who receive endocrine treatment for highly endocrineresponsive, node-positive breast cancer? International Breast Cancer Study Group Trials VII and 12-93.

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BREAST CANCER RES TR

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PLOS ONE

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DIG LIVER DIS

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EXPERT OPIN THER TAR

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EUR J CLIN MICROBIOL

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Laboratory-based management of microbiological alerts: Effects of an automated system on the surveillance and treatment of nosocomial infections in an oncology hospital

ECANCERMEDICALSCIENCE

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EUR UROL

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Adjuvant chemotherapy for young women with endocrine non-responsive early breast cancer: any issues?

PAN ARAB JOURNAL OF ONCOLOGY

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Breastfeeding after breast cancer: shifting the paradigm.

ECANCERMEDICALSCIENCE

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Cancer and pregnancy: from biology to treatment choices

PAN ARAB JOURNAL OF ONCOLOGY

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INFORMER IN ONCOLOGIA

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FERTIL STERIL

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FIGO COMMITTEE ON GYNECOLOGIC ONCOLOGY. FIGO staging for uterine sarcomas

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FIGO COMMITTEE ON GYNECOLOGIC ONCOLOGY. Revised FIGO staging for carcinoma of the vulva, cervix, and endometrium.

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Robotic surgery: changing the surgical approach for endometrial cancer in a referral cancer center.

J MINIM INVAS GYN

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Clinical and pharmacological phase I evaluation of exherintm (ADH-1), a selective anti ncadherin peptide in patients with n-cadherin expressing solid tumours.

ANN ONCOL

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Nipple-sparing mastectomy: risk of nippleareolar recurrences in a series of 579 cases.

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Nipple sparing mastectomy with nipple areola intraoperative radiotherapy: one thousand and one cases of a ﬁve years experience at the European institute of oncology of Milan (EIO).

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Intra- and Interobserver Agreement and Impact of Arterial Input Selection in Perfusion CT Measurements Performed in Squamous Cell Carcinoma of the Upper Aerodigestive Tract.

AM J NEURORADIOL

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Perfusion computed tomography for monitoring induction chemotherapy in patients with squamous cell carcinoma of the upper aerodigestive tract: correlation between changes in tumor perfusion and tumor volume.

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Circular clamp excision: a new technique for lung metastasectomy.

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Synchronous pleuro - renal solitary ﬁborous tumors: a new clinical-pathological ﬁnding

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EUR J CARDIO-THORAC

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ESPEN Guidelines on Parenteral Nutrition: Central Venous Catheters (access, care, diagnosis and therapy of complications).

CLIN NUTR

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Tolerance and M2 (alternative) macrophage polarization are related processes orchestrated by p50 nuclear factor {kappa}B.

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TUMORI

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Dietary Changes After Breast Cancer in Women Without Hot Flashes: A Simple and Inexpensive Way to Target Tumor and Host?

J CLIN ONCOL

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EUR J CANCER

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PUPPA G, CANEVA A, COLOMBARI R, SONZOGNI A. M., PELOSI G.

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ARCH PATHOL LAB MED

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Tumor deposits are encountered in advanced colorectal cancer and other adenocarcinomas: an expanded classiﬁcation with implications for colorectal cancer staging system including a unifying concept of in-transit metastases.

MODERN PATHOL

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Bone fractures among postmenopausal patients with endocrine-responsive early breast cancer treated with 5 years of letrozole or tamoxifen in the BIG 1-98 trial.

ANN ONCOL

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J HEPATOL

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FancyGene: dynamic visualization of gene structures and protein domain architectures on genomic loci.

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Interleukin-23: Linking Mesenteric Lymph Node Dendritic Cells With Th1 Immunity in Crohn’s Disease.

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Normal-release Oral Morphine Starting Dose in Cancer Patients With Pain

CLIN J PAIN

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Benign splenosis mimicking peritoneal seeding in a bladder cancer patient: a case report

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A phase I- II study of the histone deacetylase inhibitor valproic acid plus chemoimmunotherapy in patients with advanced melanoma

BRIT J CANCER

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SCAND J SURGERY

72 - 75

0,775

ROCCO B. M., JERECZEK B. A., MATEI D. V., VERWEIJ F., SANTORO L., VAVASSORI A., OSPINA RODRIGUEZ J. C., CEDEIRA F., CIOCCA M., ORECCHIA R., DE COBELLI O.

Intraoperative radiotherapy during radical prostatectomy for intermediate-risk to locally advanced prostate cancer: treatment technique and evaluation of perioperative and functional outcome vs standard radical prostatectomy, in a matched-pair analys

BJU INT

104

1624 1630

2,704

ROCCO B. M., ROCCO F

Re: Assessment of early continence after reconstruction of the periprostatic tissues in patients undergoing computer assisted (robotic) prostatectomy: results of a 2 group parallel randomized controlled trial: M. Menon, F. Muhletaler, M. Campos and J

J UROLOGY

181

1500 - 1501

3,952

ROCCO B. M., MATEI D. V., MELEGARI S., OSPINA RODRIGUEZ J. C., MAZZOLENI F., ERRICO G., MASTROPASQUA M. G., SANTORO L., DETTI S., DE COBELLI O.

Robotic vs open prostatectomy in a laparoscopically naive centre: a matched-pair analysis.

BJU INT

104

991 - 995

2,704

ROCCO F, ROCCO B. M.

Surgery Illustrated - Focus on Details Anatomical reconstruction of the rhabdosphincter after radical prostatectomy

BJU INT

104

274 - 281

2,704

RODRIGUEZ FERNANDEZ J., MARTELLA S., TRIFIRO’ G., CALISKAN M., CHIFU C., PALERMO BRENELLI F., BOTTERI E., ROSSETTO F., ROTMENSZ N., RIETJENS M., VERONESI P.

Sentinel node biopsy in patients with previous breast aesthetic surgery.

ANN SURG ONCOL

989 - 992

3,898

ROSSI G, PELOSI G., GRAZIANO P, BARBARESCHI M, PAPOTTI M

A reevaluation of the clinical signiﬁcance of histological subtyping of non--small-cell lung carcinoma: diagnostic algorithms in the era of personalized treatments.

INT J SURG PATHOL

206 - 218

0,879

ROSSI G, PAPOTTI M, BARBARESCHI M, GRAZIANO P, PELOSI G.

Re:Morphology and a limited number of immunohistochemical markers may efﬁciently subtype non-small-cell lung cancer

J CLIN ONCOL

E141-E142

17,157

ROTGER A, TRIFIRO’ G., TRAVAINI L. L., DE CICCO C., PAGANELLI G.

Carcinoma, tuberculosis and elastoﬁbroma in one patient: is (18F) FDG-PET/CT helpful?

REV ESP MED NUCL

22 - 25

ROTTOLI M, BONA S, ROSATI R, ELMORE U, BIANCHI P. P., SPINELLI A, BARTOLUCCI C, MONTORSI M

Laparoscopic rectal resection for cancer: effects of conversion on short-term outcome and survival.

ANN SURG ONCOL

1279 1286

3,898

RUGGIERO T, TRABUCCHI M, DE SANTA F., ZUPO S, HARFE B.D, MCMANUS MT, ROSENFELD MG, BRIATA P, GHERZI R

LPS induces KH-type splicing regulatory protein-dependent processing of microRNA-155 precursors in macrophages.

FASEB J

2898 2908

7,049

RUSTIGHI A, TIBERI L, SOLDANO A, NAPOLI M, NUCIFORO P, ROSATO A, KAPLAN F, CAPOBIANCO A, PECE S., DI FIORE P. P., DEL SAL G

The prolyl-isomerase Pin1 is a Notch1 target that enhances Notch1 activation in cancer.

NAT CELL BIOL

133 - 142

17,774

SANDRI M. T., RIGGIO D., SALVATICI M., PASSERINI R., ZORZINO L., BOVERI S., RADICE D., SPOLTI N., SIDERI M. G.

Typing of human papillomavirus in women with cervical lesions: prevalence and distribution of different genotypes.

J MED VIROL

271 - 277

2,576

SANTAGUIDA S., MUSACCHIO A.

The life and miracles of kinetochores.

EMBO J

2511 - 2531

8,295

SANTORIELLO C, DEFLORIAN G, PEZZIMENTI F, KAWAKAMI K, LANFRANCONE L. M., D’ADDA DI FAGAGNA F., MIONE M.

Expression of H-RASV12 in a zebraﬁsh model of Costello syndrome causes cellular senescence in adult proliferating cells.

DIS MODEL MECH

56 - 67

272 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

SAUER S, ERBA PA, PETRINI M, MENRAD A, GIOVANNONI L, GRANA C. M., HIRSCH B, ZARDI L, PAGANELLI G., MARIANI G, NERI D, DURKOP H, MENSSEN HD

Expression of the oncofetal ED-B-containing ﬁbronectin isoform in hematologic tumors enables ED-B-targeted 131I-L19SIP radioimmunotherapy in Hodgkin lymphoma patients.

BLOOD

113

2265 2274

10,432

SAVINO MT, ORTENSI B., FERRO M, ULIVIERI C, FANIGLIULO D, PACCAGNINI E, LAZZI S, OSTI D., PELICCI G., BALDARI CT

Rai acts as a negative regulator of autoimmunity by inhibiting antigen receptor signaling and lymphocyte activation.

J IMMUNOL

182

301 - 308

6,000

SCHWARTZ G.F., HUGHES K.S, LYNCH H.T, FABIAN C.J., FENTIMAN I.S., ROBSON M.E., DOMCHEK S.M, HARTMANN L.C., HOLLAND R, WINCHESTER D.J., THE CONSENSUS CONFERENCE COMMITTEE, BONANNI B.

Proceedings of the International Consensus Conference on Breast Cancer Risk, genetics, & Risk Management, April, 2007

BREAST J

004-16

2,091

SESSA C, CRESTA S, NOBERASCO C., CAPRI G, GALLERANI E, DE BRAUD F., ZUCCHETTI M, D’INCALCI M, LOCATELLI A, MARSONI S, CORRADINO I, MINOIA C, ZINTL P, GIANNI L

Phase I clinical and pharmacokinetic study of trabectedin and cisplatin in solid tumours.

EUR J CANCER

2116 - 2122

4,475

SESSA C, PEROTTI A, NOBERASCO C., DE BRAUD F., GALLERANI E, CRESTA S, ZUCCHETTI M, VIGANO’ L, LOCATELLI A, JIMENO J, FEILCHENFELDT JW, D’INCALCI M, CAPRI G, IELMINI N, GIANNI L

Phase I clinical and pharmacokinetic study of trabectedin and doxorubicin in advanced soft tissue sarcoma and breast cancer.

EUR J CANCER

1153 - 1161

4,475

SHAKED Y, TANG T, WOLOSZYNEK J, DAENEN LG, MAN S, XU P, CAI SR, ARBEIT JM, VOEST EE, CHAPLIN DJ, SMYTHE J, HARRIS A, NATHAN P, JUDSON I, RUSTIN G, BERTOLINI F., LINK DC, KERBEL RS

Contribution of Granulocyte ColonyStimulating Factor to the Acute Mobilization of Endothelial Precursor Cells by Vascular Disrupting Agents.

CANCER RES

7524 7528

7,514

SICHETTI D, DI BIAGIO K, ANINO N, BUTTA’ R, VENTURI C, BANDIERI E, RIPAMONTI C, TOGNONI G, ROMERO M, GRUPPO DI LAVORO ECAD, OMODEO SALE’ E.

[The ECAD_O project: clinical caring epidemiology of pain in the hospital: the nurses’ contribution]

ASSIST INFERM RIC

73 - 81

SIDERI M. G., CRISTOFORONI P., CASADIO C., BOVERI S., IGIDBASHIAN S., SCHMITT M, GHEIT T, TOMMASINO M

Distribution of human papillomavirus genotypes in invasive cervical cancer in Italy: a representative, single institution case series.

VACCINE

A30 - A33

3,298

SIGNORELLI M, CHIAPPA V, MINIG L. A., FRUSCIO R, PEREGO P, CASPANI G, BATTISTELLO M, COLOMBO N.

Platinum, Anthracycline, and Alkylating Agent-Based Chemotherapy for Ovarian Carcinosarcoma.

