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Centre for Dermatology and Genetic Medicine


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Centre for Dermatology and Genetic Medicine: A multi-disciplinary research initiative translating basic science discoveries in genetic skin disease into clinical application.

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Contents Dermatology and Genetic Medicine..................................................................................................... 06 Environment........................................................................................................................................................07 International Medical and Scientific Advisory Board (IMSAB)............................................................................................................................. 08 Research Profiles.............................................................................................................................................. 10

Prof Irwin McLean..................................................................................................................................... 12 Prof Irene Leigh......................................................................................................................................... 13 Prof John McGrath................................................................................................................................... 14 Prof Paul Wyatt......................................................................................................................................... 15 Prof Geoffrey Barton............................................................................................................................... 16 Dr Paul Campbell...................................................................................................................................... 17 Dr Sara Brown............................................................................................................................................ 18 Dr Albena Dinkova-Kostova................................................................................................................ 19 Dr Robyn Hickerson................................................................................................................................20 Dr Deena Leslie Pedrioli......................................................................................................................... 21 Prof Charlotte Proby.............................................................................................................................. 22 Dr Aileen Sandilands.............................................................................................................................. 23 Dr Mark Saville........................................................................................................................................... 24 Dr Frances Smith..................................................................................................................................... 25 Dr Andrew Woodland............................................................................................................................ 26

Genomic Sequencing Unit (GSU)............................................................................................................28 Training................................................................................................................................................................. 30 The Schwartz Lecture.................................................................................................................................... 32 Patient Support Activities and Outreach............................................................................................33 Financial Support.............................................................................................................................................34 Contact..................................................................................................................................................................35

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Dermatology and Genetic Medicine The Centre for Dermatology and Genetic Medicine (DGEM) is a cross-college, multidisciplinary research initiative which has brought together highly research active, internationally renowned clinical dermatologists, biologists and geneticists with specialists in drug discovery, medicinal chemistry, biomedical physics and bioinformatics. Within DGEM there is also significant bidirectional collaboration between the National Health Service diagnostic laboratories and the University research groups, which is a prerequisite for both recruitment of patients for research, as well as the rapid translation of genetic discoveries into diagnostic practice. Close links to clinical dermatology and genetics also enables building of genetically-defined case collections to enable future clinical trials of experimental medicines. The inception of this research grouping has already led to formation of several new translational collaborations, a number of which have been awarded programme-level research grants from Medical Research Council, European Research Council, Cancer Research UK, The Wellcome Trust and a number of skin-disease charities such as DEBRA. Many of these research programmes are aimed at developing and delivering small molecule and gene-silencing therapies in skin disease.

Clinical Physics

Chemistry

Biology

Bioinformatics

A multidisciplinary research initiative translating basic science discoveries in genetic skin disease into clinical application.

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Environment DGEM research groups are housed in state-of-the art laboratory facilities in the College of Life Sciences (CLS) situated on the main University of Dundee campus, and the College of Medicine, Dentistry & Nursing (CMDN) at Ninewells Hospital. Our research benefits from the wider support structure of these Colleges which provide centralised services for the scientific community. This includes access to the most advance technologies including, mass-spectrometry-based proteomics, fluorescent microscopy, flow cytometry, compound and siRNA screening and high performance computing. In addition, other centralised support facilities for activities such as biological waste sterilisation and disposal, media preparation, glass washing and sterilisation, DNA sequencing, DNA cloning service and antibody production facility are available. The colleges also provide comprehensive administrative support to their respective PIs and associated staff and PhD Students. Core staffing is in place to assist with financial management of awards and HR-related issues. DGEM is specifically supported by two divisional secretaries, a dedicated centre manager and also laboratory managers.

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International Medical and Scientific Advisory Board (IMSAB) The DGEM International Medical and Scientific Advisory Board (IMSAB) is comprised of five outstanding individuals who are internationally renowned in their respective fields. The IMSAB members are all highly experienced in complementary aspects of dermatology and human genetics, making them ideally placed to help and advise DGEM. The IMSAB members visit the Centre to review progress and meet with the staff and students. They are invited and encouraged to attend the annual DGEM symposium. We are very grateful to the IMSAB members for devoting their time to help facilitate the successful development of DGEM and their contributions are greatly appreciated.

John Dart Dystrophic Epidermolysis Bullosa Research Association (DEBRA), Crowthorne, UK Areas of expertise: Chief Operating Officer of DEBRA International – a leading genetic skin disease patient advocacy organisation
 www.debra.org.uk/contact-debra.html

Professor Chris Griffiths MD FRCP FMedSci University of Manchester, UK
 (Professor of Clinical Dermatology & Associate Dean for Research) Areas of expertise: psoriasis, photodermatology, clinical trials
 www.medicine.manchester.ac.uk/staff/ChrisGriffiths

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Professor John Todd PhD FRS FMedSci Cambridge Institute for Medical Research, UK (Professor of Human Genetics) Areas of expertise: complex trait genetics and autoimmune inflammatory disease
 www.cimr.cam.ac.uk/investigators/todd/index.html

Professor Leena Bruckner-Tuderman MD PhD Freiburg Institute for Advanced Studies; Germany (Professor of Dermatology)
 Areas of expertise: inherited skin blistering diseases, collagen biochemistry & mouse models of human disease
 www.frias.uni-freiburg.de/lifenet/fellows/fellows-leenabruckner-tuderman

Professor Masayuki Amagai MD PhD Department of Dermatology, Graduate School of Medicine, Keio University, Tokyo, Japan (Professor of Dermatology) Areas of expertise: cutaneous immunology, skin barrier and atopic eczema
 www.gcoe-stemcell.keio.ac.jp/english/member/amagai.html

