Issue 36 by The Meducator

DECEMBER 2019 | ISSUE 36

BIOTECH BLUEPRINT 3D P R I N T I N G I N S U RG E RY A N D M E D I C I N E

VIEWPOINTS D O E S T H E C M PA S E RV E T H E I N T E R E S T S O F PAT I E N T S?

INTERVIEW SPOTLIGHT I N T E RV I E W W I T H D R. G O R D O N G U YAT T, F O U N D E R O F E V I D E N C E-B A S E D M E D I C I N E

WWW.MEDUCATOR.ORG

table of CONTENTS DECEMBER 2019

ISSUE 36

01 INTRODUCTION 02 MEDPULSE 04 MEDBULLETIN 06 PATHOPROFILE 08 BIOTECH BLUEPRINT

table of contents

10 NEUROABSTRACTS 12 VIEWPOINTS CRITICAL REVIEW 14 Tick-borne diseases: From well-known Lyme disease to deadly Powassan encephalitis 18 A critical review of pyruvate kinase deficiency RESEARCH INSIGHT 22 Developing a qualitative chemotaxis-sensitive assay for bacteriophage-host interactions on E. coli BW2511

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26 INTERVIEW SPOTLIGHT 29 CONTRIBUTORS TOC ARTIST MANREET DHALIWAL COVER ARTIST PERI REN

dear reader,

Our mandate of facilitating critical discourse within the health sciences remains front and centre in Issue 36, which showcases an exciting array of student exploration, debate, and inquiry. Kashyap Patel provides a critical overview of mitapivat, a novel therapy for pyruvate kinase deficiency, calling for higher quality evidence on the safety and efficacy of mitapivat. Dr. Gordon Guyatt, a Professor in the Department of Health Research Methods, Evidence, and Impact at McMaster University, recounts his role in establishing the current paradigm of evidence-based medicine and discusses its future. Rohan Aananth, Milena Cioana, and Nuri Song deliberate whether medical malpractice insurance enhances quality of care, providing viewpoints on both benefits and harms to patients. Through comparison of the most common and most lethal forms of tick-borne diseases, Jerry Chang identifies primary prevention as the most effective measure against the spread of disease. Finally, Aaron Wen discusses how the optimization of a chemotaxis-detecting assay may help to elucidate the impact of bacteriophage vicinity on bacterial movement. In line with this issue’s focus on the intersection of technology and health sciences, we are excited to introduce an all-new article, Biotech Blueprint. Biotech Blueprint shines a spotlight on a recent biotechnological advancement that has the potential to reshape the health sciences landscape. Adrian Wong and Albert Stancescu provide a brief history of 3D printing before delving into its application to human surgery. To parallel the increasing clinical accessibility of 3D printing, Adrian and Albert suggest that regulations and protocols be established for related technologies.

Sincerely,

JIM XIE

| DECEMBER 2019

EDWARD CUI

Bachelor of Health Sciences (Honours) Class of 2021

M E D U C AT O R

We are incredibly grateful for the opportunity to work alongside such talented and passionate individuals for the 36th publication cycle. The dedication and resilience of our staff, authors, and post-graduate partners are integral to the continued excellence of this journal. In particular, we would like to thank our Managing Editors, Kevin and Saba; Creative Directors, Suffia and Karishma; Video Manager, Adrian; MeduCollab Directors, Daniel and Annisa; and MeduPromo Director, Zahra, for their leadership and initiative. Finally, we would also like to express our gratitude to you, our reader, for your continued support of our publication. We hope something in this issue piques your interest, sparks a question, and inspires innovation of your own.

introduction

INTRODUCTION ISSUE 36

Welcome to Issue 36 of The Meducator! The vast, interdisciplinary field of health sciences has evolved closely alongside other branches of knowledge such as engineering, and more recently, biotechnology. As displayed in Peri Ren’s cover illustration of an artificially enhanced human in fetal development, we are witnessing the birth of an era where technology is not simply a tool for health care provision, but rather a vital component of our well-being. As precision medicine is enabling increasingly personalized treatment and 3D printing is revolutionizing surgical possibilities, how can we leverage their potential while taking ethical considerations into account? How can healthcare professionals adopt new innovations to improve quality of care and prevent harm to patients? How can society at large employ technology to elevate the health and well-being of all citizens? As you explore this issue, we invite you to grapple with similar questions and contemplate technology’s growing implications for the health sciences.

Bachelor of Health Sciences (Honours) Class of 2020

MEDPULSE Taking the Pulse of the World

AUTHORS: Meera Chopra & James Yu ARTIST: Kien Nagales

Ovarian Cancer Drug Approved in UK October 2019 – United Kingdom

Ovarian cancer can be incredibly difﬁcult to treat due to a lack of outward symptoms. As a result, the disease is estimated to be the 6th most common form of cancer in the United Kingdom. A pharmaceutical product called Rubraca can slow down the progression of the disease by killing cancer cells. This medication works by inhibiting poly ADP-ribose polymerase (PARP), a protein that helps repair damaged DNA. Cancer cells are unable to ﬁx their damaged DNA, rendering them incapable of replicating. Rubraca was recently approved for prescription in the UK and is thought to be able to improve the quality of life of patients with ovarian cancer.

Teen Vaping Doubles in the United States September 2019 – United States of America

Since 2017, the rate of e-cigarette use in teens has doubled in the US. In 2019, 25% of twelfth graders, 20% of tenth graders, and 9% of eighth graders reported having vaped nicotine in the past month. This increase is attributed to the variety of appealing ﬂavours and perception of safety of vaping. The ﬁrst death from e-cigarettes occurred in September 2019 and, since then, over 1479 cases with 33 deaths have been reported. The injuries are typically chemically induced and have been largely attributed to illicit vaping liquids and vaping products containing THC. While the majority of cases have been in the US, an increasing number of cases have been reported in Canada.

Dengue Fever Stings in Honduras July 2019 – Honduras

Dengue is a viral infection that causes causes high fever and joint pain. It can develop into a potentially lethal complication called severe dengue. Over 40,000 cases and 135 deaths have been reported in Honduras in 2019, marking the worst outbreak of the virus in over 50 years. In comparison, only 8,000 cases were reported in 2018. The increase has led Honduras’s government to declare a national emergency and fumigate the breeding grounds of yellow fever mosquitoes which spread the disease. Experts speculate that climate change and Honduras’s three-month rainy season may have contributed to the epidemic.

Madariaga Virus Discovered in Argentina September 2019 – Argentina

Madariaga virus is a mosquito-borne disease originating in Panama. The virus has slowly spread across Central and South America since 2010. Little is known about the symptoms and fatality of this disease, as less than a dozen cases have been documented. However, it is thought to be similar to equine disease. The Madariaga virus has recently been discovered in mosquitoes in Argentina. Scientists working with the Institute of Virology with National University of Córdoba are currently looking at how this virus is spreading across South American countries and how future outbreaks can be prevented.

Polypills Prevent Heart Disease in Iran August 2019 – Iran

A polypill is a combination of multiple medications in a single pill. The PolyIran cluster-randomized trial enrolled 6,838 people in the Golestan province of Iran. Half of the patients received a polypill containing aspirin, a cholesterol-lowering statin, and two blood pressure drugs. Over 80% of patients adhered to the treatment and major cardiovascular events were cut by 34%. Among patients that had the highest adherence, events decreased by 57%. More than 18 million people die every year from cardiovascular disease and 80% of them are in low- and middle-income countries. The success of this trial may help open the door for the widespread use of polypills which promise healthier lives for millions of people.

Air Pollution Results in Missed Miscarriages October 2019 – China

Researchers in China have found that increased levels of pollution lead to higher rates of “missed” miscarriages in pregnant women. Missed miscarriages occur when a fetus dies during early pregnancy without any noticeable symptoms. A study from the Chinese Academy of Sciences found that increased levels of certain airborne particles, including carbon monoxide and sulfur dioxide, can exponentially increase rates of missed miscarriages. Previously, researchers understood that dangerously high levels of pollution can lead to pregnancy complications but this new development reveals another consequence of dangerously high levels of air pollution.