INT J GYNECOL CANCER

1142 - 1146

1,932

SIMONETTA M., MANZONI R, MOSCA R, MAPELLI M., MASSIMILIANO L., VINK M., NOVAK B, MUSACCHIO A., CILIBERTO A

The inﬂuence of catalysis on mad2 activation dynamics

PLOS BIOL

E10

12,683

SKENE L, TESTA G., HYUN I, JUNG K.W, MC NAB A, ROBERTSON J, SCOTT C.T, SOLBAKK J.H, TAYLOR P, ZOLOTH L

Ethics report on interspecies somatic cell nuclear transfer research.

CELL STEM CELL

27 - 30

16,826

SMITH A. P., VERRECCHIA A., FAGA’ G. F., DONI M., PERNA D. F., MARTINATO F., GUCCIONE E., AMATI B.

A positive role for Myc in TGFB-induced Snail transcription and epithelial-to-mesenchymal transition

ONCOGENE

422 - 430

7,216

SOBTI R.C., PUSHPINDER K., SATINDER K., ASHOK K. J., SURINDER K. J., JAI K., RAIMONDI S.

Impact of Interaction of Polymorphic Forms of p53 Codon 72 and N-Acetylation Gene (NAT2) on the Risk of Lung Cancer in the North Indian Population.

DNA CELL BIOL

443 - 449

1,988

SPAGGIARI L.

Invited commentary

ANN THORAC SURG

397 - 398

2,689

SPITALERI G., MATEI D. V., CURIGLIANO G., DETTI S., VERWEIJ F., ZAMBITO S., SCARDINO E., ROCCO B. M., NOLE’ F., ARIU L., DE PAS T., DE BRAUD F., DE COBELLI O.

Phase II trial of estramustine phosphate and oral etoposide in patients with hormonerefractory prostate cancer.

ANN ONCOL

498 - 502

4,935

SUN Z, GOLDHIRSCH A., PRICE KN, COLLEONI M. A., RAVAIOLI A, SIMONCINI E, CAMPBELL I, GELBER RD, TOWLER M

Bone Quality Test (BQT) scores of ﬁngernails in postmenopausal patients treated with adjuvant letrozole or tamoxifen for early breast cancer.

BREAST

84 - 88

2,155

TESTA G.

What to do with the Grail now that we have it? iPSCs, potentiality, and public policy.

CELL STEM CELL

358 - 359

16,826

273 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

TESTORI A., DE SALVO G.L., MONTESCO MC, TRIFIRO’ G., MOCELLIN S, LANDI G, MACRIPO’ G, CARCOFORO P, RICOTTI G, GIUDICE G, PICCIOTTO F, DONNER D, DI FILIPPO F, SOTELDO J. A., CASARA D, SCHIAVON M, VECCHIATO A, PASQUALI S, BALDINI F., MAZZAROL G., ROSSI CR, ON BEHALF OF THE ITALIAN MELANOMA INTERGROUP (IMI)

Clinical considerations on sentinel node biopsy in melanoma from an Italian multicentric study on 1,313 patients (SOLISM-IMI).

ANN SURG ONCOL

2018 2027

3,898

TESTORI A., SOTELDO J. A., SANCES D., MAZZAROL G., TRIFIRO’ G., ZONTA M. J., RASTRELLI M., SCHENONE F., VERRECCHIA F.

Cutaneous melanoma in the elderly.

MELANOMA RES

125 - 134

2,336

TESTORI A., RUTKOWSKI P, MARSDEN J, BASTHOLT L, CHIARION SILENI V, HAUSCHILD A, EGGERMONT AM

Surgery and radiotherapy in the treatment of cutaneous melanoma.

ANN ONCOL

VI 22 - VI 29

4,935

THURLIMANN B., PRICE K.N, GELBER R.D., HOLMBERG S.B., CRIVELLARI D., COLLEONI M. A., COLLINS J., FORBES J.F., CASTIGLIONE-GERTSCH M., COATES A.S., GOLDHIRSCH A., INTERNATIONAL CANCER STUDY GROUP, MARTINELLI G., PERUZZOTTI G., ORLANDO L., NOLE’ F., TORRISI R., LUINI A., ORECCHIA R., VIALE G., PECCATORI F. A., DE BRAUD F., COSTA A., ZURRIDA S., VERONESI P., SACCHINI V., GALIMBERTI V. E., INTRA M., VERONESI U.

Is chemotherapy necessary for premenopausal women with lower-risk node-positive, endocrine responsive breast cancer? 10-year update of International Breast Cancer Study Group Trial 11-93.

BREAST CANCER RES TR

113

137 - 144

5,684

TORRISI R.

Preoperative chemo- and endocrine therapy.

CANCER TREAT RES

151

103 - 120

TOSTI G., DI PIETRO A., FERRUCCI P. F., TESTORI A.

HSPPC-96 vaccine in metastatic melanoma patients: from the state of the art to a possible future.

EXPERT REV VACCINES

1513 - 1526

2,979

TRAVAINI L. L., TRIFIRO’ G., PAGANELLI G.

A parathyroid carcinoma within a cold thyroid nodule

ECANCERMEDICALSCIENCE

150

TRAZZI S., PERINI G, BERNARDONI R, ZOLI M, REESE J C, MUSACCHIO A., DELLA VALLE G

The C-terminal domain of CENP-C displays multiple and critical functions for mammalian centromere formation.

PLOS ONE

E5832

TRINEI M., BERNIAKOVICH I., BELTRAMI E., MIGLIACCIO E., FASSINA A, PELICCI P. G., GIORGIO M.

P66Shc signals to age.

AGING

503 - 510

TROVATO C. M., SONZOGNI A. M., RAVIZZA D., PRUNERI G., ROSSI M., DE ROBERTO G., TAMAYO D., VANAZZI A., FIORI G., CROSTA C.

Confocal laser endomicroscopy diagnosis of gastric adenocarcinoma in a patient treated for gastric diffuse large-B-cell lymphoma.

DIGEST LIVER DIS

447 - 449

2,577

TROVATO C. M., SONZOGNI A. M., FIORI G., RAVIZZA D., TAMAYO D., BOTTI F, CARRARA A, ZEFELIPPO A, CONTESSINI-AVESANI E, CROSTA C.

Confocal laser endomicroscopy for the detection of mucosal changes in ileal pouch after restorative proctocolectomy.

DIGEST LIVER DIS

578 - 585

2,577

VAN ESSEN D, ENGIST B, NATOLI G., SACCANI S

Two modes of transcriptional activation at native promoters by NF-kappaB p65.

PLOS BIOL

E1000073

12,683

VAN NOORD C, STRAUS SM, STURKENBOOM MC, HOFMAN A, AARNOUDSE AJ, BAGNARDI V., KORS JA, NEWTON-CHEH C, WITTEMAN JC, STRICKER BH

Psychotropic drugs associated with corrected QT interval prolongation.

J CLIN PSYCHOPHARM

009-15

4,371

VANAZZI A., CREMONESI M., PAGANELLI G., MARTINELLI G.

Hematologic toxicity and double autografting of Stem Cells after myeloablative activities of yttrium-90 ibritumomab tiuxetan

J CLIN ONCOL

1145

17,157

VAVASSORI A., JERECZEK B. A., ZERINI D., DE CICCO L., CAMBRIA R., CATTANI F., GARIBALDI C., CIOCCA M., ORECCHIA R.

Intraprostatic ﬁducial markers: a potential application for ultrasound-guided radiotherapy in prostate cancer

ECANCERMEDICALSCIENCE

143

VERONESI P., GENTILINI O.D., RODRIGUEZ-FERNANDEZ J., MAGNONI F.

Breast conservation and sentinel node biopsy

BREAST

S90-S95

2,155

VERONESI U., GALIMBERTI V. E., PAGANELLI G., MAISONNEUVE P., VIALE G., ORECCHIA R., LUINI A., INTRA M., VERONESI P., CALDARELLA P., RENNE G., ROTMENSZ N., SANGALLI C. A., DE BRITO L. N., TULLII M. V., ZURRIDA S.

Axillary metastases in breast cancer patients with negative sentinel nodes: A follow-up of 3548 cases.

EUR J CANCER

1381 - 1388

4,475

VERONESI U., ZURRIDA S., GOLDHIRSCH A., ROTMENSZ N., VIALE G.

Breast cancer classiﬁcation: time for a change

J CLIN ONCOL

2427 2428

17,157

VERONESI U., ZURRIDA S.

Preserving life and conserving the breast.

LANCET ONCOL

736

13,283

274 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

VERONESI U., ZURRIDA S., VIALE G., GALIMBERTI V. E., ARNONE P., NOLE’ F.

Rethinking TNM: a breast cancer classiﬁcation to guide to treatment and facilitate research.

BREAST J

VERONESI U.

The European Health Care System as Network of Centres of Excellence

INTERNATIONAL CONFERENCE HEALTH CARE POLICY AND FUNDAMENTAL RIGHTS IN EUROPE - RANGONI MACHIAVELLI B., VELO F.

VIALE A., PELICCI P. G.

Awaking stem cells from dormancy: growing old and ﬁghting cancer.

EMBO MOL MED

VIALE A., DE FRANCO F., ORLETH A. N., CAMBIAGHI V., GIULIANI V., BOSSI D., RONCHINI C., RONZONI S., MURADORE I., MONESTIROLI S. V., GOBBI A., ALCALAY M., MINUCCI S., PELICCI P. G.

Cell-cycle restriction limits DNA damage and maintains self-renewal of leukaemia stem cells

VIALE G.

291 - 295

2,091

17 - 34

88 - 91

NATURE

457

51 - 56

31,434

Be precise! The need to consider the mechanisms for CEP17 copy number changes in breast cancer

J PATHOL

219

001-002

5,121

VIALE G.

Giuseppe Viale

AIS, ATTUALITA’ IN SENOLOGIA

004-08

VIALE G.

Integrating molecular profiling, histological type and other variables: Defining the fingerprint of responsiveness to treatment

BREAST

s32 - s36

2,155

VIALE G., ROTMENSZ N., MAISONNEUVE P., BOTTIGLIERI L., MONTAGNA E., LUINI A., VERONESI P., INTRA M., TORRISI R., CARDILLO A., CAMPAGNOLI E., GOLDHIRSCH A., COLLEONI M. A.

Invasive ductal carcinoma of the breast with the “triple-negative” phenotype:prognostic implications of EGFR immunoreactivity.

BREAST CANCER RES TR

116

317 - 328

5,684

VIALE G., ROTMENSZ N., MAISONNEUVE P., ORVIETO E., MAIRANO E., GALIMBERTI V. E., LUINI A., COLLEONI M. A., GOLDHIRSCH A., COATES A.S.

Lack of prognostic signiﬁcance of “classic” lobular breast carcinoma: a matched, single institution series.

BREAST CANCER RES TR

117

211 - 214

5,684

VIALE G., BOTTIGLIERI L.

Pathological deﬁnition of triple negative breast cancer.

EUR J CANCER

005-10

4,475

VILLA F., CAPASSO P, TORTORICI M, FORNERIS F, DE MARCO A, MATTEVI A, MUSACCHIO A.

Crystal structure of the catalytic domain of Haspin, an atypical kinase implicated in chromatin organization.

P NATL ACAD SCI USA

106

20204 20209

9,380

VITOLO V., MILLENDER L.E, QUIVEY J.M., YOM S.S, SCHECHTER N.R., JERECZEK B. A., MILANI F, ORECCHIA R., XIA P

Assessment of carotid artery dose in the treatment of nasopharyngeal cancer with IMRT versus conventional radiotherapy

RADIOTHER ONCOL

213 - 220

3,990

WESTHOFF B, COLALUCA I. N., D’ARIO G, DONZELLI M., TOSONI D., VOLORIO S, PELOSI G., SPAGGIARI L., MAZZAROL G., VIALE G., PECE S.

Alterations of the Notch pathway in lung cancer

PNAS, PROC NATL ACAD SCI

106

22293 22298

9,380

YANG C, HOELZLE M, DISANZA A., SCITA G., SVITKINA T

Coordination of membrane and actin cytoskeleton dynamics during ﬁlopodia protrusion.