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Research Profiles

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Prof Irwin McLean PhD, DSc, FRS, FRSE, FMedSci Scientific Director DGEM Professor of Human Genetics | w.h.i.mclean@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Dermatology and Genetic Medicine: Treating genetic disease The central focus of my research group is the study of human inherited diseases that affect epithelial cells and epithelial tissues. The epidermis is the largest and most complex epithelial tissue and includes a number of appendages such as hair, nail, sweat glands and sebaceous glands, each of which is a miniature epithelial organ system. For this reason, most of the genetic diseases we work on are within the field of dermatology, although we also have an active interest in the anterior corneal epithelium, oral mucosa and other epithelial tissues. Much of our research to date has involved identification of the causative genes for epithelial disorders. These studies used a combination of candidate gene analysis focusing on molecules of the keratin intermediate cytoskeleton and its attachment or modification proteins, as well as a positional cloning approach using genomewide genetic linkage analysis and recently, next generation sequencing methodology (service provided by the Genomic Sequencing Unit which is described on pages 28-29 in this brochure). In complex trait genetics, we used a combination of candidate gene, genetic linkage and casecontrol analysis methods to identify the gene encoding filaggrin as the first definitive predisposing gene for atopic eczema, the most common skin disorder with a strong genetic component. Filaggrin mutations not only predispose individuals to atopic eczema but also to atopic asthma, hay fever and other allergies, including peanut allergy. This work showed for the first time that antigen/allergen/irritant exposure via a defective skin barrier is the key first step in triggering a whole range of allergic conditions.

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In recent years our main focus has been targeted towards therapy development. Specifically, we are developing RNA-interference therapy systems for the inherited skin blistering disorder epidermolysis bullosa simplex (EBS) and also for pachyonychia congenita (PC), a rare keratin disorder where painful thickening and blistering of the soles of the feet is highly painful for those affected. In the eye, we are developing siRNA therapy for corneal dystrophy. In addition to siRNA therapeutic strategies, we have a number of advanced small molecule drug discovery programmes. One of these is aimed at improving expression of the filaggrin gene in the very common skin conditions ichthyosis vulgaris and atopic eczema.

Atopic eczema - filaggrin

EB Simplex - keratins K5/K14

Protein expression in skin: filaggrin (green), K5 (red), nuclei (blue).


Prof Irene Leigh CBE, DSc, FRCP, FRSE, FMedSci Clinical Director DGEM Professor of Cellular and Molecular Medicine | i.m.leigh@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Keratinocyte biology and clinical networks The focus of my research has centred around all aspects of keratinocyte biology from studying basic epithelial differentiation into disease applications, particularly the molecular mechanisms of skin cancer, genetic disease and tissue engineering. I have directed the Cancer Research UK Skin Tumour Laboratory since 1983, which continues as a joint London-Dundee activity identifying genetic changes and molecular mechanisms in squamous cell carcinogenesis in both immunosuppressed and immunocompetent patients. Previous research carried out by my group identified the genetic basis of multiple genodermatoses, some with sensorineural deafness and cardiomyopathy. This work is now leading to research in therapeutic developments through Dermatology and Genetic Medicine (DGEM). a

keratinocytes

gel

medium b

More recently I have been instrumental in initiating BADGEM (the British Association of Dermatologists Dermatology & Genetic Medicine) which is a newly formed UK-wide clinical network dedicated to rare genetic diseases of the skin. BADGEM is registered as a company and I am currently acting as the Director. An executive committee has been convened to guide BADGEM and determine its projects, monitor performance and coordinate activities for the best outcomes for UK patients with rare genetic diseases of the skin. To date three subgroups of BADGEM have been established and are focused on the development of: (i) informatics infrastructure for clinical registers; (ii) diagnostic and clinical signposting; and (iii) clinical trials for genodermatoses. One of the initial objectives of BADGEM includes the creation of a national registry for genodermatoses. The database will be developed and hosted on a secure NHS server by the Health Informatics Centre (HIC) in Dundee and start-up funding is being provided by the Wellcome Trust strategic grant awarded to DGEM. The British Association of Dermatologists (BAD), which I am currently the Academic Vice President for, is also helping to facilitate this project.

grid c

(a) Organotypic culture set-up. Comparison of organotypic culture grown skin equivalent (b) to normal skin (c) using H&E staining.

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Prof John McGrath MD, FRCP, FMedSci Professor of Molecular Dermatology | john.mcgrath@kcl.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Discovery and Recovery: Improving the health of people living with inherited skin diseases One of the key research themes of my group is to discover the molecular basis of inherited skin diseases (especially fragile skin disorders). Using a combination of candidate gene approaches, genetic linkage and next generation sequencing (whole exome sequencing), we have identified 12 novel conditions, including the first inherited disorders of hemidesmosome and desmosome cell junctions. Recently, we discovered novel skin fragility disorders resulting from mutations in the Rab-GTPase associated protein, exophilin-5, and the first germline mutations in EGFR (epidermal growth factor receptor) that provide novel insights into skin biology and disease.

We are testing clinical applications of intradermal fibroblasts and intravenous mesenchymal stromal cells in patients with recessive dystrophic EB, and are involved in international efforts to explore how bone marrow progenitor cells can improve skin wound healing. We are also looking to exploit the phenomenon of revertant mosaicism (also known as natural gene therapy) through punch grafting, making sheets of reverted cultured keratinocytes, and in regenerative medicine by making iPS cells that can be used to generate progenitor skin repair cells without the need for additional gene therapy or genomic editing.

My group has been able to use its expertise in mutation detection to set up a national diagnostic laboratory for the skin fragility disorder, epidermolysis bullosa (EB). This work involves skin immunohistochemistry, transmission electron microscopy, and Sanger sequencing of 18 genes implicated in the pathophysiology of EB. Thus far, diagnostic services have been provided to over 1500 families. We have also developed DNA-based prenatal testing for genodermatoses in the UK, and designed, had approved, and applied novel preimplantation genetic diagnosis/ haplotyping tests for couples at risk of recurrence of severe inherited skin diseases. Considerable translational research efforts are underway in my group to develop gene, protein, cell and drug therapies for patients with inherited skin diseases, with a particular focus on cell-based therapies.

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Transmission electron microscopy of spinous layer of epidermis in an individual with ectodermal dysplasia - skin fragility syndrome showing widening of intercellular spaces between keratinocytes as well as a reduced number of small desmosomes.