Australia Experiences Early and Severe Flu Season July 2019 – Australia

Australia suffered an early and severe flu season this year, caused by the particularly virulent influenza strain, H3N2. The strain, which causes more significant illnesses in elderly individuals, is also less affected by the flu vaccine. This year’s season, beginning in April and ending in October, led to more than 272,000 confirmed cases and 662 deaths. The flu was the worst since 2001 when the Australian government first began releasing public data. Experts warn that the flu season in Australia is a moderate predictor of what the flu will be like in Canada and the US.

Measles Outbreak in the Democratic Republic of Congo October 2019 – Democratic Republic of Congo

Over 200,000 people in the Democratic Republic of Congo have been diagnosed with measles, 4,000 of which have died. According to UNICEF, only 57% of the Congolese population has been vaccinated against the measles. This low vaccination rate, along with unstable health infrastructure and economic uncertainty, contributes to the rapid spread of the virus. In order to combat the spread of measles, the government of Democratic Republic of Congo is collaborating with World Health Organization to vaccinate 825,000 children in the region. Citations can be found on meducator.org

MEDBULLETIN ANTIBIOTICS

GLIOBLASTOMA

SHADI SADEGHIAN Recent in vivo experiments have demonstrated that, upon treatment with oral antibiotics, mice become increasingly susceptible to West Nile (WNV), Dengue (DNV), and Zika (ZIKV) viral infections.1 Using ampicillin for as little as three days yielded exaggerated WNV symptoms.2 Specifically, the experimental data demonstrated that, after antibiotic exposure, there was a decrease in the number of murine immune cells in gut-distal lymphoid tissue. Specifically, there were fewer CD8+ T-cells relative to antibiotic-free controls.2 In previous experiments, the transfer of CD8+ T-cells has been said to protect alymphoid mice against a lethal WNV injection.3 To test whether unprimed CD8+ T-cells can provide protection against lethal WNV, splenic CD8+ T-cells were transferred into mice, along with a lethal dose of WNV.3 While all control recipients died, 75% of those who received a transfer of CD8+ T-cells survived.3 This survival rate suggests that naïve CD8+ T-cells could provide protection against WNV. In addition to this finding, mice were treated with a broad-spectrum antibiotic cocktail for 14 days preand post-inoculation with WNV, ZIKV, and DNV.2 While 80% of the control mice survived at least 21 days post-inoculation, only one experimental mouse survived this time frame.2

SOPHIE ZARB Glioblastoma is an aggressive form of brain cancer that is often fatal within 18 months of initial diagnosis. These malignant, web-like brain tumours develop from astrocytes, a specific type of brain cell. The tumors spread rapidly from the supportive tissue throughout the brain in finger-like projections.1 As there is currently no cure for glioblastoma, treatment is palliative in nature, consisting mainly of chemotherapy, radiation, and/or surgery.1

The authors argued that reducing the robustness of the microbiota is a factor in compromising immunity. This reasoning is consistent with previous studies that have examined this relationship. One experiment demonstrated that commensal bacteria-derived signals stimulate the innate immune system to attain optimal antiviral immunity.4 The specific mechanisms are poorly understood, but it is speculated that commensal bacteria influence the activation threshold of innate antiviral response pathways, such as the interferon signaling pathway.5

These findings suggest that surfen treatment may make it easier to isolate and surgically remove glioblastoma tumours. If surgical intervention is pursued at an earlier stage, the need for other treatment protocols including chemotherapy and radiation may be reduced.2 Researchers are hopeful that the proven safety and beneficial nature of surfen will result in an accelerated path to drug approval, paving the way for the medical community to change the prognosis of this lifethreatening disease.5

M E D U C AT O R

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THE INFLUENCE OF ANTIBIOTICS ON VIRAL IMMUNITY

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1. Williams S. Antibiotics increase mouse susceptibility to Dengue, West Nile, and Zika [Internet]. The Scientist Magazine. 2018 Mar 27. Available from: https://www.the-scientist.com/news-opinion/antibiotics-increase-mouse-susceptibility-to-dengue-west-nile-and-zika-29900 [cited 2019 Sept 8]. 2. Thackray L, Handley S, Gorman M, Poddar S, Bagadia P, Briseño C, et al. Oral antibiotic treatment of mice exacerbates the disease severity of multiple flavivirus infections. Cell Rep. 2018;22(13):3440-53.e6. Available from: doi:10.1016/j.celrep.2018.03.001. 3. Brien J, Uhrlaub J, Nikolich-Žugich J. Protective capacity and epitope specificity of CD8+ T cells responding to lethal West Nile virus infection. Eur J Immunol. 2007;37(7):1855-63. Available from: doi:10.1002/ eji.200737196. 4. Baldridge M, Nice T, McCune B, Yokoyama C, Kambal A, Wheadon M, et al. Commensal microbes and interferon-λ determine persistence of enteric murine norovirus infection. Science. 2014;347(6219):266-9. Available from: doi:10.1126/science.1258025. 5. Abt M, Osborne L, Monticelli L, Doering T, Alenghat T, Sonnenberg G, et al. Commensal bacteria calibrate the activation threshold of innate antiviral immunity. Immunity. 2012;37(1):158-70. Available from: doi:0.1016/j.immuni.2012.04.011.

NEW USE OF EXISTING DRUG COULD TREAT AGGRESSIVE BRAIN TUMOURS

In a recent study conducted at the University of Georgia’s Regenerative Bioscience Center and funded by the National Institutes of Health, researchers found that surfen, a compound molecule used to optimize insulin delivery, can potentially be used as an effective treatment to block tumour growth in the brain.2 Given the positive charge of surfen, researchers wanted to test whether it could be used to block negatively charged sugars in brain tissue that are known to aid in cancer proliferation.2-4 When tested in lab animals, the surfen-treated specimens developed smaller, less branched tumours, suggesting that the molecule hindered the tumour cells’ ability to spread.2

1. 2. 3. 4. 5.

National Centre for Advancing Translational Sciences. Gliobastoma [Internet]. 2015 Jan 19. Available from: https://rarediseases.info.nih.gov/diseases/2491/glioblastoma [cited 2019 Sep 12]. Logun MT, Wynens KE, Simchick G, Zhao W, Mao L, Zhao Q, et al. Surfen-mediated blockade of extratumoral chondroitin sulfate glycosaminoglycans inhibits glioblastoma invasion. FASEB. 2019;33(11):1197392. Available from: doi:10.1096/fj.201802610RR. Murray S, Tulloch A, Criscitelli K, Avena NM. Recent studies of the effects of sugars on brain systems involved in energy balance and reward: Relevance to low calorie sweeteners. Physiol Behav. 2016;164(Pt B):504–8. Available from: doi:10.1016/j.physbeh.2016.04.004. On the Brain. Sugar and the Brain [Internet]. Available from: https://neuro.hms.harvard.edu/harvard-mahoney-neuroscience-institute/brain-newsletter/and-brain-series/sugar-and-brain [cited 2019 Sep 12]. Science Daily. Existing drug could treat aggressive brain cancer: New use of insulin compound could block tumors [Internet]. 2019 Sept 10. Available from: https://www.sciencedaily.com/releases/2019/09/190910154701.htm [cited 2019 Sep 12].

HEART REGENERATION

PARKINSON’S

NEW STUDY BACKS UP INTESTINAL ORIGIN THEORY

MEERA CHOPRA When human heart cells die during a heart attack, a cellular process to replace lost cells is initiated by cytokines. To prevent mitosis, an enzyme called protein tyrosine phosphatase 1B (PTP1B) can halt this signalling pathway and avert the regeneration of cardiac tissue.1 In contrast, when cardiac tissue dies in aquatic animals, an allosteric inhibitor named MSI-1436 hinders PTP1B, leading to cell regeneration.1,2 Furthermore, mice treated with MSI-1436 following a myocardial infarction had 600% more heart tissue regrowth than those without the treatment.2

ADRIAN WONG Parkinson’s disease is a neurological disorder involving various symptoms, including slow movement, tremors of the limbs, stiffness, and deformities in posture.1 The disease currently affects approximately 100,000 Canadians.2 Parkinson’s is characterized by a reduction of dopamine in the striatum of the brain due to a loss of dopamine-releasing neurons. However, the exact cause remains unknown, leading to many theories regarding this matter.3

The regeneration of dead tissue begins once cytokines bind to receptor tyrosine kinase (RTK) on the cell membrane, triggering a signal transduction pathway.2 Generally, this process is terminated in humans by the enzymatic inactivation of RTK by PTP1B.1,2 However, when MSI-1436 binds to the allosteric site of PTP1B, the enzyme is unable to bind to RTK, allowing signal transduction to proceed.1,3 This pathway ends with the activation of a protein called CDK1 cyclin-B that enters the nucleus to stimulate DNA replication. Organelles then duplicate in the G2 phase of the cell cycle and the cell undergoes mitosis and cytokinesis.3 This process is repeated until all required tissue cells are replaced.