PLOS ONE

E5678

ZAMPINO M. G., MAGNI E., LEONARDI M. C., PETAZZI E., SANTORO L., LUCA F., CHIAPPA A., PETRALIA G., TROVATO C. M., FAZIO N., ORECCHIA R., NOLE’ F., DE BRAUD F.

Capecitabine Initially Concomitant to Radiotherapy Then Perioperatively Administered in Locally Advanced Rectal Cancer.

INT J RADIAT ONCOL

421 - 427

4,639

ZAMPINO M. G., MAGNI E., SONZOGNI A. M., RENNE G.

K-ras status in squamous cell anal carcinoma (SCC): it’s time for target-oriented treatment?

CANCER CHEMOTH PHARM

197 - 199

2,740

ZAMPINO M. G., LABIANCA R., BERETTA G.D., MAGNI E., GATTA G, LEONARDI M. C., CHIAPPA A., BIFFI R., DE BRAUD F., WILS J

Rectal cancer

CRIT REV ONCOL HEMAT

160 - 182

4,589

ZANAGNOLO V., MINIG L. A., GADDUCCI A, MAGGINO T, SARTORI E, ZOLA P, LANDONI F.

Surveillance procedures for patients for cervical carcinoma: a review of the literature.

INT J GYNECOL CANCER

306 - 313

1,932

ZANARDI A, PUNTONI M, MAFFEZZINI M, BANDELLONI R, MORI M, ARGUSTI A, CAMPODONICO F, TURBINO L, BRANCHI D, MONTIRONI R, DECENSI A.

Phase I-II trial of weekly bicalutamide in men with elevated prostate-speciﬁc antigen and negative prostate biopsies.

CANCER PREV RES

377 - 384

ZANETTI M, BIOLO G, BIFFI R., DEUTZ N E, GUGLIELMI F W, PALMO A, SINGER P, TETAMO R, MUSCARITOLI M

Reactive oxygen species and antioxidants: implications for clinical nutrition

NUTRITIONAL THERAPY & METABOLISM

62 - 72

275 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Clinical Research Projects in Progress during 2009 Data of trials and patient enrolement are collected and elaborated by the Clinical Trials and Regulatory Activities Ofﬁce

Title

Principal Investigator

Division

Patients enrolled in 2009

Total patients enrolled

Follow-up of patients treated with sentinel node biopsy at the IEO: the Galimberti node-negative patients

Senology

2904

The sentinel node biopsy performed with local anaesthesia in palpable Luini breast tumors with maximum diameter 2.5 cm

Senology

1210

The radio guided sentinel node biopsy at the internal mammary chain: Luini pilot study on patients affected by inﬁltrating breast carcinoma located in the internal mammary quadrants

Senology

663

Radio guided occult lesion localization (ROLL) in breast surgery

Luini

Senology

4900

Low-dose tamoxifen and 4-HPR in early breast cancer

Bonanni

Cancer Prevention & Genetics

214

A phase III study to evaluate Letrozole as adjuvant endocrine therapy for Goldhirsch Colleoni postmenopausal women with receptor (ER and/or PgR) positive tumors

Medicine

276

Randomized phase III trial of Herceptin followed by Taxol plus Herceptin Goldhirsch Nolè versus the combination of Herceptin and Taxol as ﬁrst-line chemotherapy in patients with HER2-overexpressing advanced breast cancer

Medicine

The HOT Study: Hormone replacement therapy opposed by low dose Veronesi Bonanni tamoxifen. A phase III trial of breast cancer prevention with low dose tamoxifen in HRT users

Cancer Prevention & Genetics

478

Breast ductal lavage in subjects with microcalciﬁcations or at risk for breast Bonanni cancer: identiﬁcation of atypical cells and assessment of proliferation

Cancer Prevention & Genetics

460

Efﬁcacy of intraoperative radiotherapy compared to conventional external Veronesi Orecchia radiotherapy to prevent local relapse of breast carcinoma after breast conserving surgery

Senology Radiotherapy

1306

Adjuvant therapy for patients with breast cancer whose tumors are Goldhirsch judged to require cytotoxic therapy (ER-negative and PgR-negative). Low-dose cytotoxics as “anti-angiogenesis treatment” following induction chemotherapy

Medicine

290

A randomized trial of axillary dissection versus no axillary dissection for Veronesi Goldhirsch patients with clinically node negative breast cancer and micrometastases in the sentinel node

Senology Medicine

580

Phase II study with dose ﬁnding of oral Vinorelbine in combination with Nolè Capecitabine in patients with metastatic breast cancer

Day Hospital

BREAST

276 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

HERA: A randomised three-arm multi-centre comparison of 1 year and Goldhirsch 2 years of Herceptin versus no Herceptin in women with HER2-positive primary breast cancer who have completed adjuvant chemotherapy

Medicine

An international multi-centre study of tamoxifen vs anastrozole in Bonanni postmenopausal women with Ductal Carcinoma in Situ (DCIS)

Cancer Prevention & Genetics

100

An international multi-centre study of anastrozole vs placebo in Bonanni postmenopausal women at increased risk o breast cancer (Prevention)

Cancer Prevention & Genetics

Phase II biomarker trial in patients with ER positive breast cancer before Bonanni surgery.

Cancer Prevention & Genetics

250

A multicenter phase I open-label dose-escalation vaccine trial of dHER2 Colleoni protein with AS15 adjuvant in HER2-overexpressing patients with high-risk breast cancer.

Research in Medical Senology

Suppression of Ovarian Function Trial (SOFT) A phase III trial evaluating Goldhirsch the role of ovarian function suppression and the role of Exemestane as adjuvant therapies for premenopausal women with endocrine responsive breast cancer tamoxifen versus ovarian function suppression + tamoxifen versus ovarian function suppression + exemestane

Medicine

Tamoxifen and Exemestane Trial (TEXT) A phase III trial evaluating Goldhirsch the role of Exemestane plus GnRH analogue as adjuvant therapy for premenopausal women with endocrine responsive breast cancer ovarian function suppression + tamoxifen versus ovarian function suppression + exemestane

Medical Oncology

291

A randomized phase II prevention trial in subjects at high risk for hormone Bonanni non-responsive breast cancer.

Cancer Prevention & Genetics

120

A multicenter open label phase I/II trial of the safety and efﬁcacy of the Curigliano dHER2 recombinant protein combined with immunological adjuvant AS15 in patients with metastatic breast cancer over espressing HER2/neu

Medical Oncology

A randomised phase III trial of exemestane vs anastrozole in post Goldhirsch menopausal women with receptor positive primary breast cancer

Medicine

122

A phase II study of pegylated liposomal doxorubicin as preoperative Colleoni treatment for patients >65 years old with locally advanced primary breast cancer

Research in Medical Senology

A randomized phase II study to assess the activity and tolerability of Colleoni two regimens of metronomic oral chemotherapy (methotrexate plus cyclophosphamide and cyclophosphamide plus capecitabine) combined with bevacizumab in advanced breast cancer

Research in Medical Senology

A phase II randomized study to investigate the effects of Bevacizumab on Colleoni cell proliferation and on pCR in patients with locally advanced operable breast cancer

Research in Medical Senology

Role of PET in the prediction of patients candidates to sentinel node biopsy Paganelli after primary treatment for breast cancer

Nuclear Medicine

Double blind study to compare the efﬁcacy of Palonosetron with or without Nolè the use of Dexamethasone on days 2 and 3, in the prevention of nausea and vomiting induced by moderately emetogenic chemotherapy given to female patients with breast cancer

Day Hospital

A Randomized phase II study of SU011248 Versus Standard-of-care for Goldhirsch patients with Previously Treated, advanced, triple receptor negative (ER, PR, and Her2) breast cancer

Medicine

A phase II study of Lapatinib for brain metastases in subjects with ErbB2 Nolè positive breast cancer following Trastuzumab based systemic therapy and cranial radiotherapy

Day Hospital

A phase II study of GW572016 (Lapatinib TM) on cell proliferation in HER- Bonanni Colleoni 2/neu (ErbB2) positive breast cancer before surgery

Chemoprevention - Medical Oncology

A phase II study to evaluate efﬁcacy and tolerability of concomitant Goldhirsch or sequential administration of Bevacizumab with oral vinorelbine and capecitabine in the treatment of advanced breast cancer

Medicine

277 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

A survey of the Breast International Group (BIG) to assess the attitude of Colleoni patients aged less than 35 years, with early breast cancer, toward the risk of loss of fertility related to adjuvant therapies

Research in Medical Senology

New frontiers of breast diagnostic: optical mammography

Breast Imaging

An open label expanded access study of Lapatinib and Capecitabine Nolè therapy in subjects with ErbB2 overexpressing locally advanced or metastatic breast cancer

Day Hospital

A clinical trial for the evaluation of the tolerability of hypofractionated Orecchia accelerated radiotherapy compared to the conventional scheme in the adjuvant treatment of breast cancer after breast conserving surgery

Radiotherapy

242

Study of intermittent letrozole as adjuvant endocrine therapy

Balduzzi

Research in Medical Senology

109

Phase II study of metronomic oral Vinorelbine in advanced breast cancer

Nolè

Day Hospital

ALTTO (Adjuvant Lapatinib and/or Trastuzumab Treatment Optimisation) Colleoni study. A randomized multicentre open-label phase III study of adjuvant lapatinib, trastuzumab, their sequence and their combination in patients with HER2/ErbB2 positive primary breast cancer

Research in Medical Senology

An observational study of cardiac events in patients with HER2 positive Goldhirsch early breast cancer treated with Herceptin

Medicine

Diagnostic surveillance in women at genetic-familial high risk for breast Bellomi cancer

Radiology

A phase III trial evaluating the role of continuous letrozole versus Colleoni intermittent letrozole following 4 to 6 years of prior adjuvant endocrine therapy for postmenopausal women with hormone-receptor positive, node positive early stage breast cancer. Sole study

Research in Medical Senology

A phase II study of metronomic oral chemotherapy with cyclophosphamide Colleoni plus capecitabine combined with bevacizumab and erlotinib (BEXE), plus trastuzumab in HER2/neu positive tumours (BEXET) in advanced breast cancer

Research in Medical Senology

Preoperative endocrine treatment with Letrozole ± Triptorelin in patients Torrisi with ER and PgR positive locally advanced breast cancer

Research in Medical Senology

A randomised, multicentre, open-label, phase III study of neoadjuvant Colleoni lapatinib, trastuzumab, and their combination plus paclitaxel in women with HER2/ErbB2 positive primary breast cancer

Research in Medical Senology

Evaluation of acquired overexpression of HER2/neu on circulating tumor Nolè cells in patients with advanced breast cancer during chemotherapy and assessment of activity of Trastuzumab-based therapy: a phase II trial

Day Hospital

TOP Trial. A randomized phase III clinical trial of Trastuzumab (Herceptin) Nolè optimization in patients with locally advanced and/or metastatic breast cancer over expressing HER2 after a ﬁrst line chemotherapy plus Trastuzumab

Day Hospital

Phase II study of Exemestane dose intensiﬁcation (150 mg/die) in Goldhirsch postmenopausal patients with advanced breast cancer progressing under standard dose exemestane (25 mg/die) given as adjuvant or palliative therapy

Medicine

A pre-surgical phase II study on activity of Metformin on breast cancer Bonanni cells proliferation

Cancer Prevention & Genetics

Multicentre randomized open label study evaluating an anti insulin like Goldhirsch growth factor I receptor (IGF-1R/CD221) monoclonal antibody AVE1642, administered every 4 weeks in combination con fulvestrant (Faslodex) in postmenopausal patients with advanced hormone dependent breast cancer

Medicine

278 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Cassano

A pre-surgical phase II study on activity of Metformin on breast cancer Bonanni cells proliferation

Cancer Prevention & Genetics

144

150

Allogeneic Haematopoietic Cell Transplantation Using a Non-myeloablative Pastano Preparative Regimen of Total Lymphoid Irradiation and Anti-Thymocyte Globulin for Patients with Refractory “Triple Negative” Breast Cancer

HaematoOncology

A phase II randomized study evaluating the role of 8 courses of primary Colleoni chemotherapy versus 4 courses of primary chemotherapy in combination with endocrine therapy in locally advanced breast cancer

Research in Medical Senology

A randomized double blind, multicenter phase 2 trial of Exemestane Goldhirsch (Aromasin) plus Dasatinib versus Exemestane plus placebo in advanced estrogen receptor positive breast cancer after disease progression on non steroidal aromatase inhibitor (NSAI)