Prof Paul Wyatt PhD Head of Drug Discovery Unit, Professor of Drug Discovery | p.g.wyatt@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Translation of novel biology through small molecule drug discovery The Drug Discovery Unit (DDU) tackles unmet medical needs through innovative small molecule drug discovery, bridging the gap between academic scientific research and commercial drug discovery and development. The Unit, which currently has 75 dedicated staff with > 580 years of collective industrial experience, is the only fully operational and integrated drug discovery team in the UK university system working across multiple therapeutic areas. It contains all the disciplines required to produce drug leads and pre-clinical drug candidates: compound profiling, medicinal and computational chemistry, structural biology, pharmacokinetics and in vivo efficacy. The DDU collaborates with partners at a local, national and international level to identify lead compounds, potential drug targets and novel tools and approaches to develop improved treatments for a wide range of debilitating and deadly diseases. Within DDU, I have overall responsibility for the Unit’s direction, strategy, scientific quality, delivery of outputs to milestones and building collaborations.

One of our most exciting programmes is the cutaneous drug discovery group (CDDG) which is embedded within both the DDU and DGEM. The CDDG been established to address the unmet medical need for dermatology translational research. This unique initiative builds on our expertise in cutaneous drug design, anti-drug design, understanding of exposurepharmacodynamic relationships, and topical formulations. The cutaneous drug discovery portfolio (CDDP) progresses early stage projects bridging the translation gap and generating high quality data packages suitable for partnerships with pharmaceutical development organisations. The CDDP projects are managed on a day-to-day basis by our ITP Portfolio Manager, Dr Andrew Woodland (page 26 of this brochure).

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Prof Geoffrey Barton PhD, FSB Head of Computational Biology, Professor of Bioinformatics | g.j.barton@dundee.ac.uk

Clinical Physics

Biology

Chemistry

Bioinformatics

Computational analysis of biological systems Key research themes in my group are the analysis of large collections of biological data and the development of tools and techniques to predict the properties of proteins and other biomolecules from their sequence and related information. The analysis of sequences relies on techniques for sequence alignment and is carried out by virtually all scientists interested in the functions of genes and proteins. Working with alignments requires powerful, but easy-to-use interactive tools to manipulate and visualize the data. Our Jalview (www.jalview.org) open-source, GPL-licenced multiple sequence alignment editor and analysis workbench is widely regarded as the de facto standard tool for these tasks. We are continuing to develop Jalview to meet the challenge of working with alignments containing tens of thousands of protein sequences, genome-length alignments and RNA editing, while also extending the range of sequence analysis services available to the program through our new “JABAWS” web services framework. Visualisation Alignment

Interactive Editing

Analysis www.jalview.org

Data Integration

Prediction Services provided by JAVA BIOINFORMATICS ANALYSIS WEB SERVICES

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Figure Generation

As a group we do not do conventional laboratory experiments. For this reason we collaborate extensively with “wet lab” scientists who can either test our predictions, or provide datasets that we analyse in new ways. In this way our complementary skills often enhance understanding of biology while also leading to improved predictive methods. Over the last five years we have developed a lot of experience in analysing the results of highthroughput DNA sequencing technologies, known as “Next Generation Sequencing” (NGS) methods, both as part of DGEM projects and work outside of this group. NGS is able to determine millions of DNA sequences of up to 150 base pairs long in a single experimental run. In order to address the needs and requirements for DGEM, we have been developing techniques for the analysis of whole exome datasets to enable resolution of unsolved monogenic skin conditions. In addition, we have been addressing the analysis of repetitive genes and homologous gene families in complex disease traits such as eczema. As NGS data on its own is not very tractable for the majority of end-users unless they have access to bioinformatics expertise, we are also implementing a Galaxy-based system to deliver data and make certain analyses (e.g. RNA-seq) more accessible. This will be of benefit to all users of the Genomic Sequencing Unit (described on pages 28-29 in this brochure).


Dr Paul Campbell

PhD

Reader in Physics | p.a.campbell@dundee.ac.uk

Clinical Physics

Biology

Chemistry

Bioinformatics

The application of physics in biomedicine The cell membrane in human cells performs the remarkable task of regulating all molecular traffic into and out of the cytosol - and does this to such a refined extent, that water soluble therapeutic species (i.e. many drug formulations) will not readily traverse into the cytoplasm where they can then elicit some intended bioeffect. This is a key challenge in drug delivery. My group seeks to exploit the power of ultrasound and lasers for non-invasive molecular delivery, with a particular emphasis on delivering therapeutics for the treatment of skin diseases. In the context of ultrasound as a potential therapy, it is feasible to introduce microscopic bubbles surrounded by a thin lipid shell into the blood circulation. These ‘microbubbles’, which can carry both therapeutic molecules embedded within the shell, as well as chemical tethers to target and accumulate at specific tissues, are then stimulated to react to the periodic pressure wave as the ultrasound pulse traverses their locale. Vigorous bubble responses generate shear flows and other more exotic fluid structures that have the potential to transfer momentum on a microscopic scale, thus permeabilizing local tissues in a controllable manner, and underscoring the potential of this technology to deliver drugs in a non-invasive fashion.

We have developed significant expertise in the controlled observation of such microscopic/ microsecond interactions, as well as the production of the microbubble constructs themselves using in-house designed microfluidic technologies. We are presently engineering our next-generation microbubble to include siRNA as the molecular payload, with a view to targeting specific skin diseases. In addition, we are also active in the development of novel instrumentation for imaging at refined spatial and temporal scales. Here, we have developed the concept for an ultrafast framing camera technology that effectively doubles the temporal resolution of existing hardware within the MHz regime.

Microbubbles made of lipids, 5-10 microns in diameter, can be used to deliver drugs into the skin using ultrasound.