1. Jankovic J. Parkinson’s disease: Clinical features and diagnosis. J Neurol Neurosurg Psychiatry. 2008;79(4):368-76. Available from: doi:10.1136/jnnp.2007.131045. 2. Parkinson Canada. Parkinson Canada priorities for the 2018 federal budget. 2017. 5 p. 3. Lotharius J, Brundin P. Pathogenesis of Parkinson’s disease: Dopamine, vesicles and α-synuclein. Nat Rev Neurosci. 2002;3(12):932-42. Available from: doi:10.1038/nrn983. 4. Aarhus University. Parkinson’s disease may originate in the intestines [Internet]. Faculty of Health. 2019 Sept 2. Available from: http://health.au.dk/en/#news-16099 [cited 2019 Sept 14]. 5. Van Den Berge N, Ferreira N, Gram H, Mikkelsen TW, Alstrup AGO, Casadel N, et al. Evidence for bidirectional and trans-synaptic parasympathetic and sympathetic propagation of alpha-synuclein in rats. Neuropathol. 2019;138(4):535–50. Available from: doi:10.1007/s00401-019-02040-w. 6. Siddiqui IJ, Pervaiz N, Abbasi AA. The Parkinson disease gene SNCA: Evolutionary and structural insights with pathological implication. Sci Rep. 2016;6:24475. Available from: doi:10.1038/srep24475. 7. Stefanis L. a-Synuclein in Parkinson’s disease. Cold Spring Harb Perspect Med. 2012;2(2):a009399. Available from: doi:10.1101/cshperspect.a009399. 8. Zadikoff C, Kianirad Y. Managing PD Mid-Stride. Miami: Parkinson’s Foundation; 2018. 44 p.

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1. Smith AM, Maguire-Nguyen KK, Rando TA, Zasloff MA, Strange KB, Yin VP. The protein tyrosine phosphatase 1B inhibitor MSI-1436 stimulates regeneration of heart and multiple other tissues. NPJ Regen Med. 2017;(1):4. Available from: doi:10.1038/s41536-017-0008-1. 2. Strange K, Yin V. A shot at regeneration [Internet]. Scientific American. 2019 Apr. Available from: https://www.scientificamerican.com/magazine/sa/2019/04-01/ [cited 2019 Sept 10]. 3. Daub H, Weiss FU, Wallasch C, Ullrich A. Role of transactivation of the EGF receptor in signalling by G-protein-coupled receptors. Nature. 1996;379(6565):557–60. Available from: doi:10.1038/379557a0. 4. Krishnan N, Tonks NK. Anxious moments for the protein tyrosine phosphatase PTP1B. Trends Neurosci. 2015;38(8):462–5. Available from: doi:10.1016/j.tins.2015.06.006.

This research may have implications for the treatment of Parkinson’s, providing scientific evidence towards an origin for the disease.4 While treatments for Parkinson’s exist, they only replace dopamine in the brain to suppress symptoms and cannot halt the disease.8 A better understanding of the way Parkinson’s arises may allow the development of treatments that can stop it before it causes harm to a patient.4

M E D U C AT O R

Using MSI-1436 is a promising technique for tissue regrowth. This compound has undergone comprehensive toxicity testing and it is believed that injection into organs would not cause significant side effects.2,4 Unlike other regenerative treatments, this technique has a lower risk of electrical impulses from regenerated tissue misaligning with the innate cardiac rhythm.2 Thus, MSI-1436 shows promise for the future of biotechnology and could improve treatment for various heart diseases.

One such theory, proposed in 2003, states that Parkinson’s may originate from the gut and move to the brain.4 A recent study conducted by Van Den Berge et al. presents evidence supporting this theory.5 The researchers engineered transgenic rats that express extra copies of the SNCA gene, the gene most involved in early-onset Parkinson’s.5,6 These rats were used to study the movement of preformed alpha-synuclein fibrils —the intermediate form of which may cause neuronal death by affecting synapses and other targets— to the brain.5,7 After injection of fibrils into the duodenum of the rats, alphasynuclein was found in the major structures of the sympathetic and parasympathetic routes to the brainstem, including the substantia nigra, where the loss of dopamine-releasing neurons occurs.4 These findings suggest that alpha-synuclein spread from the duodenum to the brain.5

medbulletin

REGENERATING TISSUES AFTER A HEART ATTACK

Multiple Myeloma AUTHORS: Michal Moshkovich & Matthew Lynn ARTIST: Wendy Zhang

Pathoprofile doi:10.35493/medu.36.6

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Robiou-Du-Pont S, Cleynen A, Fontan C, Attal M, Munshi N, Corre J, et al. Genomics of multiple myeloma. J Clin Oncol. 2017;35(9):9637. Available from: doi:10.1200/ JCO.2016.70.6705. Hauksdóttir B, Klinke ME, Gunnarsdóttir S, Björnsdóttir K. Patients’ experiences with multiple myeloma: A meta-aggregation of qualitative studies. Oncol Nurs Forum. 2017;44(2):E64-E81. Available from: doi:10.1188/17.ONF.E64-E81. Manier S, Salem KZ, Park J, Landau DA, Getz G, Ghobrial IM. Genomic complexity of multiple myeloma and its clinical implications. Nat Rev Clin Oncol. 2017;14(2):100-13. Available from: doi:10.1038/nrclinonc.2016.122. International Myeloma Foundation. Understanding MGUS and smoldering multiple myeloma [Internet]. 2018. Available from: https://www.myeloma. org/sites/default/files/resource/umgus_smm.pdf [cited 2019 Oct 22]. Michels TC, Petersen KE. Multiple myeloma: Diagnosis and treatment. Am Fam Physician. 2017;95(6):373-83. Available from: https://www.aafp.org/ afp/2017/0315/p373. Koshiaris C, Van den Bruel A, Oke JL, Nicholson BD, Shephard E, Braddick M, et al. Early detection of multiple myeloma in primary care using blood tests: A case–control study in primary care. BR J Gen Pract. 2018;68(674):58693. Available from: doi:10.3399/bjgp18X698357. Dehghanifard A, Kaviani S, Abroun S, Mehdizadeh M, Saiedi S, Maali A, et al. Various signaling pathways in multiple myeloma cells and the effects of treatment on these pathways. Clin Lymphoma, Myeloma Leuk. 2018;18(5):31120. Available from: doi:10.1016/j. clml.2018.03.007. Oyajobi BO. Multiple myeloma/ hypercalcemia. Arthritis Res Ther. 2007;9(1):S4. Available from: doi:10.1186/ar2168. Mittelman M. The implications of anemia in multiple myeloma. Clin Lymphoma. 2003;4(1):23-9. Available from: doi:10.3816/clm.2003.s.005. Fandrey J, Jelkmann WEB. Interleukin-1 and tumor necrosis factor-alpha inhibit erythropoietin production in vitro. Annals of the New York Academy of Sciences. 1991;628(1): 250-55. Available from: doi:10.1111/j.1749-6632.1991. tb17252.x Kim M, Ju YS, Lee EJ, Kang HJ, Kim HS, Cho HC, et al. Abnormalities in Chromosomes 1q and 13 independently correlate with factors of poor prognosis in multiple myeloma. Ann Lab Med. 2016;36(6):573-82. Available from: doi:10.3343/alm.2016.36.6.573. Kazandjian D. Multiple myeloma epidemiology and survival, a unique malignancy. Semin Oncol. 2016;43(6):67681. Available from: doi:10.1053/j. seminoncol.2016.11.004. Kuznetsova IS, Labutina EV, Hunter N. Radiation risks of leukemia, lymphoma and multiple myeloma incidence in the mayak cohort: 1948–2004. PLoS One. 2016;11(9):e0162710. Available from: doi:10.1371/journal. pone.0162710. Smith-Field A, Morgan GJ, Davies F. Bortezomib (Velcade™) in the treatment of multiple myeloma. Ther Clin Risk Manag. 2006;2(3):271–9. Available from: doi:10.2147/ tcrm.2006.2.3.271. Rajkumar SV. Multiple myeloma: 2018 update on diagnosis, risk-stratification, and management. Am J Hematol. 2018;93(8):981-1114. Available from: doi:10.1002/ajh.25117. Thordardottir M, Lindqvist E, Lund S, Costello R, Burton D, Steingrimsdottir L, et al. Dietary intake is associated with risk of multiple myeloma and its precursor disease. PLoS One. 2018;13(11):e0206047. Available from: doi:10.1371/journal.