Medicine

A randomized multicenter phase II study of the efﬁcacy and safety Goldhirsch of trastuzumab-mcc-dm1 vs. trastuzumab (Herceptin) and docetaxel (Taxotere) in patients with metastatic her2 positive breast cancer who have not received prior chemotherapy for metastatic disease

Medicine

Evaluation of a new educational approach in patients who underwent Simoncini axillary dissection aimed to improve compliance to a rehabilitation program

Fisiotherapy

156

Metronomic Capecitabine plus Docetaxel as ﬁrst line treatment for Nolè metastatic breast cancer patients: a phase II trial

Day Hospital

Randomized trial of diet, physical activity and breast cancer recurrences Bonanni DIANA 5 study

Cancer Prevention & Genetics

GIM8 (OVER): A randomized trial with factorial design comparing Nolè Fulvestrant ± Lapatinib ± Aromatase inhibitors in metastatic breast cancer progressing after aromatase inhibitor therapy

Day Hospital

Randomized placebo controller phase III trial with low dose tamoxifen in Bonanni women with intraepithelial breast neoplasm

Cancer Prevention & Genetics

Phase II study with epirubicin, cisplatin and infusion of ﬂuorouracil Colleoni (ECF) followed by weekly paclitaxel plus metronomic cyclophosphamide ± trastuzumab as preoperative treatment of locally advanced ER e PgR negative breast cancer

Research in Medical Senology

Randomized phase III trial of surgery alone or surgery plus preoperative Spaggiari gemcitabine-cisplatin in clinical early stages (T2N0 T1-2N1 T3N0 e T3N1) non small cell lung cancer

Thoracic Surgery

Validation of low-dose spiral CT for early diagnosis of lung cancer in a Bellomi Spaggiari high risk population

Radiology

7154

Cromogranine A as a marker for the diagnosis and follow-up of NET. The De Braud CROMaNET observational study

Clinical Pharmacology and New Drugs

Phase II trial of neoadjuvant Alimta plus cisplatin followed by surgery and Spaggiari radiation in the treatment of pleural mesothelioma

Thoracic Surgery

Phase III trial evaluating the role of adjuvant pegylated liposomal Colleoni doxorubicin (PLD Caelyx, Doxil) for women (age 66 years or older) with endocrine non responsive breast cancer who are not suitable for being offered a standard chemotherapy regimen

Research in Medical Senology

Budesonide Turbohaler 800 μg b.i.d. versus placebo in high risk patients Veronesi G. with lung nodules of undetermined origin identiﬁed through spiral TC. A randomized phase II study

Thoracic Surgery

202

A phase II study comparing sequential Satraplatin and Erlotinib to single De Braud agent Erlotinib in patients 70 years or older with unresectable stage III or IV NSCLC as ﬁrst line therapy

Clinical Pharmacology and New Drugs

LUNG CANCER

279 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

NGR010: A phase II study of NGR-hTNF administered as single agent De Braud every 3 weeks in patients affected by advanced or metastatic malignant pleural mesothelioma previously treated with no more than one systemic therapeutic regimen

Clinical Pharmacology and New Drugs

A phase I/II study to assess the safety and immunogenicity of recMAGE- De Pas A3+AS15 cancer immunotherapeutic given as adjuvant therapy, with or without adjuvant chemo(-radio) therapy, to patients with MAGE-A3positive Non Small Cell Lung cancer (stage IB, II or III)

Clinical Pharmacology and New Drugs

A double blind, randomised placebo controlled phase III study to assess the De Pas efﬁcacy of recMAGE-A3 + AS15 antigen speciﬁc cancer immunotherapeutic ad adjuvant therapy in patients with resectable MAGE-A3 positive non small cell lung cancer

Clinical Pharmacology and New Drugs

Analysis of the expression of a speciﬁc set of genes and tumor antigens in De Pas patients with non-small cell lung cancer and melanoma

Clinical Pharmacology and New Drugs

A phase III randomized double blind, placebo controlled multicenter study De Braud of ASA404 in combination with Paclitaxel and Carboplatin as ﬁrst line treatment for locally advanced or metastatic (stage IIIb/IV) non small cell lung cancer (NSCLC)

Clinical Pharmacology and New Drugs

A randomised, multicenter phase II study to explore whether biomarkers De Pas correlate with treatment outcome in chemo-naive patients with advanced or recurrent non-squamous, non small cell lung cancer, who receive treatment with bevacizumab (at a dose of either 7.5 mg/kg or 15 mg/kg) in addition to carboplatin-based chemotherapy (gemcitabine or paclitaxel)

Clinical Pharmacology and New Drugs

Evaluation of the accuracy of a serological biomarker (proGRP) in the Sandri differential diagnosis in the monitoring of small cell lung cancer

Laboratory Medicine

Comparison between intensive and minimal follow-up in colorectal Andreoni carcinoma (Dukes B2-C) after treatment with curative intent: GILDA multicentric Italian randomised study.

General Surgery

A phase III trial of preoperative vs postoperative chemotherapy with Fazio Taxotere-Cisplatin-5FU (TCF) in patients with locally advanced operable gastric carcinoma.

Medicine

A phase II trial to evaluate the efﬁcacy and safety of ZD1839 (IRESSA TM) De Braud in combination with Fluorouracil and Oxaliplatin as ﬁrst-line treatment in patients with metastatic colorectal cancer

Clinical Pharmacology and New Drugs

An Open–label Randomized Phase III Study of Intermittent Oral capecitabine De Braud in combination with Intravenous Oxaliplatin (Q 3 W) ( “XELOX”) versus Fluorouracil / Leucovorin as Adjuvant Therapy for patients who have undergone surgery for colon carcinoma UICC/AJCC Stage III (Dukes stage C)

Clinical Pharmacology and New Drugs

Roche sample repository research project in association with protocol De Braud NO16968: An open–label randomized phase III study of intermittent oral capecitabine in combination with intravenous Oxaliplatin (Q3W) (“XELOX”) versus Fluorouracil/Leucovorin as adjuvant therapy for patients who have undergone surgery for colon carcinoma UICC/AJCC Stage III (Dukes stage C).

Clinical Pharmacology and New Drugs

A phase III, randomized, double-blind, placebo-controlled study of SU De Braud 011248 in the treatment of patients with imatinib mesylate (GleevecTM, Glivec®)-resistant or intolerant malignant gastrointestinal stromal tumor

Clinical Pharmacology and New Drugs

A phase II open label study of PTK787/ZK222584 in the treatment of De Braud metastatic gastrointestinal stromal tumors (GISTs) resistant to Imatinib mesylate

Clinical Pharmacology and New Drugs

A randomized three arm multinational phase III study to investigate De Braud Bevacizumab (q3w or q2w) in combination with either intermittent capecitabine plus oxaliplatin (XELOX) (q3w) or ﬂuorouracil/leucovorin with oxaliplatin (FOLFOX-4) versus FOLFOX-4 regimen alone as adjuvant chemotherapy in colon carcinoma

Clinical Pharmacology and New Drugs

COLORECTAL & GASTRIC CARCINOMA

280 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Predictive value of sentinel node analysis obtained through mini-invasive Andreoni vs laparotomic procedure in the staging of initial colorectal cancer

General Surgery

121

A treatment protocol for patients with Gastro-intestinal Stromal Tumor De Braud who are ineligible for participation in other SU011248 Protocols and are refractory to or intolerant of Imatinib Mesylate

Clinical Pharmacology and New Drugs

OPUS: Oxaliplatin and cetuximab in ﬁrst-line treatment of mCRC Open, De Braud randomized, controlled, multicenter phase II study comparing 5-FU/FA plus oxaliplatin (FOLFOX-4) plus cetuximab versus 5-FU/FA plus oxaliplatin as ﬁrst-line treatment for epidermal growth factor receptor-expressing metastatic colorectal cancer

Clinical Pharmacology and New Drugs

Open label randomised multicentre phase III study of adjuvant Fazio chemotherapy in radically resected adenocarcinoma of the stomach or gastroesophageal junction: comparison of a sequential treatment (CPT11+5-FU/LV - TXT+CDDP) versus 5-FU/LV regimen

Medicine

Localized, completely resected, gastrointestinal stromal tumors (GIST) De Pas expressing KIT receptor: a controlled randomized trial on adjuvant Imatinib mesylate (Glivec) versus no further therapy after complete surgery

Clinical Pharmacology and New Drugs

A single arm, multicenter phase II study of RAD001 in patients with De Braud metastatic colorectal adenocarcinoma whose cancer has progressed despite prior therapy with an anti-EGFR antibody (if appropriate), bevacizumab, ﬂuoropyrimidine, oxaliplatin, and irinotecan-based regimens

Clinical Pharmacology and New Drugs

Clinical Study Phase I-II: Prevention of Colorectal Cancer with Allopurinol Crosta in subjects with Adenomatous Polyps. APAC study

Endoscopy

Identiﬁcation and possible prognostic role of circulating tumor cells (CTC) Sandri in peripheral venous blood of patients with locally advanced rectal cancer diagnosed through traditional clinical workout and treated with radical surgical approach (open surgery or mini invasive laparoscopy)

Laboratory Medicine

A multinational randomized, double blind study comparing the efﬁcacy Zampino of aﬂibercept once every 2 weeks vs placebo in patients with metastatic colorectal cancer (MCRC) treated with irinotecan/5-FU combination (FOLFIRI) after failure of an oxaliplatin based regimen

Day Hospital

A randomised trial investigating the role of FOLFOX-4 regimen duration (3 Zampino versus 6 months) and bevacizumab as adjuvant therapy for patients with stage II/III colon cancer

Day Hospital

Surgical sites infections following colorectal cancer surgery. A randomized Andreoni prospective trial comparing standard and advanced antimicrobial dressing containing ionic silver

General Surgery

A blinded randomized phase I/II study of Brivanib Alaninate vs placebo in De Braud combination with Erbitux and Irinotecan in K-Ras wild type subjects with metastatic colorectal cancer

Clinical Pharmacology and New Drugs

Chromoscopy-guided endomicroscopy with the PENTAX EC 3870CIFK/ Crosta EC 3870-CILK confocal colonoscopes for the detection of intraepithelial neoplasia in subjects with long standing ulcerative colitis

Endoscopy

Evaluation of a new reduced volume bowel preparation before Crosta colonoscopy

Endoscopy

Evaluation of colorectal cancer patients in terms of quality of the surgical Andreoni oncological treatment in the time period after surgery

General Surgery

129

Preservation of genito-urinary function in colorectal cancer patients Valvo undergoing surgery through robotic technique

General Surgery

Brachytherapy

378

PROSTATE CANCER

Ultrasound-guided brachytherapy with permanent implant in cancer of Marsiglia the prostate

281 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

3D conformal radiotherapy in prostate cancer

Orecchia

Radiotherapy

207

1536

An open randomised, parallel group, multicentre, phase III study De Cobelli comparing the efﬁcacy of 2 different oral Nolvadex dosing and scheduling regimen (20mg/day versus 20mg/day for 8 weeks and then 20mg/ week) in preventing gynaecomastia induced by oral Casodex 150mg/die monotherapy in prostate cancer subjects

Urology

Diagnostic accuracy of TRIMprobe in prostate cancer. A conﬁrmative De Cobelli study.

Urology

A randomized multicenter phase II trial of Patupilone (EPO906) plus De Braud Prednisone versus Docetaxel (Taxotere) plus Prednisone in patients with metastatic hormone refractory prostate cancer

Clinical Pharmacology and New Drugs

The importance of monitoring of patients with prostate cancer receiving Orecchia hormonal treatment

Radiotherapy

A Phase III, Randomised, Placebo-controlled, Double-blind Study to Assess Nolè the Efﬁcacy and Safety of Once-daily Orally Administered ZD4054 10 mg in Non-metastatic Hormone-resistant Prostate Cancer Patients

Day Hospital

A randomized double blind phase III trial comparing docetaxel combined Nolè with dasatinib to docetaxel combined with placebo in castration resistant prostate cancer

Day Hospital

Multicenter randomized trial of chlorambucil versus chlorambucil plus Martinelli Rituximab versus Rituximab alone in extranodal marginal-zone B cell lymphoma of mucosa associated lymphoid tissue (MALT lymphoma)

HaematoOncology

Multicenter pilot phase II study for use of 90 Y ibritumomab tiuxetan Martinelli (Zevalin) in refractory or relapsed gastric non Hodgkin’s lymphoma previously treated by chemotherapy, adjuvant radiotherapy or immunotherapy with rituximab

HaematoOncology

Pilot phase II study for use of 90Y Ibritumomab Tiuxetan (Zevalin®) in Martinelli refractory or relapsed gastric non Hodgkin’s lymphoma previously treated by chemotherapy with or without antimicrobial therapy for helicobacter pylori

HaematoOncology

A prospective open randomized trial on the efﬁcacy of gonadotropin- Martinelli releasing hormone agonist depot-triptorelin to prevent chemotherapyinduced premature ovarian failure for lymphoma.