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Dr Sara Brown MBChB, MRCP, MD Wellcome Trust Intermediate Clinical Fellow, Clinical Senior Lecturer & Hon Consultant Dermatologist | s.j.brown@dundee.ac.uk Clinical Physics

Biology

Chemistry

Bioinformatics

Genetic mechanisms in atopic eczema Atopic eczema is an itchy inflammatory skin disease affecting 1 in 4 school-age children in the UK. It is commonly associated with other atopic illnesses including asthma, hay fever and food allergy. Eczema is a complex disorder resulting from the interplay of multiple genetic and environmental effects. In 2006 the McLean laboratory and collaborators identified loss-of-function mutations in the gene encoding filaggrin (FLG) that greatly increase the risk of atopic eczema. Filaggrin is expressed in the outer layers of skin and this discovery has placed new emphasis on the role of skin barrier function in the pathogenesis of atopic eczema. Approximately 40% of patients with severe atopic eczema carry one or more FLG null mutations. A fuller understanding of genetic mechanisms in eczema would clarify the pathogenesis of this complex trait and facilitate the development of novel treatments.

Skin from areas without active inflammation was sampled in order to study the underlying molecular mechanisms leading to atopic inflammation. The data generated by whole transcriptome analysis was stratified by FLG genotype. This revealed a network of stress response genes up-regulated in the FLG mutation carriers and dysregulated lipid metabolic pathways in eczema cases independent of FLG mutations. The strong effect of FLG on eczema represents a tool which is now being applied to stratify clinical trials of novel eczema treatments. The detailed insight provided by my skin transcriptome analysis offers many opportunities for future therapy development.

My work aims to identify genetic factors affecting FLG expression in human skin and to define other genes involved in the development of eczema and atopic disease. We have shown that individuals carrying FLG mutations have a 3-times increased risk of developing peanut allergy. This finding demonstrates the importance of a functional skin barrier to prevent the development of allergic disease. I have also shown that copy number variation within the FLG gene contributes to eczema risk in a dose-dependent way, indicating that treatments aimed at increasing filaggrin expression may be useful therapeutically. I have investigated the genes expressed in atopic skin using single-molecule direct RNA sequencing of samples from children with atopic eczema.

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Paediatric case with atopic eczema Clinical photographs courtesy of the University of Dundee Computing and Media Services, Ninewells Hospital and Medical School; reproduced with patient/parental consent.


Dr Albena Dinkova-Kostova PhD Senior Lecturer | a.dinkovakostova@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

The role of Nrf2 in skin carcinogenesis: mechanisms of cytoprotection Non-melanoma skin cancers are the most common human malignancies, with more than two million new cases diagnosed globally each year. In addition, cutaneous squamous cell carcinomas are among the most highly mutated human cancers, carrying 1 mutation per ~30,000 bp of coding sequence. Solar ultraviolet (UV) radiation, is the main factor in the etiology of cutaneous squamous cell carcinomas, both in the general and the immunosuppressed populations, causing direct DNA damage, generation of reactive oxygen species, inflammation, and immunosuppression. Most of these damaging processes are counteracted by the endogenous cytoprotective mechanisms that are regulated by the cap’n’collar (CNC) basic-region leucine zipper (bZIP) transcription factor NF-E2 p45related factor 2 (Nrf2, also called Nfe2l2). Nrf2 orchestrates an inducible transcriptional programme comprising nearly 500 genes encoding cytoprotective proteins, which allow adaptation and survival under conditions of electrophilic, oxidative, and inflammatory stress. Experimental evidence has demonstrated that the absence of Nrf2 increases the sensitivity to numerous carcinogens and accelerates disease progression. Conversely, Nrf2 activation by pharmacological agents protects against cancer in various animal models, including a high-risk mouse model of UV radiation-induced cutaneous squamous cell carcinoma. Paradoxically however, Nrf2 is frequently activated in established human tumours and contributes to resistance to chemotherapy and radiation therapy. Our work aims to understand the role of Nrf2 in UV radiation-induced skin carcinogenesis. We use genetic mouse models in which Nrf2 is either disrupted or constitutively activated, as

well as pharmacological Nrf2 activators with a range of potencies to investigate how Nrf2 activity impacts on the development of cutaneous squamous cell carcinoma after chronic exposures to low doses of solar-simulated UV radiation, which are relevant to human occupational or recreational exposures to sunlight. In collaboration with our clinical colleagues in the Photobiology Unit of the Ninewells Hospital and Medical School at the University of Dundee, we are also testing the potential protective properties of pharmacological Nrf2 activators against skin erythema development caused by solar-simulated UV radiation in human subjects.

Model of Nrf2 regulation. (a) Under basal conditions, dimeric Keap1 (blue) binds through its Kelch domains and continuously targets transcription factor Nrf2 (purple) for ubiquitination via association with Cullin 3 (Cul3)-based E3 ubiquitin ligase (orange). (b) Inducers (exemplified by sulforaphane) bind and chemically modify reactive cysteine residues of Keap1, leading to loss of its ability to target Nrf2 for ubiquitination. Consequently, the binding sites of Keap1 are saturated with unmodified Nrf2, and the newly synthesized Nrf2 accumulates, heterodimerizes with a small Maf transcription factor (green), and the complex drives transcription of cytoprotective genes.

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Dr Robyn Hickerson PhD Marie Curie Research Fellow | r.p.hickerson@dundee.ac.uk

Clinical Physics

Biology

Chemistry

Bioinformatics

Identification of molecular transporters that facilitate siRNA delivery

One specific goal of my research is to address these obstacles by identifying and developing novel siRNA modifications that enhance cellular uptake and thus improve in vivo siRNA delivery. In order to achieve this we are developing a simple, yet efficient, method to conjugate a library of small molecules to siRNAs. The proposed methodology takes advantage of standard thiol and amine chemistries allowing any primary amine-containing molecule to be covalently attached to the 3’-end of the passenger strand of a siRNA in a one-step reaction. This novel conjugation system will be used to screen chemical libraries to identify modifications that improve siRNA delivery. Development of molecular transporters to deliver siRNA more efficiently has high practical significance in the development of nucleic acid therapeutics for the treatment of genetic disorders.

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In addition to nucleic acid-based therapeutics, I am also interested in developing small molecules to treat genetic skin disease. The ability to locally deliver small molecule gene modulators would be a boon to patients suffering from a number of genetic skin disorders including eczema, epidermolysis bullosa simplex (EBS), epidermolytic hyperkeratosis (EHK), pachyonychia congenita (PC), Hailey-Hailey, Darier’s disease, and loricrin keratoderma (LK) to name a few. In collaboration with the Drug Discovery Unit within CLS, we have developed a 96-well format primary keratinocyte screen against the clinical compound set (~1,500 compounds) evaluating gene modulation by quantitative RT-PCR. An advantage of this type of screen is the ability to evaluate multiple targets from a single screen. Preliminary data suggests a number of “hits” that will be further evaluated for efficacy and safety before moving into the clinic.