INTRODUCTION

Multiple myeloma (MM) is a build-up of tumour plasma cells in the bone marrow.1 Although considered a rare cancer, making up only ~1% of all cancer types, MM is the second most common hematologic malignancy.2 This disease starts out in the premalignant stages as monoclonal gammopathy of undetermined significance (MGUS) defined by the presence of plasma cell-made monoclonal protein (M-protein) in the blood. MGUS leads to smouldering multiple myeloma, an intermediate stage which presents a greater amount of bone marrow plasma cells and M-protein in the blood.3,4 Finally, symptomatic multiple myeloma takes the form of cancer and causes damage to the body.3

SYMPTOMOLOGY: CRAB

MM exhibits heterogeneous clinical symptoms.1 The cause for this remains uncertain, but it has been proposed that the variety of symptoms experienced is related to chromosomal abnormalities located on the tumour plasma cells.1 There is a standard set of symptoms that MM patients experience, represented by the CRAB criteria: calcium (hypercalcemia), renal impairment, anemia, and bone involvement.5 Proliferating plasma cells produce immunoglobulins, specifically paraproteins (myeloma proteins), which can result in plasma hyperviscosity and kidney damage.6 This proliferation can also cause bone marrow suppression and hypercalcemia.6 These resulting conditions are reflected in the symptoms, such as fatigue as a result of anemia, or headaches due to hyperviscosity.6 Due to the heterogeneity of symptoms, diagnosis of MM requires patients to exhibit these general symptoms, as well as abnormal plasma cells in the bone marrow, bone lesions, and/or a monoclonal protein in the serum or urine.5 Of all common cancers, MM has the longest interval of time between presentation of primary symptoms and diagnosis.

SIGNALING PATHWAYS

There is a variety of signalling pathways involved in apoptosis, cell growth, survival, and drug resistance of MM cells. One family of dimer transcription factors called NF-KB, is involved in intracellular functions such as apoptosis, inflammation, angiogenesis, and immunity.7 Mutations and pro-inflammatory cytokines in the microenvironment of MM cells activate this NF-KB pathway, while several clinical drugs aim to inhibit it to overcome drug resistance and prompt apoptosis.7 Hypercalcemia present in MM occurs due to an increase of osteoclastic bone resorption as a result of local cytokine release by MM cells.8 Bone resorption further initiates the entrance of calcium into extracellular fluid.8 While the development of anemia in MM patients can be explained through several factors, the most common factor is inadequate production of erythropoietin (EPO), a hormone synthesized by the kidney that plays a significant role in the synthesis of red blood cells.9 The production of inflammatory cytokines such as Interleukin-1 and tumor necrosis factor in MM patients downregulates EPO production, which leads to anemia.9,10

Risk Factors

Although the cause of MM is uncertain, recent studies have sought to investigate the complex interrelation between genetic, demographic, and environmental factors that may increase the risk of developing MM.3 Genetic findings often vary across studies due to the complexity of the disease; however, research has primarily focused on chromosomes 1 and 13. 35-40% of MM cases exhibited trisomy 1q, where three copies of the q arm on chromosome 1 are present instead of the normal two.3,11 Furthermore, an additional 45-50% of cases exhibited chromosome 13q deletion, possessing only 1 copy of the q arm on chromosome 13.11 Trisomy 1q was associated with increased immunoglobin levels, while 13q deletion was correlated with lower levels of tumour-suppressor gene RB1. These are characteristics associated with MM and cancer development.11 Considering these abnormalities were not present in all cases, genetic factors alone may not be sufficient in determining MM risk. MM is most common among elderly populations with median age of diagnosis ranging from 66 to 70; patients under 30 comprised only 0.02% of cases.12 Studies show that MM occurs twice as frequently in African-American populations than among European-Americans, suggesting that ethnicity may also play a role. As with most cancers, exposure to external radiation appears to increase the risk of MM, as seen in a 1948-2004 cohort study of Russian plutonium production workers.13

suppression of tumour suppression gene RB1

Treatment

pathoprofile

Treatment of MM depends on a patientâ&#x20AC;&#x2122;s eligibility for autologous stem cell transplantation (ASCT) as assessed by the clinician. In ASCT- eligible patients, blood-forming stem cells are harvested from a patientâ&#x20AC;&#x2122;s bone marrow following 3-4 cycles of pharmaceutical induction therapy. This is often achieved with Bortezomib, a proteasome inhibitor that disrupts the cell cycle and induces apoptosis.14 Patients may then be given dexamethasone to improve the sensitivity of myeloma cells, before treatment with high doses of chemotherapy.14 The stem cells are then released back into the bloodstream to allow the bone marrow to produce new blood cells.14 Patients ineligible for ASCT will be provided with 8-12 cycles of Bortezomib and dexamethasone.14 Both ASCT and non-ASCT patients then undergo maintenance therapy with lenalidomide in order to prevent the growth of residual myeloma cells.15 Although these treatments have proved quite effective in improving prognosis, almost all MM patients eventually relapse and require multiple treatment cycles throughout their lives. In addition to pharmaceutical interventions, studies have shown that low inflammatory diets, high in fruits and vegetables, may also decrease risk for MM.16 REVIEWED BY: TOM KOUROUKIS Dr. Kouroukis is an Associate Professor in the Department of Oncology and Division Head of Malignant Hematology at McMaster University. His clinical interests include the treatment of older patients with hematological malignancies. pone.0206047.

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3D PRINTING IN SURGERY AND MEDICINE AUTHORS: Adrian Wong & Albert Stancescu

Biotech Blueprint

INTRODUCTION

Three-dimensional (3D) printing involves the creation of a 3D product through combining and depositing materials (i.e. plastic, metals, and ceramics) in layers.1 Since the first patent was made by Charles Hull in 1984, advances in 3D printing technology have allowed it to be utilized in a variety of fields, including engineering and medicine.2 Specifically, 3D printing technology has been useful in refining surgical techniques. 3D-printed anatomical models, surgical implants, and prosthetic limbs have proven to be beneficial in daily medical practice. Organ resection, reconstruction surgeries, reparative procedures, and organ transplantations have also been improved through this technology.1,2,3

biotech blueprint

3D PRINTING METHODS IN A SURGICAL CONTEXT

The first step in 3D printing in a surgical context involves creating the 3D model itself, usually through computer-aided design (CAD) software or the conversion of data from computerized tomography (CT) or magnetic resonance (MR) scans.3 Next, the images are set through post-processing and segmentation steps to extract anatomical regions of interest. These regions are then transcribed into a computerized model with which tessellated triangles are used to approximate the contours of the model.4 This file can then be used by CAD software for further processing to improve the modelâ&#x20AC;&#x2122;s design, before being sent to the printer for producing a tangible 3D object.3 The quality of the model is highly dependent on the quality of the original scans, with higherresolution models scans producing higher-quality models.3

ARTIST Michelle Chen

Chromosomal Translocation

M E D U C AT O R

| DECEMBER 2019

A NEW AGE FOR IMPLANTS Research surrounding 3D-printed bone implants and scaffolds have shown advances in combating osseointegration.9,10 Any potential bone substitute requires a high degree of osseointegration to facilitate the bond between new bone and the scaffold biomaterial.10 Although this requirement has limited the available pool of biomaterials for use in bone implants, advances in inkjet 3D printing with titanium alloy powders have enabled research into osseointegration with 3D modeled structures.9