HaematoOncology

Comparing two schedules of rituximab maintenance in rituximab- Martinelli responding patients with untreated, chemotherapy resistant or relapsed follicular lymphoma: A randomized phase III trial

HaematoOncology

Phase II Study of VELCADE TM in patients with extranodal marginal Martinelli zone B-cell lymphoma of MALT-type pre treated with one prior systemic therapy regimen

HaematoOncology

Pegﬁlgrastim in the treatment of lymphomas

recurrent or refractory malignant Martinelli

HaematoOncology

A randomized Phase III multicenter study for the treatment of young Martinelli patients with diffuse cell B lymphoma with unfavourable prognosis (IPI 2-3). Dose-dense chemotherapy + rituximab +/- intensive high dose chemoimmunotherapy supported by autologous peripheral stem cell

HaematoOncology

Phase II trial of bortezomib (Velcade) in combination with chlorambucil Martinelli in patients with relapsed/refractory waldenstrom’s macroglobulinemia or small lymphocytic lymphoma. A multi center study

HaematoOncology

A phase II study about the use of intensiﬁed hybrid chemotherapy regimen Martinelli ChLVVP/ABVVP in advanced Hodgkin lymphoma patients

HaematoOncology

Prospective evaluation of the predictive value of PET in patients with Martinelli diffuse large B-cell-lymphoma under R-CHOP-14. A multicentre study

HaematoOncology

LYMPHOMAS

282 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Prospective collection of data in patients with peripheral T-cell lymphoma Laszlo

HaematoOncology

An open-label single arm multicenter phase II trial with Ofatumumab in Martinelli patients with relapsed diffuse large B-cell lymphoma (DLBCL) ineligible for transplant or relapsed after autologous transplant

HaematoOncology

Master protocol for mantle cell lymphoma. A multicenter stratiﬁed phase II Martinelli trial testing Everolimus (RAD001) for the treatment of patients with newly diagnosed and relapsed or therapy resistant mantle cell lymphoma

HaematoOncology

Nonmyeloablative PBSC allografting from HLA matched related Martinelli Pastano donors using ﬂudarabine and/or low dose TBI with disease-risk based immunosuppression, for patients with acute and chronic myeloprolipherative and lymphoprolipherative disorders

HaematoOncology

5 vs 7 days treatment with 2-CDA (cladribine) s.c. in the treatment of Martinelli hairy cell leukemia

HaematoOncology

Clinical phase II trial to evaluate the safety and efﬁcacy of treosulfan based Martinelli conditioning prior to allogeneic haematopoietic stem cell transplantation in patients with haematological malignancies

HaematoOncology

Brachytherapy

364

Randomized phase III study of neoadjuvant chemotherapy followed by Landoni surgery vs. concomitant radiotherapy and chemotherapy in FIGO Ib2, IIa>4cm or IIb cervical cancer

Gynaecology

Role of FDG PET-CT, RM and lymphoscintigraphy in the staging of cervical Travaini cancer in the pelvic lymphonodes

Nuclear Medicine

Diagnostic accuracy and acceptability of a self testing procedure in the Sideri identiﬁcation of high risk HPV cervical infection

Gynaecology

102

1018

A phase II study of AP23573, a mTOR inhibitor, in female adult patients Colombo with recurrent or persistent endometrial cancer

Gynaecology

A multicentre randomized double blind parallel arm two stage study of the Colombo efﬁcacy and safety of AVE0005 (VEGF Trap) administered intravenously every 2 weeks in patients with platinum resistant and topotecan and/or liposomal doxorubicin resistant advanced ovarian cancer

Gynaecology

An open label, multicenter randomized phase III study comparing the Colombo combination of Doxil/Caelyx and Yondelis with Doxil/Caelyx alone in subjects with advanced relapsed ovarian cancer

Gynaecology

A randomized, parallel group, open-label, active controlled multicenter, Colombo phase III trial of Patupilone (EPO0906) versus pegylated liposomal doxorubicin (Doxil/Caelix) in taxane/platinum, refractory/resistant patients with recurrent epithelial ovarian, primary fallopian or peritoneal cancer

Gynaecology

A phase II, open-label randomized multicenter trial of Pazopanib Colombo (GW786034) in combination with Lapatinib (GW572016) compared to Pazopanib monotherapy and Lapatinib monotherapy in subjects with FIGO stage IVB or recurrent or persistent cervical cancer with zero or one prior chemotherapy regimen for advanced/recurrent disease CALYPSO - A multi-national randomized phase III GCIG (Gynecologic Colombo Cancer Inter Group) study comparing pegylated liposomal Doxorubicin (CAELYX®) and Carboplatin vs Paclitaxel and Carboplatin in patients with epithelial ovarian cancer in late relapse (> 6 months)

Gynaecology

LEUKEMIA

GYNECOLOGICAL CANCERS

Pulsed-dose-rate brachytherapy in gynecological cancers and sarcomas

Marsiglia

283 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

A randomised, multicentre, phase III study of Erlotinib (TARCEVA) versus Colombo observation in patients with no evidence of disease progression after ﬁrst line, platinum based chemotherapy for high risk stage I and stage II -IV ovarian epithelial, primary peritoneal, or fallopian tube cancer

Gynaecology

A randomized double blind placebo controlled multicentre trial of Colombo Abagovomab maintenance therapy in patients with epithelial ovarian cancer after complete response to ﬁrst line chemotherapy

Gynaecology

A Double-Blind, Randomized Phase II Study Evaluating the Efﬁcacy and Colombo Safety of Sorafenib Compared to Placebo in Ovarian Epithelial Cancer or Primary Peritoneal Cancer Patients who have achieved a Complete Clinical Response after Standard Platinum/Taxane Containing Chemotherapy

Gynaecology

Clinical evaluation of the CINtec Cytology Dual Stain Kit as well as a reﬂex Sideri test in the triage of women with a Pap cytology result of ASC-US or LSIL. The Palms trial

Gynaecology

Two-arm, randomized, open-label, phase IIIb study investigating the safety Colombo of a 3 hour i.p. infusion of catumaxomab with and without prednisolone premedication in patients with malignant ascites due to epithelial cancer.

Gynaecology

A phase II, double-blind placebo-controlled multicentre randomised study Colombo of ZD4054 plus Carboplatin and Paclitaxel or Placebo plus Carboplatin and Paclitaxel, in patients with advanced ovarian cancer sensitive to Platinumbased chemotherapy

Gynaecology

A phase III study to evaluate the efﬁcacy and safety of Pazopanib Colombo monotherapy versus placebo in woman who have not progressed after ﬁrst line chemotherapy for epithelial ovarian, fallopian tube or primary peritoneal

Gynaecology

A randomized controlled study comparing Synergo System adjuvant De Cobelli hyperthermia treatment in conjunction with Mitomycin C (LHT + MMC) versus BCG immunotherapy (BCG) adjuvant treatment in patients with Superﬁcial Transitional Cell Carcinoma of the Bladder (STCCB).

Urology

Phase II trial of intravescical Gemcitabine in patients with transitional cell De Cobelli carcinoma of the bladder.

Urology

Sutinib either before or after cytoreductive nephrectomy. A phase II trial Nolè in patients with metastatic renal cell carcinoma. GIR 1.

Day Hospital

Medical optimization of TORisel (MoTOR): multicenter phase II evaluation Nolè of Torisel as II-line treatment for metastatic RCC patients progressing after cytokine therapy, tyrosine kinase or angiogenesis inhibitors.

Day Hospital

PEG-Intron versus observation after regional lymph node dissection in Testori AJCC stage III (TxN1-2M0) melanoma patients: a randomized phase III trial

Melanoma

Post operative adjuvant ganglioside GM2-KLH/QS-21 vaccination treatment Testori vs observation after resection of primary cutaneous melanoma (AJCC Stage II, T3-T4N0M0). A 2-arm multicenter randomized phase III trial

Melanoma

A phase II, multicentre, open, randomised, dose ranging study to investigate Testori the efﬁcacy of combination therapy containing Dacarbazine (DTIC) plus low dose Interferon alpha (alfaIFN) plus Thymosin alfa1 versus both DTIC plus Thymosin alfa1 and DTIC plus alfaIFN in patients with advanced-stage metastatic malignant melanoma

Melanoma

Intravenous versus intra-arterial fotemustine chemotherapy in patients Testori with liver metastases from uveal melanoma: a randomized phase III study of the EORTC Melanoma Group

Melanoma

UROLOGICAL CANCER

MELANOMA + SARCOMAS

284 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Multicenter Selective Lymphadenectomy Trial II (MSLT II): A Phase III Testori Multicenter Randomized Trial of Sentinel Lymphadenectomy and Complete Lymph Node Dissection versus Sentinel Lymphadenectomy Alone in Cutaneous Melanoma Patients with Molecular or Histopathological Evidence of Metastases in the Sentinel Node

Melanoma

Phase II study of dacarbazine with anti-vascular endothelial growth Munzone factor antibody (bevacizumab) in patients with unresectable/metastatic melanoma

Medicine

Pilot phase II study of a combined immunotherapy protocol based on oral Testori Rescigno vaccination and direct intratumoral injection of Salmonella Typhi Ty21a (Vivotif) in metastatic cutaneous melanoma patients

Melanoma Experimental Oncology

A phase 3, open label, randomized, comparative study of Ticilimumab and Testori either dacarbazine or temozolomide in patients with advanced melanoma

Melanoma

A multicenter, randomized, double-blind, two-arm, phase III study in Testori patients with untreated stage III (unresectable) or IV melanoma receiving Dacarbazine plus 10 mg/kg of Ipilimumab (MDX-010) vs Dacarbazine with placebo

Melanoma

Prospective study of the natural history of subjects who were entered in Testori the MMAIT-03-001 and MMAIT-04-001 trials and underwent post-operative therapy with BCG plus placebo or BCG plus CanvaxinTM.

Melanoma

Dose deﬁnition and activity evaluation study of the tumor-targeting human De Braud L19IL2 monoclonal antibody-cytokine fusion protein in combination with Dacarbazine in patients with metastatic melanoma.

Clinical Pharmacology and New Drugs

Prospective multicentre study of elettrochemotherapy, for patients with Testori cutaneous and subcutaneous metastases unresponsive to, or ineligible for standard treatments

Melanoma

A phase III clinical trial to evaluate the safety and efﬁcacy of treatment Testori with 2 mg intralesional Allovectina-7 compared to Dacarbazine (DTIC) or Temozolamide (TMZ) subjects with recurrent metastatic melanoma.

Melanoma

A double blind randomized placebo controlled phase III study to assess the Testori efﬁcacy of recMAGE-A3 + AS15 ASCI as adjuvant therapy in patients with MAGE-A3 positive resected stage III melanoma.

Melanoma

Adjuvant immunotherapy with anti-CTLA-4 monoclonal antibody Testori (Ipilimumab) versus placebo after complete resection of high-risk stage III melanoma: A randomized double-blind phase 3 trial of the EORTC melanoma group.