Delta Ct (target gene Ct - reference gene Ct)

The discovery of RNA interference (RNAi) is a major breakthrough in biology and has the potential to make an enormous impact on the treatment of many diseases. Due to its accessibility, skin is an attractive target for siRNA therapeutics. However, there are barriers to overcome to enable successful delivery of siRNA to skin: (i) penetration through the stratum corneum barrier; and (ii) inefficient uptake of functional siRNA by the different classes of cells e.g. keratinocytes, fibroblasts, etc. Previous programmes of work have examined ways to overcome these problems, including different methodologies to facilitate stratum corneum penetration (e.g. intradermal injection, electroporation and microneedle application) and modification of siRNAs with varying transporters (e.g cholesterol, proteins and small molecules).

Toxic compounds 12-fold up 14-fold up 23-fold up Raw Ct (reference gene)

Filaggrin upregulation in quantitative PCR based drug screen


Dr Deena Leslie Pedrioli PhD Independent Investigator | d.lesliepedrioli@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Targeted therapies for keratinizing skin disorders The epidermis is the largest, most complex epithelial tissue in the human body and its mechanical integrity is vital in protecting the human body from harm. My research focuses on a large group of highly debilitating, difficultto-manage hereditary skin conditions called keratinizing skin disorders. Most keratinizing disorders arise from single nucleotide mutations in one of several critically important structural molecules of the skin, which leads to the development of weakened, thick skin - epidermal fragility and hyperkeratosis. Collectively these diseases affect ~1 in 2000 people, but because they are individually rare very little progress has been made towards developing effective treatments. We have begun to tackle the major challenge of developing effective, long-term treatments for these conditions. The biggest obstacle in developing treatments for these disorders lies in selectively repairing or silencing the causative mutation. Harnessing the therapeutic potential of the RNA interference (RNAi) pathway, we have identified several siRNAs that specifically target keratinizing skin disorder mutations. We now aim to develop a patient-friendly way to delivery siRNA into the skin. This is done using unique in vivo reporter and keratinizing skin disorder models that allow us to monitor siRNA delivery into the skin in real-time and determine whether this delivery alleviates human diseaselike symptoms. These studies will provide the preclinical evidence required to progress siRNA therapeutics into clinical trials and, ultimately, patient prescribed treatments.

Because each condition is individually rare it is important to explore the possibility of developing one common treatment for all keratinizing skin disorders. New medicines aimed at a common disease feature, like hyperkeratosis, may allow treatment of these disorders regardless of the genetic abnormality. In line with this, another goal of my research programme is to identify the molecules and biological pathways that cause a single gene mutation to develop into the phenotypic end product of weak, thickened, blistering, painful skin. We are using innovative, state-of-the-art molecular profiling techniques to assemble an in-depth inventory of the individual cellular components that are present in normal but not diseased states, and vice versa. These studies aim to identify the molecular mechanisms that trigger hyperkeratosis and shed light on the cellular processes that regulate hyperkeratosis. The hope is that these findings will seed the development of a generic treatment for most, if not all, keratinizing skin disorders.

Keratinizing Skin Disorders. (a) Clinical features of the keratinizing disorder Epidermolytic Palmoplantar Keratoderma (EPPK). (b-c) Histological comparison of normal (b) vs. disease (c) model skin shows EPPK-like footpad localised hyperkeratosis and acanthosis.

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Prof Charlotte Proby

MBBS, FRCP

Professor of Dermatology | c.proby@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Combating the skin cancer epidemic; understanding the molecular mechanisms driving SCC Within NHS Tayside, and generally in the UK, there has been a 4-fold increase in skin cancers in the past 20 years and my work is directed towards our fight against this skin cancer epidemic. My research is focused especially on the molecular mechanisms driving squamous cell carcinoma (SCC) and the identification of biomarkers and therapeutic targets. Genetic studies confirm the massive burden of UV-induced mutations in cutaneous SCC and challenge us to identify the few genetic ‘drivers’ (such as Notch & p53 mutations) from multiple genetic ‘passengers’. We aim to identify, validate and trial targeted therapies in in vitro and in vivo models of skin cancer prior to translation into human trial.

Microinvasive SCC Cross section of an early SCC showing a small area of ‘invasion’ where the cancer is invading down into the underlying dermis.

Drugs or diseases that compromise the immune system lead to a hugely increased skin cancer risk, presumably allowing the many skin cells already harboring tumour initiating mutations to escape immune surveillance and progress to invasive cancers.

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Together with my colleague, Professor Catherine Harwood (Queen Mary’s University of London), I have defined the risks and developed comprehensive prevention strategies in high-risk patients such as the immunosuppressed organ transplant recipients (OTR) who show a 150fold increased risk for SCC skin cancers. These prevention strategies include topical approaches to treat the ‘field’ of sun-damaged skin aimed at reversing much of the pre-cancerous abnormality. We are undertaking clinical trials of new agents funded by the pharmaceutical industry and head to head comparisons of established topical agents funded nationally by the NIHR Research for Patient Benefit (SPOT study). A new clinical trial (the first in the UK) for use of MEK inhibitor in NRAS-mutation positive melanoma (NEMO trial) commenced in Dundee in December 2013 (PI Dr Tim Crook, Medical Oncologist). In collaboration with Professor Irene Leigh and Dr Andrew South, my group are working with Dundee University’s Drug Discovery Unit, as part of DGEM’s cutaneous drug discovery portfolio, to examine drug sensitivities in 20 primary SCC cancer cell lines aiming to identify suitable candidates to take forward for topical delivery to pre-cancerous skin. This approach takes advantage of an asset unique to Dundee, our large panel of patient-derived skin SCC cell lines, which are also the basis of our in vitro organotypic (skin equivalent) cultures and in vivo mouse xenograft and xenotransplantation skin cancer models.