8 9

Beyond bone graft physiology, it is important to consider the functionality and survival of bone grafts post-implantation. All bone implants must minimize the impact of Wolff â&#x20AC;&#x2122;s Law: under physical stress, bone is deposited in areas where stress occurs the most and is removed where it occurs the least.11 Consequently, any differences between the stress-bearing properties of the implant and surrounding bone tissue may result in long-term bone loss. Therefore, while bone graft integration is critical in promoting bone regrowth, it is also important to consider the load-bearing and stress-shielding mechanical properties as well as orientation of the graft in question.12, 13 To date, clinical applications of 3D-printed implants are few and far between, with most surgeons in this field applying bone tissue models to preoperational planning and surgical training.11 However, studies utilizing 3D-printed scaffolds for bone regeneration seem promising, showing evidence that cellular proliferation and bone matrix formation are supported by these bone scaffolds.12 Yet, with multiple methods to create 3D-printed tissue scaffolds that maximize osseointegration, more research is still required to generate reliable tissue scaffolds that facilitate bone regrowth while avoiding host rejections.10,14,15,16

MOVING TOWARDS IMPROVING SURGICAL PREPARATION To better understand the extent of a disease in a patient prior to surgery, surgeons will rely on two-dimensional (2D) images obtained from X-rays, CT scans, or MRI scans.4 However, 2D depictions may differ from what the surgeon actually encounters during an operation.5 This process intrinsically requires a high degree of visualization on the surgeon’s part, which may be difficult for inexperienced surgeons.4 Furthermore, a surgeon’s understanding of a patient’s anatomy may be complicated by variation or changes in shape due to disease progression.5 While 3D renderings of radiography, CT, and MRI scans have been developed to better visualize complex pathologies, they lack tangibility.4,5 These problems have prompted the use of 3D printing technology in pre-operational settings. Patient-specific anatomical models provide a tangible structure to manipulate —as opposed to 2D and 3D renderings of images —while maintaining accuracy.5 The introduction of 3D-printed anatomical models has markedly improved surgical preparation. For instance, they can be used by surgeons to perform “trial runs” for surgeries, allowing surgeons to predict potential technical challenges due to anatomical variations.2,5 3D-printed anatomical models have been used to prepare for surgeries in many disciplines, including cardiac and orthopedic surgery.6, 7 In these settings, researchers report that the models had been useful in planning and improving physician understanding of surgical procedures.6,7 Additionally, a study showed that out of 22 patients undergoing hepatectomy, 20 operations were successful the first time with the help of 3D model preparation with only two requiring further open surgery.3 There are limitations to the use of 3D printing technology for surgical preparation. Firstly, utilizing this technology necessitates specific skills that are not typically taught to surgeons.8 Furthermore, it has been reported that the accuracy of 3D-printed models was insufficient in more precise surgeries, as certain anatomical structures are too fine to be successfully replicated.3 Another concern among researchers is the cost of implementing 3D printing technology; however, with the increasing accessibility and the decreasing cost of 3D printing, this concern will likely diminish over time.3,8

Despite recent advancements in 3D printing technology, regulations for its use in surgery are virtually nonexistent. Currently, there are no formal protocols for using surgical 3D printing that can ensure a high degree of quality control. Such regulations, if implemented, must ensure that only the highest quality implants are used during surgery.18 The adoption of these protocols is therefore the key to increasing surgeons’ trust in this technology, so that one day it may truly find its place in the surgical toolbox.

biotech blueprint

THE FUTURE OF 3D PRINTING IN SURGERY Given the successes of 3D printing in many aspects of surgery, there is much opportunity for growth. The money and time needed to print 3D models, implants, and prostheses are likely to decrease in comingyears, improving the clinical accessibility of this technology.5 Furthermore, as research advances, a wider range of materials will permit better simulation of realistic tissues; surgeons will increasingly have access to better practice specimens, and consequently better surgical outcomes will be produced.17

Reviewed By: Dr. Janie Wilson Dr. Wilson is a Professor in the Department of Surgery at McMaster University, having completed a Ph.D. in Biomedical Engineering from Dalhousie University. Her expertise includes orthopaedic biomechanics, musculoskeletal biomechanics, gait analysis, osteoarthritis, orthopaedic surgery, implants and surgery outcomes.

14. Tellisi N, Ashammakhi NA, Billi F, Kaarela O. Three dimensional printed bone implants in the clinic. J Craniofac Surg. 2018;29(8):2363-7 Available from: doi:10.1097/ SCS.0000000000004829. 15. Helguero CG, Mustahsan VM, Parmar S, Pentyala S, Pfail JL, Kao I, Komatsu DE, Pentyala S. Biomechanical properties of 3D-printed bone scaffolds are improved by treatment with CRFP. J Orthop Surg Res. 2017;12(1):195. Available from: doi:10.1186/s13018-017-0700-2. 16. Bose S, Vahabzadeh S, Bandyopadhyay A. Bone tissue engineering using 3D printing. Mater Today. 2013;16(12):496-504. Available from: doi:10.1016/j. mattod.2013.11.017. 17. Ganguli A, Pagan-Diaz GJ, Grant L, Cvetkovic C, Bramlet M, Vozenilek J, et al. 3D printing for preoperative planning and surgical training: A review. Biomed Microdevices. 2018;20(3):65. Available from: doi:10.1007/s10544-018-0301-9. 18. Kwok JKS, Lau WHL, Zhao ZR, Yu PSY, Ho JYK, Chow SCY, et al. Multidimensional printing in thoracic surgery: Current and future applications. J Thorac Dis. 2018;10(Suppl 6):S756-63. Available from: doi:10.21037/jtd.2018.02.91.

| DECEMBER 2019

from: doi:10.3941/jrcr.v6i1.889. 8. Martelli N, Serrano C, van den Brink H, Pineau J, Prognon P, Borget I, et al. Advantages and disadvantages of 3-dimensional printing in surgery: A systematic review. Surgery. 2016;159(6):1485-500. Available from: doi:10.1016/j. surg.2015.12.017. 9. Popov VV, Muller-Kamskii G, Kovalevsky A, Dzhenzhera G, Strokin E, Kolomiets A, et al. Design and 3D-printing of titanium bone implants: Brief review of approach and clinical cases. Biomed Eng Lett. 2018;8(4):33744. Available from: doi:10.1007/s13534-018-0080-5. 10. Martinez-Marquez D, Mirnajafizadeh A, Carty CP, Steward RA. Application of quality by design for 3D-printed bone prostheses and scaffolds. Plon. 2018;13(4):e0195291. Available from: doi:10.1371/journal.pone.0195291. 11. Salter RB. Textbook of disorders and injuries of the musculoskeletal system. Baltimore: Williams & Wilkins; 1970. 12. Van Oosterwyck H, Duyck J, Vander Sloten J, Van der Perre G, De Coomans M, Lieven S, et al. The influence of bone mechanical properties and implant fixation upon bone loading around oral implants. Clin Oral Implants Res. 1998;9(6):407-18. Available from: doi:10.1034/j.1600-0501.1996.090606.x. 13. Oppenheimer AJ, Tong L, Buchman SR. Craniofacial bone grafting: Wolff’s law revisited. Craniomaxillofac Trauma Reconstr. 2008;1(1):49-61. Available from: doi:10.1055/s-0028-1098963.