Melanoma

MISCELLANEA

Computer-aided positioning in radiotherapy

Orecchia

Radiotherapy

294

997

Stereotactic radiotherapy in brain and body metastases

Orecchia

Radiotherapy

184

937

Early detection of myocardial cell damage in high-dose chemotherapy Cardinale treated patients through troponine I determination

Cardiology

1616

MR lymph node staging in patients affected by malignant neoplasia of Bellomi oropharynx: comparison of diagnostic efﬁcacy of Magnetic Resonance with Sinerem versus plain MRI

Radiology

Receptor radiotherapy with Dota-D-Phe1-Tyr3-Octrotate labelled with Paganelli 177Lu

Nuclear Medicine

Port washing with no heparin solution: perspective study in patients with Zencovich solid tumors treated with chemotherapy or other treatments

Medical Oncology

218

Radiotherapy and bisphosphonate combined treatment in patients with Orecchia bone metastasis due to breast, prostate or renal cancer. Observational study

Radiotherapy

285 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Phase II study of imatinib mesylate in Chordoma

Clinical Pharmacology and New Drugs

Endoscopic surveillance vs radical surgery after complete polypectomy Andreoni Crosta of malignant polyp

General Surgery Endoscopy

Phase III randomized trial of concomitant radiation, cisplatin and Orecchia tirapazamine (SR259075) versus concomitant radiation and cisplatin in patients with advanced head and neck cancer

Radiotherapy

A phase II study of combined modality treatment with primary Chiesa chemotherapy followed by endoscopic laser resection in intermediate stage laryngeal squamous cell carcinoma

Head & Neck

Talidomide in the treatment of patients with primitive cancer of the liver. Fazio A clinical/biological study

Medicine

A dose ﬁnding pharmacokinetic study of the tumour-targeting human De Braud L19IL2 monoclonal antibody-cytokine fusion protein in patients with advanced solid tumour

Clinical Pharmacology and New Drugs

Locoregional pre-targeted antibody guided radio immuno therapy Paganelli (PAGRIT) with biotinylated antitenascin monoclonal antibody (ST2146), avidin and 90Y labelled biotin in recurrent glioblastoma multiforme: a phase I/II study

Nuclear Medicine

Phase I dose escalation study to determinate the safety, pharmacokinetics De Braud and pharmacodynamics of BMS 582664 in patients with advanced or metastatic solid tumor

Clinical Pharmacology and New Drugs

Phase I dose escalation safety and pharmacokinetic study of SSR244738 De Braud administered as one hour intravenous infusion every three weeks in patients with solid tumors refractory to all known standard anticancer treatments

Clinical Pharmacology and New Drugs

A phase I dosimetric study of 131I-L19-SIP in solid tumors

Paganelli

Nuclear Medicine

Crosta

Endoscopy

301

Catania

Medical Oncology

139

Clinical Pharmacology and New Drugs

General Surgery

A new induction therapy (ThalDoDex) for multiple myeloma. A phase II Martinelli study

HaematoOncology

Early feeding after gynaecologic laparotomy: a randomized controlled Maggioni trial

Gynaecology

193

NGR8 A phase II study of NGR-hTNF administered a single agent every De Braud 3 weeks in patients affected by advanced or metastatic hepatocellular carcinoma (HCC) previously treated with no more than one systemic therapeutic regimen

Clinical Pharmacology and New Drugs

A randomized double-blind placebo-controlled multicenter phase III in Fazio patients with advanced carcinoid tumor receiving Sandostatin LAR® e RAD001 10 mg/d or Sandostatin LAR® and placebo

Medicine

Phase I dose-escalation study of vinﬂunine hard capsules administered De Braud once a day from D1 to D5 and from D8 to D12 every 3 weeks in patients with advanced/metastatic solid tumors who failed standard therapy

Clinical Pharmacology and New Drugs

NGR004: A phase IB study of NGR-hTNF in combination with cisplatin in De Braud patients affected by advanced o metastatic solid tumors

Clinical Pharmacology and New Drugs

Application of confocal endomicroscopy in the staging of pre malignant and neoplastic lesions of the gastrointestinal tract Fears, prejudices, expectations and perceptions of cancer patients deriving from the proposal to participate in a clinical trial.

De Pas

An open label, stratiﬁed, single-arm phase II study of RAD001 in patients De Braud with advanced pancreatic neuroendocrine tumor (NET) after failure of cytotoxic chemotherapy Home enteral nutrition in malnourished patients after major surgery for gastrointestinal malignancy

286 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Bifﬁ

Phase I/II study of the tumor-targeting human L19IL2 monoclonal antibody- De Braud cytokine fusion protein in combination with gemcitabine in patients with advanced pancreatic cancer

Clinical Pharmacology and New Drugs

A phase II, multicenter, randomised, open label study of Velcade, Melphalan, Martinelli Prednisone and Thalidomide (V-MPT) versus Velcade, Melphalan; Prednisone (V-MP) in elderly untreated multiple myeloma patients

HaematoOncology

Phase I/II study of the tumor-targeting human L19TNF alfa monoclonal De Braud antibody-cytokine fusion protein in patients with advanced solid tumors

Clinical Pharmacology and New Drugs

Effect of mechanical bowel preparation with polyethylene glycol plus Andreoni Bifﬁ bowel enema (glycerine 5%) vs bowel enema alone in patients candidates to colorectal resection for malignancy. Prospective randomized trial.

General Surgery

222

A randomized double-blind phase III study of RAD001 10 mg/d plus best Fazio supportive care versus placebo plus best supportive care in the treatment of patients with advanced pancreatic neuroendocrine tumor (NET)

Medicine

Study on the impact of GARDASIL vaccination program within a population Sideri of 18th years old girls.

Gynaecology

196

427

Prevention of atrial ﬁbrillation in patients undergoing thoracic surgery for Cardinale lung cancer (PRESAGE study)

Cardiology

A randomised, double blind, placebo controlled, multicentre, phase III study Nolè of post-operative adjuvant Lapatinib or placebo and concurrent chemo radiotherapy followed by maintenance Lapatinib or placebo monotherapy in high risk subjects with resected squamous cell carcinoma of the Head and Neck (SCCHN)

Day Hospital

An Open-Label, Dose-escalation, Phase I Study to Determine the De Braud Maximum Tolerated Dose, Recommended Dose, Pharmacokinetics, and Pharmacodynamics of the Proteosome Inhibitor CEP-18770 Given Intravenously as Single Agent in Patients With Advanced Solid Tumours or non-Hodgkin’s Lymphomas

Clinical Pharmacology and New Drugs

A phase I pharmacokinetic and pharmacodynamic study of PF-03446962 in De Braud patients with advanced solid tumors

Clinical Pharmacology and New Drugs

Histoscanning for echography based detection of thyroid cancer (“THS”) Bellomi THS 01

Radiology

Phase III, randomized, double blind trial on Vitamin D supplementation for De Pas resected stage II melanoma patients

Clinical Pharmacology and New Drugs

Neoadjuvant Docetaxel plus Cisplatin and 5-Fluorouracil (TPF) followed by Nolè radiotherapy plus concomitant chemo or Cetuximab versus radiotherapy plus concomitant chemo or Cetuximab in patients with locally advanced squamous cell carcinoma of the head & neck. A randomized phase III factorial study

Day Hospital

A dose ﬁnding pharmacokinetic phase Ib/II study of the tumor-targeting De Braud human F16IL2 monoclonal antibody-cytokine fusion protein in combination with doxorubicin in patients with advanced solid tumours

Clinical Pharmacology and New Drugs

A dose-ﬁnding pharmacokinetic phase Ib/II study of the tumor-targeting De Braud human F16IL2 monoclonal antibody-cytokine fusion protein in combination with Paclitaxel in patients with advanced solid tumours

Clinical Pharmacology and New Drugs

Differences in dynamic contrast-enhanced (DCE) MRI measurements using Bellomi Magnevist (Gd-DTPA) and Vasovist in patients with untreated squamous cell head & neck cancer.

Radiology

A multi center randomized blinded efﬁcacy and safety study of pasireotide Fazio LAR versus ocreotide LAR in patients with metastatic carcinoid tumors whose disease-related symptoms are inadequately controlled by somatostatin analogues

Clinical Pharmacology and New Drugs

Eps8 gene and deafness: from mouse to man through genomic and Goldhirsch proteomic study

Medicine

287 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Phase I/II trial with sorafenib in combination with RAD001 administered De Braud orally in patients with advanced solid tumors, selected on the base of molecular targets

Clinical Pharmacology and New Drugs

Allogenic hematopoietic cell transplantation using a non myeloablative Martinelli preparative regimen of total lymphoid irradiation and anti-thymocyte globulin for patients with hematologic malignancies

Ematoncology

An open label single arm trial investigating zalutumumab, a human Nolè monoclonal anti-EGF receptor antibody, in combination with best supportive care , in patients with non curable squamous cell carcinoma of the head and neck who have failed standard platinum based chemotherapy

Day Hospital

Dose ﬁnding study of CAELIX and RAD001 in patients with advanced solid Colombo tumors

Gynaecology

Evaluation of the ability of multiple serum biomarkers in the assessment of Sandri risk of ovarian carcinoma in patients with a pelvic mass

Laboratory Medicine

Evaluation of the perception of pain symptoms in patients undergoing Martinelli haemopoietic stem cell transplantation

Ematoncology

A non interventional observational post authorisation safety study of Martinelli subjects treated with lenalidomide

Ematoncology

An open label single arm phase II study of combination RAD001 and Fazio octreotide LAR in patients with advanced neuroendocrine tumors as ﬁrst line treatment

Medicine

Pan-European registry on emesis. PEER

Medicine

Ematoncology

3.154

34.407

Locatelli

An open-label, multi-centre, dose escalating, phase I/ randomized phase Martinelli II study to investigate the safety and tolerability of RO5072759 given as monotherapy in patients with CD20+ malignant disease TOTALE

288 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

IEO Foundation Better care where research grows To make a donation is a kindness that counts so much for us. Thank you.

European Institute of Oncology Foundation Via Ripamonti 435 20141 Milan T +39.02 57489.798 F +39.02 94379.269 E fondazione@ieo.it W www.ieo.it

To make a donation is simple, it’s enough: • by credit card through our website www.ieo.it • by bank transfer to account no. 098407800383 at Banca Intesa-Sanpaolo – Branch no. 4258, Via G. Ripamonti 205 – 20141 Milan (routing codes: CAB 09545 - ABI 03069 IBAN IT08 C030 6909 5450 9840 7800 383) in the name of “Fondazione IEO” • by bank transfer to account no. 28615243 at Poste Italiane (routing codes: IBAN IT37K0760101600000028615243) in the name of "Fondazione IEO" • by postal giro transfer to account no. 28615243 in the name of Fondazione IEO, Via Ripamonti 435, 20141 Milan.

290 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

For those at the IEO, via Ripamonti Milan, it is possible to make a donation directly at the Foundation front ofﬁce, open Monday-Friday 09.00-17.00. The ofﬁce is in the aisle between the two entrances (Hall A/Hall B)

IEO Foundation.

Better care where research grows.

IEO foundation takes part in the Scientific Report of the Institute as an entity that has contributed to finance research and scholarship programs. The principal statement of the foundation IEO is to continuously guarantee to IEO researchers the opportunity to express, always at their best level, their creativity, experience and dedication. They also are in charge of the care of patients and of every person going to the IEO. That’s why we operate with the Institute as a non-profit, to collect funds and contribute to train and develop in clinical and experimental research. Also the founders of the Institute where committed to the idea of being near people’s health. They’re big Italian groups that find in research, cure and technology innovation, a concrete expression of progress.During the years, many projects proposed by the Foundation IEO have become a reality thanks to the financial aid of companies that had believed in this social mission and had continued to sustain them. Sustaining the Foundation IEO is fundamental for the future of the defeat of cancer and offers the chance to participate and become aware of research progress. How can you help us? Research and care projects are many and all need economic support. As well as purchase of machines and creation of scholarships, IEO is active in projects on improvement of assistance for patients still in hospital and on support after going home. Main ways to make a donation are: • to bestow a scholarship • to sustain a research project • to participate in an investment in hightech machines for research • to promote internal initiatives (events and activities) • to organize manifestations and develop ideas • to contribute to the 5x1000 campaign of the IEO Other ways • IEO libraries • Biobank • didactic materials to contribute to the instruction of IEO operators In Italy, every year, there are 720.000 new cases of cancer and 160.000 deaths. The estimate of experts for the near future is growth. To fight against cancer, it is essential to develop new research. Research applied to care is the only way to discover new methods and to cure those who are ill and then prevent new cases in the community through clear information. In this context, IEO is one of the great players in Italy; it represents a crowded net of specialists trained internally that work around the world and, thanks to them, in the last ten years, Italy has gone ahead in cancer research. Also IEO founder members usually re-invest IEO profits in research. The background is that of a team that believes in itself and thanks to this, year after year, it renews its trust in researchers. There’s a lot work to do. Your kindness is essential.