Dr Aileen Sandilands PhD Research Fellow | a.sandilands@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Using mouse models to investigate skin barrier function and relationship to atopic disease The outermost layer of the skin, the stratum corneum, is a highly effective physical barrier that prevents excessive loss of water from the body as well as preventing the percutaneous entry of pathogens and allergens. Disruption of the stratum corneum, through the loss of structural proteins such as profilaggrin, renders the individual highly susceptible to a variety of atopic diseases such as atopic eczema, asthma, allergic rhinitis (“hayfever�) and food allergy. I am primarily interested in using mouse models of skin disease to identify key proteins and interactions that are important for maintaining skin barrier function. For example, the filaggrindeficient mouse mutant flaky tail has been shown to be a valuable animal model for gaining mechanistic insights into the pathogenesis of human atopic disease. Recently, we identified the gene defect underlying another mouse mutant, matted, which develops spontaneous skin inflammation. Using positional mapping on chromosome 3, coupled with transcriptome analysis, we identified a nonsense mutation in a previously uncharacterized gene, Tmem79, in this mutant strain. Tmem79 encodes a multi-pass transmembrane protein which may play a role in lamellar granule secretion during the process of cornification. During the course of this work we also identified a missense mutation in the human TMEM79 gene that is significantly associated with atopic eczema – highlighting the usefulness of mouse models of skin dysfunction for identifying hitherto unknown genes that are associated with skin disease in humans.

It is hoped that continuing the characterization of these mutant mice and the generation of other mouse models will yield further valuable insights into the importance of skin barrier proteins in human atopic disease.

Immunofluorescence image of a section of human skin (scalp) stained with antibodies to transmembrane protein 79 (green). Nuclei are counterstained in blue. The red arrow indicates staining of the keratinocyte plasma membrane.

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Dr Mark Saville PhD Lecturer | m.k.saville@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Understanding the role of the ubiquitin system in skin cancer and developing therapeutic strategies Ubiquitination can act as a signal which targets proteins for degradation by the proteasome and can also control protein activity and localisation. Through these mechanisms ubiquitination regulates many fundamental processes relevant to cancer and its therapy including: signalling pathways, cell cycle progression, apoptosis and DNA-repair. My group is exploring the potential of components of the ubiquitin system as targets for the treatment of cancer and characterising the roles of these proteins. We have a major focus on improving the treatment of cutaneous squamous cell carcinoma (cSCC). The transcription factor p53 plays a key role in protecting against tumour development. p53 and its repressors Mdm2 and MdmX are ubiquitinated and are degraded by the proteasome. We are studying how the ubiquitin system regulates the balance between p53, Mdm2 and MdmX. p53 mutation can result in loss of its normal tumour protective activity and in some circumstances can result in tumour-promoting gain of function. This gain of function activity is dependent on elevated mutant p53 protein expression. Mutation and accumulation of p53 is an early step in cSCC development. We are investigating the role of mutant p53 in cSCC progression. To identify therapeutically tractable ways to reduce mutant p53 protein expression we are looking at the mechanism(s) of mutant p53 stabilisation in cSCC.

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The proteasome plays a major role in ubiquitindependent protein degradation. The 26S proteasome is made up of two multisubunit complexes: the 19S regulatory particle and the 20S core. The proteasome inhibitor bortezomib has made a great impact on the treatment of some cancers. We are looking at the potential of bortezomib and other inhibitors of the proteolytic activity of the 20S core for cSCC therapy. We are also interested in alternative mechanisms of targeting the proteasome and proteasomal degradation. This includes targeting activities of the 19S regulatory particle and the recruitment of ubiquitinated proteins to the proteasome. There are several families of proteins in the ubiquitin-proteasome system. Ubiquitin conjugation to target proteins requires the sequential action of a ubiquitin activating enzyme (E1), a ubiquitin conjugating enzyme (E2) and a ubiquitin ligase (E3). Ubiquitin-protein linkages can be cleaved by the action of deubiquitinating enzymes (DUBs). Ubiquitin receptors are involved in mediating the outcome of ubiquitination. We are carrying out siRNA screens to identify additional components of the ubiquitin system which could be targeted for cSCC therapy.


Dr Frances Smith PhD Senior Research Fellow | f.j.d.smith@dundee.ac.uk

Clinical Physics

Biology

Chemistry

Bioinformatics

Molecular basis of human keratinizing disorders Keratinizing disorders are a large group of inherited skin conditions that cause blistering and/or hyperkeratosis (overgrowth/thickened skin) and/or ichthyosis (scaling/flaking) of the epidermis. Some of these conditions predominantly affect the palm and sole skin leading to thickened, painful calluses (keratoderma), particularly on the pressure points on the feet. Others are more widespread with for example, ichthyosis affecting the whole body. My research interest is in the discovery of genes underlying these rare genetic skin disorders and the function of these genes within the skin. I have been involved in identifying the causative genes for a number of these disorders including the first keratin disorder, epidermolysis bullosa simplex (EBS), other keratin disorders including pachyonychia congenita (PC), EBS with muscular dystrophy due to mutations in the plectin gene, the discovery of the first filaggrin mutations in ichthyosis vulgaris and atopic dermatitis and more recently in identification a new gene causing a painful and debilitating skin disorder, punctate palmoplantar keratoderma (also known as punctate PPK or PPKP1). With the recent development of new technology we are now using a whole exome sequencing (WES) approach to identify new genes as the underlying cause of cases with no known candidate genes. When we identify new genes our approach is that assays are developed so we can offer genetic testing of these disorders through our NHS Tayside partner laboratory at nearby Ninewells Hospital and Medical School, Dundee. We currently provide genetic testing for the keratin disorder pachyonychia congenita (PC), for all individuals registered with the Pachyonychia Congenita Project (www.pachyonychia.org).