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1. Ventola CL. Medical applications for 3D printing: Current and projected uses. P T. 2014;39(10):704-711. Available from: https://www.ncbi.nlm.nih.gov/ pubmed/25336867 [cited 2019 Nov 6]. 2. Shilo D, Emodi O, Blanc O, Noy D, Rachmiel A. Printing the future – Updates in 3D printing for surgical applications. Rambam Maimonides Med J. 2018;9(3):e0020. Available from: doi:10.5041/RMMJ.10343. 3. Li C, Cheung TF, Fan VC, Sin KM, Wong CWY, Leung GKK. Applications of three-dimensional printing in surgery. Surg Innov. 2017;24(1):82-8. Available from: doi:10.1177/1553350616681889. 4. Tack P, Victor J, Gemmel P, Annemans L. 3D-printing techniques in a medical setting: A systematic literature review. Biomed Eng Online. 2016;15:115. Available from: doi:10.1186/s12938-016-0236-4. 5. Pugliese L, Marconi S, Negrello E, Mauri V, Peri A, Gallo V, et al. The clinical use of 3D printing in surgery. Updates Surg. 2018;70(3):381-8. Available from: doi:10.1007/ s13304-018-0586-5. 6. Schmauss D, Gerber N, Sodian R. Three-dimensional printing of models for surgical planning in patients with primary cardiac tumors. J Thorac Cardiovasc Surg. 2013;145(5):1407-8. Available from: doi:10.1016/j.jtcvs.2012.12.030. 7. Tam MD, Laycock SD, Bell D, Chojnowski A. 3-D printout of a DICOM file to aid surgical planning in a 6 year old patient with a large scapular osteochondroma complicating congenital diaphyseal aclasia. J Radiol Case Rep. 2012;6(1):31-7. Available

NEUROABSTRACTS EXAMINING THE INFLUENCE OF INSTAGRAM STATUS ON SENSE OF AGENCY FELICIA CHIAPPETTA1 & SUKHVINDER S. OBHI1

Department of Psychology, Neuroscience & Behaviour, McMaster University

neuroabstracts

Power is an interpersonal construct where the powerful have control over the social, economic, and physical resources of others. Previous studies have found that power priming influences one’s sense of agency (SoA). SoA is the feeling of control over your actions and the outcomes of those actions, as indexed by the intentional binding task. The intentional binding paradigm suggests that individuals experiencing a higher sense of agency tend to make shorter time interval estimates between their actions and outcomes. Previous research using the intentional binding task has found that interval estimates tend to be longer for individuals of low power compared to those of high power. With the increasing popularity of social media use, people look to these sites for social connection, opening a new avenue for social status hierarchy. This hierarchy exists on Instagram, a social media network where people follow others

and have others following them. This follower/following (F/F) ratio is an index of online power, where users can be categorized as “leaders” (have more followers than they are following) or “followers” (following more people than they have following them). The aim of the current study is to investigate whether online social status, indexed by the F/F ratio, can alter one’s SoA. Participants will complete a baseline and post-prime block of the intentional binding task. In between blocks, they will write down their F/F ratio, in place as a prime for their sense of online status. When comparing post-prime block estimates, we predict that Instagram “leaders” will give shorter overall estimates, whereas “followers” will give longer overall estimates. Such evidence could validate and extend previous research on the effects of real-life power on SoA, suggesting that online and real-life power may have concordant effects.

GENE EXPRESSION ANALYSIS UPON ADMINISTRATION OF PAOPA AND ITS POTENCY TO TREAT SYMPTOMS OF SCHIZOPHRENIA VIDHI PATEL1, HETSHREE JOSHI2, RAM MISHRA3

Department of Biochemistry, McMaster University Department of Medicine, University of Toronto Department of Psychiatry and Behavioural Neuroscience, McMaster University

M E D U C AT O R

| DECEMBER 2019

Schizophrenia is a chronic mental disorder that affects more than 21 million people worldwide. 3(R)-[(2(S)-pyrrolidylcarbonyl) amino]-2-oxo-1-pyrrolidineacetamide (PAOPA), the positive allosteric modulator of the dopamine D2 receptor, has previously demonstrated the potential to treat the negative cognitive symptoms associated with schizophrenia. To investigate the molecular effects of the drug, it was hypothesized that PAOPA administration would significantly alter gene expression in the striatum of Sprague-Dawley rats. RNA sequencing was performed to analyze differential gene expression and the results revealed a significant overexpression in the calciumbinding protein, parvalbumin (PVALP). PVALP was further investigated because previous research published by our lab has

shown decreased levels of PVALP mRNA in schizophrenia. RTqPCR demonstrated that PAOPA significantly increased mRNA expression of PVALB in vivo. PVALB can modulate gammaaminobutyric acid neuron activity and potentially control impaired gamma-band oscillations, which are an underlying cause of cognitive schizophrenic symptoms. The implication of PVALB in schizophrenia provides further support for PAOPA as a novel therapeutic which has the potential to treat schizophrenic symptoms. Further research should investigate changes in PVALB protein levels in an animal model of schizophrenia and elucidate the molecular mechanism through which the drug alters PVALB expression.

Acknowledgments: This research was supported by CIHR and NSERC and the constant effort of members of the Mishra Lab. Application of PAOPA is secured by the US patent: PCT/CA2011/000968.

ARTIST MANREET DHALIWAL

AN EXPLORATORY STUDY OF IMPULSIVITY DIFFERENCES BETWEEN RECREATIONAL AND MEDICINAL CANNABIS USERS AMIRA BENAINI1, JAMES MACKILLOP2,3,4, IRIS BALODIS2,3,4 Department of Psychology, Neuroscience & Behaviour, McMaster University Department of Psychiatry & Behavioural Neuroscience, McMaster University 3 Peter Boris Centre for Addictions Research, St. Josephâ&#x20AC;&#x2122;s Healthcare Hamilton 4 Michael G. DeGroote Centre for Medicinal Cannabis Research

Methods: To investigate differences in impulsivity between medicinal and recreational cannabis users, a community sample of 55 regular cannabis users was recruited from the Hamilton area. Participants were grouped into a recreational cannabis use group (n=20), a combined medicinal+recreational use group (n=29) and a medicinal-only use group (n=6). Participants completed the Barratt Impulsivity Scale (BIS-11), a validated self-report measure of impulsivity including an attentional, motor, and non-planning subscale.

Results: Total and subscale impulsivity scores did not significantly differ between the three groups. There was a trend for the medicinal-only users to display lower total BIS-11 and motor impulsivity scores than the other two groups (p-values<0.075). Conclusions: These exploratory trends suggest that impulsivity might differ between recreational, medicinal+recreational, and medicinal-only cannabis users. Medicinal-only users may have lower impulsivity rates, driven by lower motor impulsivity. Larger, demographically-matched samples should be studied to better understand the difference in impulsivity amongst recreational and medicinal cannabis users, as well as transitions to problematic use. Future studies can also examine specific features of cannabis use, including motives for use, drug potency, and other forms of impulsivity. Uncovering differences between cannabis-using subpopulations can help develop better policies and regulations surrounding the use of the drug in different settings.

neuroabstracts

Background: Individuals who use cannabis demonstrate high levels of impulsivity, which is defined by rapid action without careful consideration and planning. Previous studies have examined a link between cannabis use and increased impulsivity, but differences between medicinal and recreational cannabis users have not yet been explored. It is important to understand impulsivity differences in cannabis-using subgroups, as this trait is associated with increased vulnerability to substance use disorders.

GENDER EFFECTS IN THE HUMAN BRAIN: A VOLUMETRIC IMAGING ANALYSIS CONNOR M. SCHMID1, CHRISTIE E. MACLEOD1, NICHOLAS A. BOCK1 Department of Psychology, Neuroscience & Behaviour, McMaster University

| DECEMBER 2019

between gender and brain structure volume, while considering age as a cofactor. Statistical analysis revealed a significant difference in volume between genders in 37 of the regions analyzed, with males having larger volumes by 3-28%. Significant effects of age were identified in 21 regions, with no significant age by gender interactions identified. Our findings may provide insight into different areas of research, such as cognitive function and gender-based psychiatric disorders.

M E D U C AT O R

A major ongoing research question in neuroscience is how the structure of the brain underlies and influences human behaviour. Studies have consistently found that on average, males have larger brain volumes than females. The current study analyzed 1113 whole-head anatomical magnetic resonance images from the Human Connectome Project to identify volumetric gender differences across 39 subcortical structures in the human brain. The subjects represent young adults aged 22-37 years. Using an ANCOVA statistical framework, we explored the relationship

EDITED BY: MEERA CHOPRA & JAMES YU

doi:10.35493/medu.36.12

VIEWPOINTS

Does the CMPA serve the interests of patients? ROHAN AANANTH1 , MILENA CIOANA 2 , NURI SONG 3

Bachelor of Health Sciences (Honours) Program Class of 2021, McMaster University Bachelor of Health Sciences (Honours) Program Class of 2020, McMaster University 3 Bachelor of Health Sciences (Honours) Program Class of 2022, McMaster University 1 2

M E D U C AT O R

| DECEMBER 2019

viewpoints

INTRODUCTION

Medical malpractice consists of taking legal action against a healthcare professional when the professional deviates from the standard of care, creating harm or injury to patients. In Canada, the majority of physicians receive medical malpractice insurance and legal representation from the Canadian Medical Protective Association (CMPA). The CMPA claims that effective medical liability protection enables Canadian physicians to practise medicine with confidence and focus on providing the best quality of care for their patients. However, whether the CMPA improves and maintains quality of patient care remains a debated topic. Critics say that the power and vast financial resources of the CMPA allow it to put on an aggressive legal defence with which patients cannot compete. Since the 1970s, the number of lawsuits in Canada has decreased.1 This ViewPoints piece will examine whether the CMPA truly protects the interests of patients.