291 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

Ongoing Grants in 2009 Agenzia Italiana del Farmaco • Temozolomide in the chemo-immuno-therapy of refractory acute leukaemia of adult patients: the TRIAC protocol. (P.I. E. Bonmassar) Association for International Cancer Research • Analysis of the mechanism of tumor-antigen cross-presentation induced by Salmonella typhimurium. (P.I. M. Rescigno) • Role of p31comet in the regulation and adaptation of the spindle assembly checkpoint. (P.I. A. Musacchio) • Targeting the Ndc80 kinetochore subcomplex for cancer inhibitors. (P.I. P. De Wulf) • Rai/Shc C function in GBM stem cells. (P.I. G. Pelicci) • Histone H3 lysine 27 methylation in gliomagenesis. (P.I. G. Testa) • MicroRNA in Lymphomagenesis, a system biology approach. Analysis of miRNAs targets by Quantitatives Proteomics. (P.I. T. Bonaldi) Associazione Italiana per la Ricerca sul Cancro • Evolutionary genomics of cancer. (P.I. F. Ciccarelli) • Marker-driven, patient-tailored optimization of the chemosentitizing effect of anti-angiogenic drugs. (P.I. F. Bertolini) • Multidisciplinarity: integration of surgery and other specialties as a strategy to improve breast cancer cure. (P.I. A. Luini) • Mapping the epigenome in cancer cells: identifying causes, markers and targets for new therapeutic strategies. (P.I. S. Minucci) • The mitotic checkpoint: a fundamental guard against chromosome instability. (P.I. A. Musacchio) • Phenotypic and functional characterization of circulating tumor cells. (P.I. F. Nolè) • Biological and molecular differences between normal and cancer stem cells. (P.I. P.G. Pelicci) • Post-translational modifications of histone deacetylase 1, a potential target for cancer therapy. (P.I. S. Chiocca) • Avidinated radionuclide therapy in clinically occult breast cancer. (P.I. C. De Cicco) • Role of mitochondrial permeability transition in tumorigenesis. (P.I. M. Giorgio) • Dissection of the molecular pathways involved in melanoma migration and metastasis. (P.I. L. Lanfrancone) • Cell cycle control pathways in aging and cancer. (P.I. E. Migliaccio) • Epigenetic disruption in inflammation and inflammation-associated cancer: mechanisms and molecular players. (P.I. G. Natoli) • Role of Numb and Notch in human lung tumorigenesis and assessment of their relevance as clinical biomarkers. (P.I. S. Pece) • Study of a new pathway of tumor antigen loading via gap-junctions between tumor cells and dendritic cells. (P.I. M. Rescigno) • Histone demethylation in carcinogenesis and lineage commitment. (P.I. G. Testa) • The timely execution of exit from mitosis. (P.I. R. Visintin) • SILAC-based Quantitative Proteomics for Functional Analysis of miRNAs cancer-related targets. (P.I. T. Bonaldi) • Genome-wide analysis of transcription factor interplay in myeloid differentiation and acute myeloid leukemia. (P.I. M. Alcalay) • Function of c-Myc in gene regulation and tumorigenesis. (P.I. B. Amati) • Molecular and functional dissection of novel kinetochore protein Cnn1 in Saccharomyces cerevisiae. (P.I. P. De Wulf) • Role of Nucleophosmin in Leukemia development and therapy design. (P.I. E. Colombo) • Molecular bases of the mitotic spindle coupling to polarity cues in asymmetric stem cell division. (P.I. M. Mapelli) • Randomized trial of diet, physical activity and breast cancer recurrences: the DIANA 5 study. (P.I. B. Bonanni) • Defining the interplay between genetic and epigenetic events in inflammation-associated cancer. (P.I. S. Ghisletti) • Molecular and cellular imaging in cancer. (P.I. U. Veronesi) • Characterization of epigenetic mechanisms of transcriptional regulation in differentiation and cancer. (P.I. D. Pasini) Crohn's & Colitis Foundation of America • Study of the defects of TSLP expression in Crohn's disease patient new therapeutic strategies. (P.I. M. Rescigno)

292 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

European Commission • A multidisciplinary approach to determine the structures of protein complexes in a model organism - 3D REPERTOIRE. (P.I. A. Musacchio) • Advancing Clinico-Genomic Trials on Cancer: Open Grid Services for Improving Medical Knowledge Discovery - ACGT. (P.I. G. McVie) • Targeting tumour-vascular/matrix interactions – ANGIOTARGETING. (P.I. F. Bertolini) • Epigenetic treatment of neoplastic disease – EPITRON. (P.I.’s P.G. Pelicci, S. Minucci) • Targeting transcriptional mechanisms in chronic inflammation-associated diseases TRANS-TAR. (P.I. G. Natoli) • Cancer stem cells and asymmetric cell division - ONCASYM. (P.I. S. Pece) • Health-promoting cross-talk between intestinal microbiota and Humans - CROSS-TALK. (P.I. M. Rescigno) • Mucosal dendritic cells in intestinal homeostasis and bacteria-related diseases -DENDROWORLD. (P.I. M. Rescigno) • European federation for systematic stem cell biology – EUROSYSTEM. (P.I. B. Amati) • Genomic instability in cancer and precancer - GENICA. (P.I. P.G. Pelicci) • Dynamic imaging the mucosal immune system – GUT. (P.I. M. Rescigno) • Mucosal protease and their inhibitors in inflammatory bowel disease: from etiopathogenetic insight to innovative therapy - IBDASE. (P.I. M. Rescigno) • Metagenomics of the human intestinal tract - METAHIT. (P.I. M. Rescigno) • Genomic determinants of inflammation: from physical measurements to system perturbation and mathematical modeling - MODEL-IN. (P.I. G. Natoli) • Early and late health risks to normal/healthy tissues from the use of existing and emerging techniques for radiation therapy ALLEGRO. (P.I. R. Orecchia) • Molecular bases of kinetochore-microtubule attachment and their implications for cell cycle control - KINCON. (P.I. A. Musacchio) • Establishing an efficient network for cancer communication in Europe - EUROCANCERCOMS. (P.I. U. Veronesi) • Structural and Functional Characterization of the RZZ Complex. (P.I. A.S. Wehenkel) • A coordinated and structured international post-doctoral program to foster trans-national mobility and independent career of scientists in biomedicine - SIPOD. (P.I. M. Rescigno) • Oncovideos - Developping vocational skills in oncology through e-learning. (P.I. R. Biffi) European Molecular Biology Organization • Role of dendritic cells in the initiation of innate and adaptive immunity to bacteria - Development of new immunotherapy protocols. (P.I. M. Rescigno) Fondazione Giancarla Vollaro • Neoplastic stem cells and innovative therapies. (P.I. P.G.. Pelicci) Human Frontier Science Program Organization • Unveiling role and functional specificity of individual kB sites in NF-kB dependent genes. (P.I. G. Natoli) • A multidisciplinary approach to microtubule-kinetochore attachment. (P.I. A. Musacchio) Istituto Superiore di Sanità • Servizio nazionale di informazione in oncologia. (P.I. F. Nascè) • Rete nazionale per studi clinici e di strutture GMP per le bioterapie dei tumori. (P.I. M. Rescigno) • Network nazionale italiano tumori eredo-familiari (in TEF): creazione di strumenti operativi condivisi per l'assistenza e la ricerca. (P.I. B. Bonanni) • Rete nazionale "Progetto START", stato dell'arte in oncologia. (P.I. F. de Braud) • Rete nazionale telepatologia - TESEO. (P.I. G. Viale) • Rete nazionale sui modelli sperimentali e "facilities" animali. (P.I. G. Bonizzi) • Rete nazionale bioinformatica in oncologia - RNBBIO. (P.I. F. Ciccarelli) • Rete nazionale delle biobanche per l'oncologia. (P.I. S. Pece) • Immunoterapia con il peptide EGFRvIII nel glioblastoma multiforme e in altri tumori solidi.

293 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

(P.I.’s N. Colombo, G. Curigliano, M. Rescigno, G. Viale ) • Nuove molecole e peptidi quali farmaci regolatori del ciclo cellulare e della risposta a chemioterapici nei tumori epiteliali e cutanei. (P.I. L. Lanfrancone) • Identificazione di marcatori per la predizione della risposta a nuovi farmaci antitumorali (inibitori di HDAC, tirosino chinasi e pompe ioniche). (P.I. P.G. Pelicci) • Il microambiente tumorale come bersaglio terapeutico. (P.I.’s B. Amati, P.G. Pelicci, G. Pelicci) • Sviluppo di nuovi farmaci capaci di alterare il microambiente tumorale e ripristinare la risposta immune anti-tumorale. (P.I. F. Bertolini) • Applicazione della chemioterapia alla rimodulazione della risposta immune antitumorale: studio dei meccanismi e "proof of concept" nell'uomo. (P.I. M. Rescigno) • Development of novel molecular approaches for the diagnosis and characterization of genetic lymphedema. (P.I. F. Bertolini) • Therapy-oriented large scale genomic and gene expression analysis in thymomas, mesotheliomas and lung carcinoids. (P.I.’s E. Belloni , F. de Braud, G. Pelicci) • Model systems to identify genes and factors implicated in pathological processess in Ataxia telangiectasia patients. (P.I. S. Minucci) • Studio sieroproteomica. (P.I. M.T. Sandri) Lega Italiana per la Lotta contro i Tumori • Breast cancer chemoprevention with fenretinide: long-term follow-up of a randomized phase III study. (P.I. B. Bonanni) • Diagnosi precoce delle neoplasie del distretto cervico-facciale mediante video autofluorescenza fibroscopica: studio pilota. (P.I. N. Tradati) • Studio randomizzato di fase II per la prevenzione del tumore alla mammella in donne ad alto rischio: focus sui tumori non ormonoresponsivi. (P.I. B. Bonanni) • Studio clinico sugli effetti anti-proliferativi della Metformina e sue implicazioni per la prevenzione del carcinoma mammario. (P.I. B. Bonanni) Ministero della Salute • New murine models of tumors: new avenues for the identification of novel oncogenes and therapeutic strategies. (P.I. G. Testa) • Surrogate markers of antiangiogenic therapy in triple receptor negative breast cancer. (P.I. A. Goldhirsch) • Biological and molecular characterization of cancer stem cells. (P.I. P.G.. Pelicci) • Tumor radioresistance and predictive markers of the response to radiotherapy. (P.I. G. Bonizzi) • Identification and therapeutic targeting of new oncogenetic molecules in human tumors: basic, preclinical and clinical studies. . (P.I. P.G.. Pelicci) • Neoangiogenesis in hepatoma and other tumors: novel therapeutical targeting of endothelial precursors. (P.I.’s M. Alcalay, F. Bertolini) • Perturbation of tissue microenvironment and pre-neoplastic epigenetic alterations: the epigenetically altered cell as a therapeutic target. (P.I.’s L. Lanfrancone, G. Natoli, G. Testa) • Integrated genomic analysis for the identification of genetic markers of breast cancer metastasis. (P.I.’s I. Dellino, S. Minucci, P.G. Pelicci) • Identification of therapeutic targets and development of new anti-tumour therapies. (P.I.’s E. Colombo, A. Musacchio) • Validation of targeted therapies and related biomarkers to predict tumour response. (P.I.’s S. Chiocca, F. de Braud, S. Minucci, S. Pece) • Cancer stem cells, drug resistance and niche in minimal residual disease. (P.I. F. Bertolini) • New protocols and targets for the treatment of brain and other solid tumors. (P.I.’s G. Pelicci, M. Rescigno) • Optimising therapy of locoregional cancer and evaluation of imaging and biomolecular techniques. (P.I.’s C. De Cicco, O. De Cobelli, G. McVie, R. Orecchia, F. Orsi) • Multidimensional characterization of solid tumors. (P.I. G. Viale) • Research of new prognostic and therapy-oriented biomarkers. (P.I. S. Pece) • Detection of the oncologic disease at early stage. (P.I. M.T. Sandri) • Functional studies and target validation on histone acetyl-transferases in cancer biology. (P.I.’s B. Amati, P. De Wulf) • Preclinical and Phase I clinical studies for glioblastoma and melanoma targeting with innovative methods. (P.I. M. Rescigno) • Analytical and clinical validation of biomarkers for non-invasive early diagnosis of lung cancer. (P.I. E. Belloni) • Analytical and clinical validation of functional imaging for early non-invasive cancer diagnosis. (P.I. G. Veronesi) • Phase IB/II trials of antivascolar/immunomodulant new drugs with chemotherapy in patients with solid tumour. (P.I. F. De Braud)