We also offer genetic testing of other rare keratoderma disorders not currently provided by NHS Molecular Genetics Lab, Ninewells. For the patients, our aim is to develop diagnostic tests for these disorders with the ultimate goal of developing new therapies to treat and improve the quality of life for these individuals. I am involved in various projects to develop therapeutics such as siRNA for pachyonychia congenita and other keratin disorders. A

B

Immunofluorescence images stained using antibodies to keratin (green) and nuclei (blue) showing (a) a dense meshwork of keratin intermediate filament bundles in normal cells (b) abnormal “clumping� of the keratin filaments due to the presence of a mutation in the K14 keratin gene.

The genetic testing provided for patients diagnosed with Pachyonychia Congenita (PC) has been an enormous help, both to individual patients and to the effort to learn more about PC in order to develop effective treatments. Through the work of the Dundee laboratory, nearly 100 mutations have been identified in genes causing PC. This has led to important new classifications and as a result better differential diagnosis guidelines have been drafted. Dr Mary Schwartz, Director of PC Project

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Dr Andrew Woodland PhD ITP Portfolio manager, DDU | a.woodland@dundee.ac.uk

Clinical Physics

Chemistry

Biology

Bioinformatics

Development of new medicines for first world disease As Portfolio Manager for the Innovative Targets Portfolio (ITP) within the Drug Discovery Unit (DDU), I lead the first world drug discovery projects within the DDU, de-risking novel targets and developing promising new therapies. Within the first two years of this initiative we partnered two projects with GSK under the DPAc initiative. Now 4 years old, the ITP supports a core team capable of progressing nine new projects each year, through hit identification and hit to lead development. The DDU’s model is that we wish to compliment biopharma strengths by de-risking and partially validating higher risk, un-validated drug targets which seek to address an unmet medical need. As part of our first world drug discovery efforts we have established a cutaneous drug discovery group (CDDG) which is embedded within both the DDU and DGEM. The CDDG been established to address the unmet medical need for dermatology translational research. This unique initiative builds on our expertise in cutaneous drug design, anti-drug design, understanding of exposurepharmacodynamic relationships, and topical formulations. The cutaneous drug discovery portfolio (CDDP) progresses early stage projects bridging the translation gap and generating high quality data packages suitable for partnerships with pharmaceutical development organisations.

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A broad range of technologies are actively under development by the CDDG. We are currently working with Prof Irwin McLean and Dr Robyn Hickerson on the development of a high throughput qPCR assay capable of accurately assessing the effects of drug like compounds on gene regulation in disease relevant cell lines. We believe this new approach will have broad applicability for rare genetic disorders. In more common diseases we are exploring new strategies to treat the underlying causes of diseases such as eczema and psoriasis. In eczema we are seeking to enhance the levels of the important skin barrier protein filaggrin. For psoriasis, we are exploring a target which will have pleotropic effects across the range of cell types involved in this disease. We believe such approaches will deliver enhanced efficacy over available treatments. In addition to our established projects we are constantly developing new opportunities and are interested in forming drug discovery collaborations with leading researchers throughout the UK and internationally.


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Genomic Sequencing Unit (GSU)

m.febrer@dundee.ac.uk http://gsu.lifesci.dundee.ac.uk +44 (0)1382 381 048

About

Equipment

To fulfill the research requirements of DGEM for genodermatology research and diagnosis, nextgeneration sequencing (NGS) and bioinformatics infrastructure has been implemented within the Centre. The NGS facility, known as the Genomic Sequencing Unit (GSU), is run as a core facility thereby also supporting world-class research across the University. A dedicated Facility Manager (Dr Melanie Febrer) and support team provide access to well maintained equipment and develop and implement the latest techniques for the benefit of its users. GSU services are also available to individuals outside of Dundee University and are offered on a competitive basis. At present, GSU provides minimal bioinformatics analysis to individuals outside of DGEM, and users are therefore requested to source their own bioinformatics support. However the facility, in collaboration with Prof Barton’s group, is in the process of developing a Galaxy-based system to deliver data and make certain analyses (e.g. RNA-seq) possible for all users.

The facility is equipped with Illumina’s next generation sequencing machines, the HiSeq 2000 and MiSeq. High throughput capabilities are achieved using the Beckman Beckman FXp Dual arm system robotics system. Other equipment used to operate this facility includes the Covaris M220 (DNA shearing), Agilent Tape Station (quality control) and Life Technologies Qubit (Fluorometric Quantitation).

Applications A wide range and ever-increasing number of different type of applications are available through GSU. Some examples are given below: • De novo/ re sequencing • RNA-seq (either random primed or directional) • Whole exome sequencing • Targeted sequencing • ChIP Seq • 16S amplicon sequencing or Metagenomics GSU welcomes collaborative projects outside of routine experiments on an ad hoc basis.

Prior to the opening of the GSU facility we were sending material across the world for processing at centres in Bejing, Bangalore and Edinburgh. Since the facility has opened in Dundee we have benefited from faster processing of our samples and the availability of local expertise aids the design of our experiments. Professor Tom Owen-Hughes, Wellcome Trust Senior Research Fellow, College of Life Sciences, Dundee

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Training The Centre for Dermatology and Genetic Medicine is dedicated to training the next generation of researchers. We have unique resources for teaching due to our multifaceted approach to translational dermatology (monogenic and complex trait genetics, clinical network, cell and animal model development, drug discovery, formulation and biomedical physics). DGEM therefore provides a vibrant environment for postgraduate teaching and currently has 7 PhD students supported by an active postgraduate society and a structured mentoring/monitoring process. GSDRC To further enhance international cross-disciplinary training and collaboration, DGEM has joined the Global Skin Disease Research Consortium (GSDRC) with 5 major dermatology research groups in the USA, Germany, Japan and Singapore. The long-term goals of this established collaborative agreement are to: • Provide joint opportunities •F  oster the creation of novel approaches and insights into cutaneous biology, disease pathogenesis and clinical dermatology by coordinating multidisciplinary research aimed at finding innovative translational approaches to improve the care of patients with skin disease • Train the scientific leaders of tomorrow Accomplishment of these aims will be supported by activities such as: symposiums and annual video conferencing; summer schools for PhD students and postdocs; exchange of students and postdocs to learn new techniques and approaches; exchange of faculty; and establishment of a web-based inventory to promote exchange of mouse models, other animal models, reagents and protocols.