THE CMPA IS BENEFICIAL FOR PATIENTS

The legal support and liability coverage from the CMPA provides physicians with greater autonomy and reduced fear of litigation. The fear of litigation can make doctors question their clinical judgement and order unnecessary tests. By discouraging defensive medicine —the practice of making treatment decisions to avoid litigation —the CMPA ultimately improves patient care.2 In the United States, physicians do not receive legal protection from a central body like the CMPA; therefore defensive medicine is highly prevalent.3 For example, in 2005, a study reported that 92% of Pennsylvanian physicians (n = 842) had ordered an unnecessary medical test at least once to cover for liabilities.4 Such measures to deter lawsuits may be justified if they were to truly benefit the health of patients, but research shows that the practice of defensive medicine does not necessarily lead to improvements in health outcomes.5 By addressing the fears of legal disputes, the CMPA protects physician autonomy to exercise clinical judgement in their patients’ best interests.6,7 Some may argue that the existence of the CMPA harms patients, as their legal prowess deters patients from pursuing legal action, and consequently enables malpractice.1 However, there is evidence to show that physicians who face legal action do not actually become less negligent.8 Malpractice is not usually the fault of one individual —it is often due to compounding issues inherent in the system.9 Hence, malpractice is best resolved through implementing systemic changes and the CMPA’s power and influence are key to doing so. The CMPA ensures patient safety through the revision of healthcare delivery guidelines while educating physicians on how to avoid procedures that would pose a high risk to patients.10 For instance, the CMPA has identified key areas of risk in obstetrical care and created 18 action-oriented recommendations for physicians focused on improving the safety of care for Canadian mothers and babies.11 Along with its risk management and education efforts, the CMPA uses its understanding of the medical liability system to advocate for improvements to patient safety. One such example is the advisory service that the CMPA provides to governmental committees on medico-legal matters.12 Notably, the CMPA has 1. CBC. As fewer patients sue their doctor, the rate of winning malpractice suits is dropping too [Internet]. 2019 Apr 18. Available from: https://www.cbc.ca/news/health/medical-malpractice-doctors-lawsuits-canada-1.4913960 [cited 2019 Oct 29]. 2. Cano-Urbina J, Montanera D. Investigating defensive medicine: The role of access. Toronto: Canadian Centre for Health Economics; 2015. 24 p. Report No.:150012. 3. American College of Physicians. Medical malpractice insurance [Internet]. 2019. Available from: https://www.acponline.org/aboutacp/about-internal-medicine/career-paths/residency-career-counseling/guidance/medical-malpractice-insurance [cited 2019 Oct 29]. 4. Studdert DM, Mello MM, Sage WM, DesRoches CM, Peugh J, Zapert K, et al. Defensive medicine among high-risk specialist physicians in a volatile malpractice environment. JAMA. 2005;293(21):260917. Available from: doi:10.1001/jama.293.21.2609.

consulted on the Canadian federal government’s “Special Joint Committee on Physician-Assisted Dying,” where it advocated for equal legal protection for patients and their physicians regarding Medical Assistance in Dying.13 Although the media recurrently portrays the CMPA as providing legal defence to negligent doctors, physicians whose cases lack merit are not actually supported by the CMPA.14 Moreover, the CMPA directly provides compensation to patients proven to have been harmed by negligent physician care.15 An evaluation of the CMPA’s entire body of work would therefore demonstrate conclusively that its interests do not clash with those of the patients —rather, both of their interests are aligned.

THE CMPA ADVERSELY AFFECTS PATIENTS

the physician’s favour.1,16 Paul Harte, a Canadian medical malpractice lawyer, says that, “it is in the CMPA’s best interest to drag out some of these cases because it acts as a deterrent for suing other doctors.”1 These hurdles imposed by the CMPA often result in victims of medical malpractice not pursuing legitimate claims.1,16 In fact, between 2013 and 2017, 55.2% of malpractice cases in Canada were ultimately dismissed, discontinued, or abandoned.1 In doing so, the CMPA buries malpractice cases that would otherwise help inform systemic changes to physician practice guidelines.18 An analysis of the past 40 years of the CMPA’s annual records reveals that, although the number of doctors has exponentially increased, the rate of patients suing has decreased.1 This might indicate that the CMPA is so effective at protecting doctors that it harms patients. Through discussing missteps and searching for root causes of failures, the medical field might see improvements in quality of care and patient safety. By defending doctors and silencing the patient voice, the CMPA prevents this reflective process necessary for the improvement of healthcare. An obvious risk of this practice is that negligent behaviours will never come to light, will never be corrected, and thus will continue to hurt patients.

viewpoints

In 2012, when Anne received a spinal injection intended to treat her chronic back pain, little did she know that it would cripple her for the rest of her life. Unknowingly using a contaminated needle, Anne’s doctor injected harmful bacteria into her spinal cord, infecting it and causing permanent nerve damage. Five years later, Anne remains burdened by the repercussions of her doctor’s negligence —she can barely climb up the stairs of her own home and struggles to pay for the medication needed to relieve complications resulting from the infection. Furthermore, Anne is unable to receive the financial compensation she Although the CMPA claims to be beneficial for both physicians rightfully deserves. She wonders why the medical malpractice and patients, it is clear that the CMPA prioritizes the protection lawsuit she filed against her physician years earlier has dragged of its physicians over improving patient care. Thus, it jeopardizes on, given there is no dispute that her doctor is responsible for the safety of patients and the future of our healthcare system.15 her injury. She realizes that the slow pace of legal proceedings Ultimately, the CMPA makes the process of achieving justice is due to the aggressive and unethical legal defence advanced and compensation more difficult for patients victimized by by her doctor’s insuring body, the CMPA. Exhausted by an medical malpractice. The association should shift its mandate unwinnable legal battle, and running out of money to pay her to better protect the interests of patients. lawyer, Anne wonders if she should simply abandon her case.16 Such a scenario is not uncommon in Canadian medical malpractice litigation. Although the CMPA claims to help maltreated individuals receive compensation, the actions of the CMPA reflect poor accountability to patients and and translates to worsened quality of care.15 The CMPA’s mandate is to defend doctors accused of medical malpractice.17 However, the length to which the CMPA will go to legally protect its physicians is problematic. The CMPA’s lawyers litigate on behalf of physicians regardless of the defensibility of their actions.1

13. The Canadian Medical Protective Association. Special joint committee consultations on physician-assisted dying [Internet]. 2016. Available from: https://www.cmpa-acpm.ca/static-assets/pdf/research-and-policy/ public-policy/2016-02-03_Submission_to_Joint_Committee_on_PAD_C-14-e.pdf [cited 2019 Nov 6]. 14. The Canadian Medical Protective Association. Medical-legal issues to consider with individual contracts [Internet]. Available from: https://www.cmpa-acpm.ca/en/membership/protection-for-members/principles-of-assistance/medico-legal-issues-to-consider-with-individual-contracts [cited 2019 Nov 6]. 15. The Canadian Medical Protective Association. What we do [Internet]. Available from: https://www.cmpa-acpm.ca/en/about/what-we-do [cited 2019 Nov 6]. 16. CTV News. Patients who suffer from medical errors face ‘rigged system,’ critics say. [Internet]. 2019 Jul Available from: https://www.ctvnews.ca/health/patients-who-suffer-from-medical-errors-face-rigged-system-critics-say-1.4507664 [cited 2019 Nov 6]. 17. The Canadian Medical Protective Association. Submissions to standing committee - Bill 87, Protecting Patients Act, 2016 [Internet]. 2017 May 8. Available from: https://www.cmpa-acpm.ca/static-assets/pdf/ research-and-policy/public-policy/2017-05-08_Submissions_to_Standing_Committee-Bill_87_Protectin. pdf [cited 2019 Nov 6]. 18. Mondaq. Less patients are suing doctors for medical malpractice - but why? [Internet]. 2019 May 14. Available from: http://www.mondaq.com/canada/x/804920/Professional+Negligence/Less+Patients+are+Suing+Doctors+for+Medical+Malpractice+But+Why [cited 2019 Nov 6].