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• Identification and validation of new therapeutic targets. (P.I. M. Alcalay) • A randomized phase III trial of Fenretinide in women at high risk for breast cancer. (P.I. B. Bonanni) • Phase III study of low dose tamoxifen in women with breast intraepithelial neoplasia. (P.I. B. Bonanni) • Genetic determinants of cardiovascular aging and diabetes. (P.I. P.G. Pelicci) • A molecular medicine project: neoplastic diseases in women. (P.I. P.G. Pelicci) • A prevention phase III trial of fenretinide in women at genetic risk for breast cancer. (P.I. B. Bonanni) • A two step protocol for infarcted myocardium repair by engineered bone marrow progentors and cardiac mesangioblasts. (P.I.’s M. Giorgio, F. Bertolini) • Palliative care network for incurable and terminally ill patients in Milan southern area. (P.I. B. Andreoni) • The role of the lymphatic system in inflammatory bowel disease pathogenesis: a novel therapeutic target. (P.I. M. Rescigno) • The role of Polycomb group proteins in oncogenesys and cell reprogramming. Applications in cancer therapy and regenerative medicine. (P.I.’s G. Testa, G. Pruneri) • Genetic and pathological dissection of the DNA damage response induced by the Myc oncogene and of its role in tumor suppression. (P.I. B. Amati) • Study of the mechanisms of resistance to Herceptin in metastatic breast cancer patients: implications for therapy. (P.I. M. Rescigno) • Identification and isolation of normal and tumoral lung stem cells as a tool for the definition of new therapeutic strategies in lung cancer. (P.I. P.G. Pelicci) • Investigations on molecular pathways involved in mammary gland morphogenesis and role of their functional subversion in human breast carcinogenesis. (P.I. S. Pece) • The validation of safety walk round as a risk assesment tool in an Italian clinical setting: The Hematoncologic. (P.I. G. Martinelli) Ministero dell'Istruzione, dell'Università e della Ricerca • Post-genomic structural biology: development of infrastructures for protein crystallography. (P.I. A. Musacchio) • High throughput genotyping of populations for whole-genome mapping of common disease genes. (P.I. P.G. Pelicci) • Identification of novel anti-cancer therapeutics: from genomics to therapy. (P.I. P.G. Pelicci) • Individuazione, caratterizzazione e valutazione preliminare dell'efficacia di strategie farmaceutiche innovative, basate sull'interferenza con vie metaboliche cellulari. Studio del loro uso per la prevenzione e la terapia di malattie ad eziologia virale. (P.I. S. Minucci) National Institutes of Health • A mitochondrial longevity pathway: p66shc. (P.I. P.G. Pelicci) Swiss Bridge • Biological and molecular characterization of cancer stem cells. (P.I. P.G. Pelicci) The Giovanni Armenise Harvard Foundation • Characterization of Cdc14 and Cfi1, two key regulators of mitotic exit. (P.I. R. Visintin) • Quantitative proteomics for the analysis of the epigenetic regulation of gene expression. (P.I. T. Bonaldi) PHARMAS: Ethical Oncology Science S.p.A. • Characterizing the HDAC inhibitors in mammalian model systems. (P.I.’s S. Minucci, S. Chiocca) Pfizer Limited • Services related to Pfizer clinical study A8471001 "A phase I pharmacokinetic and pharmacodynamic study of PF-03446962 in patients with advanced solid tumors". (P.I. F. Bertolini) OTHERS: GE Healthcare Limited • Imaging and targeting cancer stem cells. (P.I. G. McVie) Tethis S.r.l. • Studi sperimentali per testare l'applicabilità e la funzionalità di un dispositivo microfluidico per saggi di Fluorescent In Situ Hybridization (FISH) su modelli di sangue periferico e midollo osseo da paziente. (P.I. F. Bertolini)

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Seminars 2009

January 12 - Aebersold Rudolf (Zurich): “Qualitative Proteomics and Systems Biology”

February 27 – David G. Pina (European Commission): “Marie Curie Actions: Funding opportunities for researchers”

January 13 – Urban Deutsch (Bern): “Chronically Elevated Levels of Angiopoietin-2 Lead to an Altered Vascular Phenotype in Transgenic Mice”

January 16 – Thomas Surrey (Heidelberg): “Reconstitution of microtubule plus-end tracking by EB1 and CLIP-170 in vitro”

March 10 – Lucio Luzzatto (Florence): “Somatic mutations, clonal diseases and cancer”

January 19 – Anton Gartner (Dundee): “The roads to death, uncovering new DNA damage response pathways by C. elegans genetics”

March 20 – Sergio Nasi (Rome): “Inhibiting Myc to cure cancer?”

January 22 – Claudio Sunkel (Porto): “The role of SAC proteins in kinetochore and non-kinetochore signaling”

March 06 – Ruggero Pardi (Milan): “Adhesion-generated signals in cell proliferation, survival and adaptation to stress”

April 03 – Cayetano González (Barcelona): “Setting up asymmetry in Drosophila neural stem cells”

January 26 – Nancy Hynes (Basel): “Targeting receptor tyrosine kinases in breast cancer”

April 09 – Veronique Orian-Rousseau (Karlsrue): “Regulation of Receptor Tyrosine Kinases activation, signalling and internalization via CD44v6 controls angiogenesis and metastasis”

January 29 – Simon J. Boulton (South Mimms): “Restraining recombination”

April 17 – Ann Ehrenhofer-Murray (Duisburg-Essen): “Establishment of silent domains in the yeast genome”

February 04 – Derek M Yellon and Derek Hausenloy (London): “The MPTP as a target for cardioprotection”

April 24 – Miguel Angel del Pozo (Madrid): “Caveolin and Rho/Rac GTPases in membrane trafﬁcking, cell migration and cell growth”

February 05 – Bernd Pulverer (London): “Scientiﬁc Publishing: wherefore, whereof, wherein and whereto?”

February 12 – Yosef Shiloh (Tel Aviv): “The ATM-Mediated DNA Damage Response: Back to DNA Repair” February 16 – Anabelle Decottignies (Brussels): “Mapping of NUclear DNA sequences of MiTochondrial orign, NUMTs, reveals an association with DNA replication origins” February 20 – Alexander A. Mironov (Chieti): “The application of electron microscopy for the study of intracellular transport”

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May 05 – Stefano Biffo (Milan): “Translational Control of Growth and Cancer: eIF6-dependent regulation of growth factor signaling to the translational apparatus” May 11 – Glenn Marshall (Sydney): “Myc oncogenesis and therapy” May 18 – Amos Maritan (Padua): “Physics of proteins” May 21 – Tapas K. Kundu (Bangalore): “Acetylation of histones and histone chaperones in cancer manifestation: implications in therapeutics”

May 22 – Maria Pia Cosma (Naples): “Periodic activation of Wnt signalling: a toggle switch for mouse somatic cell reprogramming” May 25 – Guttorm Haraldsen (Oslo): “Interleukin-33 - a novel intranuclear cytokine involved in vascular integrity”

September 18 – Richard Lundmark (Umeå): “Coordination of cell adhesion and membrane trafﬁcking by GRAF1” September 24 – Akira Shinohara (Osaka): “Control of protein assembly in recombination”

May 26 – Naama Barkai (Rehovot): “Scaling and Robusteness in embryonic development”

September 28 – Staffan Mueller-Wille (Devon): “Heredity, Race, and Disease: Late Medieval and Early Modern Connections”

June 04 – Ted Weinert (Tucson): “Genome Instability in Yeast: A New and Perilous Twist Arising from Nearby Inverted Repeats”

June 11 – Andreas G. Ladurner (Heidelberg): “Identiﬁcation of nuclear receptors for cellular poly-ADP-ribosylation signaling: The macroH2A and Alc1 remodeler story” June 12 – Volker Haucke (Berlin): “Molecular insights into synaptic vesicle cycling” June 18 – Dominique Belin (Geneva): “Signal sequence recognition by the /E. coli/ preprotein translocase” June 22 – Laura Fontana (Rome): “Role of microRNAs in hematopoiesis and cancer” June 23 – Adam J. Dupuy (Iowa City): “The Impact of Cell of Origin on Genetic Selection in Sleeping Beauty Models of T-ALL”

July 09 – Dmitry Bulavin (Singapore): “Wip1 phosphatase in cancer and aging” July 10 – Roberto Sitia (Milan): “Proteostasis and signaling in the early secretory compartment”

August 26 – Kyungjae (KJ) Myung (Bethesda): “Evolutionary conserved pathways suppress genomic instability”

October 02 – Karl Lenhard Rudolph (Ulm): “Telomere dysfunction and DNA damage checkpoints in stem cell aging”

October 06 – Galina Selivanova (Stockholm): “Rescue of the p53 function by small molecules as a strategy to treat cancer” October 08 – Silvia Nicolis (Milan): “Sox2 is required for neural stem cell maintenance by an autochrine mechanism involving Sox2-dependent activation of the Sonic hedgehog gene” October 12 – Shao-En Ong (Cambridge, USA): “Quantitative proteomics for small-molecule target identiﬁcation” October 23 – Hugues de Thé (Paris): “New insights into acute promyelocytic leukemia therapy response” October 26 – Martin Ruthardt (Frankfurt): “Tranlsocations and their aberrant gene product in the pathogenesis and therapy of leukaemia” October 28 – Gabriela Dontu (London): “ER expression in mammary progenitor cells, biological and clinical implications” October 30 – Johanna Ivaska (Turku): “Integrin Trafﬁc in Cancer”

November 10 – Daniel Panne (Grenoble): “Integrating Signals: Structure of the interferon enhanceosome”

September 14 – Inga Igorevna Poletaeva (Moscow): “Genetic approach to animal cognition and the possible ways to enhance this capacity”

November 13 – Jesper Velgaard Olsen (Copenhagen): “Global analysis of cell signaling pathways by quantitative phosphoproteomics”

September 15 – Aoife McLysaght (Dublin): “Recent de novo origin of human protein-coding genes”

November 16 – Wolfgang Zachariae (Dresden): “Control of chromosome segregation in meiosis”

297 IEO — Scientiﬁc Report 2009 — Ongoing research 2010

November 17 – Richard Pestell (Philadelphia): “Breast Cancer Invasion and Metastasis-New Mechanisms” November 23 – Martin Holzenberger (Paris): “The IGF-1 receptor between life span and neuronal death” November 26 – Enrico V. Avvedimento (Naples): “Histone demethylation, DNA oxidation and transcription” November 27 – Atan Gross (Rehovot): “Balancing cell life and death decisions” November 30 – Kyuson Yun (Bar Harbor): “Regulation of normal and cancer stem cells from the nervous system”

December 01 – Óscar Fernández-Capetillo (Madrid): “Exploring the effects of replicative stress on cancer and ageing” December 03 – Agnes Allansdottir (Siena): “Public views on life sciences in Europe” December 09 – Muriel Moser (Gosselies): “Control of T helper differentiation by regulatory T cells in vivo” December 10 – Gustavo Mostoslavsky (Boston): “Democratizing Pluripotency: A single lentiviral STEMCCA for the generation of iPS cells free of exogenous transgenes” December 14 – Luca Gattinoni (Bethesda): “Basic principles guiding the development of new adoptive Immunotherapies” December 15 – Yves Barral (Zurich): “Spatial and temporal control of late mitotic events: How to cut the bag without damaging its content” December 18 – Buzz Baum (London): “Cell shape through the cell cycle” December 21 – Nir Gov (Rehovot): “Pattern formation inside cells and in cell cultures driven by physical forces”

298 IEO — Scientiﬁc Report 2009 — Ongoing research 2010