Irwin McLean, Irene Leigh Dundee Angela Christiano, David Bickers New York

Carien Nieseen, Thomas Krieg Cologne

Masayuki Amagai Tokyo Dennis Roop, David Norris Denver

The Global Skin Disease Research Consortium 30

Birgit Lane Singapore


I chose DGEM for my postgraduate studies because of the translational research opportunities made possible by the clinical and academic expertise. The availability and interaction between the two have fostered a supportive environment that has been invaluable to my development as a scientist and has motivated me to pursue medical studies with the aspiration of becoming a clinical academic. Dun Jack Fu, Wellcome Trust 4-year PhD student

Dermatogenetics and BADGEM DGEM is also uniquely placed to provide training opportunities to the wider clinical and academic communities in the UK through its strong links to Dermatogenetics (sub-group of the umbrella organisation BSGM) and via BADGEM (the British Association of Dermatologists Dermatology & Genetic Medicine), a newly formed UK-wide clinical network dedicated to rare genetic diseases of the skin.

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The Schwartz Lecture Mary Schwartz, long-term collaborator of researchers in Dundee, and founder of the international charity Pachyonychia Congenita (PC) Project, received an Honorary Doctor in Laws by the University of Dundee in the summer of 2013. Since she founded PC project in 2003/4, Mary Schwartz has been a major source of inspiration and a driving force behind research into pachyonychia congenita. She has identified and brought together thousands of PC patients from all corners of the globe, and has funded grants in academic labs, including Dundee, as well as spinning out the company Transderm Inc, all in the search for a treatment for PC and related skin conditions. Much of the funding has come from Mary’s own pocket and that of the Schwartz family themselves. PC Project is simply one of the cornerstones of translational research in dermatology and represents a new paradigm for how a patient-led initiative can shape and accelerate the course of therapeutic research. In recognition of her enormous contribution to the skin disease community, the University of Dundee awarded Mary with an honorary Doctorate, which was long-overdue and much deserved. To continue to honour and recognise Mary’s outstanding contributions in this area, the Schwartz Lecture has been initiated. This is an annual event where DGEM get to invite the top internationally thought-leaders in the field of skin biology to Dundee to hear the latest research from their lab. Mary kindly provided some seed funding for this lecture series and the Heads of the Colleges of Life Sciences and of Medicine, Dentistry and Nursing will make this a permanent fixture and will continue to fund this lecture series in perpetuity. Schwartz Lecture Series 2013 (Inaugural Lecture) - Professor Pierre Coulombe John Hopkins Bloomberg School of Public Health, Baltimore. “Keratin intermediate filaments: A unique perspective on epithelial biology in health and disease”. Pierre is a leading epithelial cell biologist who has very successfully used biochemistry, human genetics, transgenics and structural biology methodologies to define the structural and importantly, non-structural, physiological functions of intermediate filaments. 2014 – Professor Eli Sprecher The Tel Aviv Sourasky Medical Center, Israel. Eli has carried pioneering work on deciphering the molecular basis of numerous genodermatoses, delineated the genetic basis of acquired diseases such as pemphigus and is translating genetic findings into innovative treatment strategies for inflammatory diseases.

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Patient Support Activities and Outreach Investigators within DGEM are actively involved in patient support activities and fund-raising efforts. We currently run two patient support meetings per year for the painful incurable skin blistering disorders pachyonchia congenita (PC) and the closely related condition epidermolysis bullosa simplex (EBS). These meetings provide a unique opportunity to bring together patients with expert dermatologists and scientists. The Centre also established, and currently administers, the charity PC Project UK (http://pcprojectuk.org.uk/). Funds raised for PC Project UK are used to develop and deliver treatments for PC and EBS, as well as to financially support the patient meetings. DGEM also has strong links with other the patient associations such as DEBRA, Eczema Outreach Scotland and the Ichthyosis Support Group. As well as patient focused activities, DGEM participates in a wide variety of other events such as the annual College of Life Sciences Open Doors Day, school outreach programmes and CafĂŠ Science informing the wider public and schools about our research aims and discoveries in plain language and imaginative formats. We continually monitor and evaluate the impact of our outreach events to ensure that our activities are as successful as possible.

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Financial Support The Centre for Dermatology and Genetic Medicine is in receipt of a strategic award from the Wellcome Trust (2012-2017) which helps to support the activities of DGEM. This is augmented by joint funding and administrative support from CLS, CMDN and the University of Dundee, who provide two secretaries, laboratory manager support and a centre manager to help run DGEM effectively. A large proportion of research carried out in DGEM is funded by competitive fellowships, programme and project grants awarded to the individual group leaders by national and international funding agencies. We are enormously grateful to the charities, funding agencies and companies that have funded our work past and present. We are also especially grateful to the many individuals, patients and families, either directly or indirectly affected by the genetic disorders we work on, who have made donations towards our research efforts.

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Contact The main campus of the University of Dundee is located centrally in the City, close to both the train station with regular direct services to Newcastle, York and London, and many other major cities, and also Dundee Airport which is served by direct flights to London Stansted. Edinburgh airport, about an hours drive away, provides access to many international destinations. Ninewells Hospital, situated on the western outskirts of Dundee, is only a short drive from the main University Campus. Postal address Centre for Dermatology and Genetic Medicine Colleges of Life Sciences and Medicine, Dentistry & Nursing University of Dundee Dow Street, Dundee DD1 5EH Find out more http://dgem.lifesci.dundee.ac.uk +44 (0) 1382 381 046 DGEM@dundee.ac.uk @DGEM_Dundee

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Centre for Dermatology and Genetic Medicine Colleges of Life Sciences and Medicine, Dentistry & Nursing University of Dundee Dow Street, Dundee DD1 5EH +44 (0)1382 381 046 dgem@dundee.ac.uk

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University of Dundee Nethergate, Dundee DD1 4HN +44 (0)1382 383 000 university@dundee.ac.uk June 2014 Cover image: Protein expression in skin showing filaggrin (green) and K5 (red) - Image supplied by Declan Lunny

DGEM Brochure June 2014  

Read about current research activities in the Centre for Dermatology and Genetic Medicine at the University of Dundee (http://dgem.lifesci.d...

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