| DECEMBER 2019

5. Frakes MD, Gruber J. Defensive medicine: Evidence from military immunity. Am Econ J Econ Policy. 2019;11(3):197-231. Available from: doi:10.1257/pol.20180167. 6. Coyte PC, Dewees DN, Trebilcock MJ. Medical malpractice — The canadian experience. N Engl J Med. 1991;324:89-93. Available from: doi:10.1056/NEJM199101103240204. 7. The Canadian Medical Protective Association. 2018 CMPA annual report highlights [Internet]. 2018. Available from: https://www.cmpa-acpm.ca/en/about/annual-report [cited 2019 Nov 4]. 8. The Canadian Medical Protective Association. Reimbursement of CMPA fees: A benefit to physicians and their patients [Internet]. 2010. Available from: https://www.cmpa-acpm.ca/en/advice-publications/ browse-articles/2010/reimbursement-of-cmpa-fees-a-benefit-to-physicians-and-their-patients [cited 2019 Nov 4]. 9. Carlson RJ. A conceptualization of a no-fault compensation system for medical injuries. Law Soc Rev. 1973;7(3):329. Available from: doi:10.2307/3052919. 10. The Canadian Medical Protective Association. Medical-legal handbook for physicians in Canada [Internet]. 2016. Available from: https://www.cmpa-acpm.ca/static-assets/pdf/advice-and-publications/ handbooks/com_16_MLH_for_physicians-e.pdf [cited 2019 Nov 6]. 11. Lefebvre G, Calder LA, Gorter RD, Bowman CL, Bell D, Bow M. Recommendations from a national panel on quality improvement in obstetrics. J Obstet Gynaecol Can. 2019;41(5):653–9. Available from: doi:10.1016/j.jogc.2019.02.011. 12. The Canadian Medical Protective Association. Public policy, submissions, and responses [Internet]. Available from: https://www.cmpa-acpm.ca/en/research-policy/public-policy [cited 2019 Nov 6].

M E D U C AT O R

Patients, on the other hand, are often not able to afford legal assistance, much less endure the financial burden of paying lawyers to represent them.1,16 More often than not, the CMPA exploits its financial advantage to tip the scales of justice in

ARTIST RUBY ZHENG

CRITICAL REVIEW

Tick-borne diseases: From well-known Lyme disease to deadly Powassan encephalitis

doi:10.35493/medu.36.14

JERRY CHANG

Bachelor of Health Sciences (Honours), Class of 2022, McMaster University Correspondence: changj30@mcmaster.ca

INTRODUCTION

Throughout central to eastern Canada, ticks are frequently found during warmer times of the year, typically in late spring to early fall. In Hamilton, the most commonly encountered tick is the American dog tick (Dermacentor variabilis), which does not transmit any tick-borne diseases. However, observation of tick populations in Hamilton in 2017 reported new evidence of blacklegged tick populations.1 Blacklegged ticks, or Ixodes scapularis, are the primary vector for Lyme disease and other infections, including anaplasmosis, babesiosis, ehrlichiosis, tularemia, and Powassan encephalitis.2

LYME DISEASE

| DECEMBER 2019

Diagnosing Lyme disease can prove challenging. In early stages of Lyme disease, immunoglobulin M and immunoglobulin G may not have developed, making serologic testing a poor determinant of disease.11 Serologic testing is more useful once the disease has progressed. The Centres for Disease Control and Prevention and the Infectious Diseases Society of America recommend using an enzyme-linked immunosorbent assay (ELISA) along with a Western blot, should the ELISA yield positive or non-specific results.12 Treatment of Lyme disease typically involves oral antibiotics in early localized stages of disease and intravenous regimens in patients with neurologic symptoms. Oral doxycycline is preferred, though macrolide antibiotics can be used when a patient cannot tolerate doxycycline.11

M E D U C AT O R

Lyme disease is the most well-known tick-borne disease worldwide, and the most prevalent tickborne disease in North America. By 2020, 80% of central and eastern Canada’s population could live in high risk areas for Lyme disease.3 Borrelia burgdorferi, a spirochete bacteria transmitted to humans through tick bites, is the primary cause of Lyme disease. Ticks must be attached for at least 24 hours before infection becomes possible. Infection is certain should the tick carrying B. burgdorferi be attached for over 72 hours.4 Early discovery and removal of ticks is the best way to prevent contraction of Lyme disease.4

Lyme disease manifests through progressive stages. The earliest stage of infection is characterized by a localized skin lesion known as erythema migrans. This lesion, also commonly called a target lesion or “bulls-eye” rash, typically appears within a month of infection. Up to 80% of patients with Lyme disease develop this rash and approximately 1020% of patients develop multiple lesions.5,6 Early Lyme disease may sometimes be confused with other conditions involving similar lesions, but the condition is distinguished by specific erythema migrans lesions. Such lesions are typically found on areas of the lower extremities and trunk such as the axilla, inguinal region, and lower abdomen.7 In the early disseminated phase, Lyme disease can affect the nervous, cardiovascular, and musculoskeletal systems. If left untreated, Lyme disease may progress to the late disseminated stage, manifesting as arthritis or severe neurological damage.8 Even after successful treatment, patients may suffer from post-Lyme disease syndrome; non-specific symptoms such as fatigue may persist. As well, patients may suffer from chronic Lyme disease, a condition where the patient continues to show symptoms, yet it is unclear whether the patient still suffers from Lyme disease.9,10 Thus, early diagnosis of Lyme disease is crucial to managing and preventing long-term complications.

critical review

ABSTRACT Tick-borne diseases are prevalent throughout central and eastern Canada, including Ontario. Of the many infections transmissible through tick bites, Lyme disease is the most well-known. However, lesser known tick-borne diseases, such as Powassan encephalitis, can have debilitating, or even lethal complications. The incidence of all tick-borne infections has increased over the past decade, with reported cases of Lyme disease seeing a tenfold increase. Identifying both Lyme disease and Powassan encephalitis can be challenging, as patients only show abnormalities in the later stages of disease progression. Even following successful treatment for Lyme disease, patients may continue to suffer from chronic disease sequelae. As tick populations continue to increase, primary prevention is the most effective method of combating tick-borne diseases.

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15.

16.

Sanderson M, Lindsay LR, Campbell TM, Morshed. A case of Powassan encephalitis acquired in southern Quebec. CMAJ. 2018;190(50):E1478-80. Available from: doi:10.1503/cmaj.180905. Centers for Disease Control and Prevention. Outbreak of Powassan encephalitis—Maine and Vermont, 19992001. MMWR Morb Mortal Wkly Rep. 2001;50(35):761-4. Available from: https://www.ncbi.nlm.nih.gov/ pubmed/11787585 [cited 2019 Nov 6]. Gasmi S, Ogden NH, Leighton PA, Adam-Poupart A, Milord F, Lindsay LR, et al. Practices of Lyme disease diagnosis and treatment by general practitioners in Quebec, 2008-2015. BMC Fam Prac. 2017;18(1):65. Available from: doi:10.1186/s12875-017-0636-y. Flisiak R, Prokopowicz D. Antibodies against Borrelia garinii in diagnosis of Lyme borreliosis. Przegl Lek. 2000;57(3):147-9. Available from: https://www.ncbi.nlm.nih.gov/ pubmed/10909283 [cited 2019 Nov 6]. Tibbles CD, Edlow JA. Does this patient have erythema migrans? JAMA. 2007;297(23):2617-27. Available from: doi:10.1001/jama.297.23.2617. Aucott J, Morrison C, Munoz B, Rowe PC, Schwarzwalder A, West SK. Diagnostic challenges of early Lyme disease: Lessons from a community case series. BMC Infect Dis. 2009;9:79. Available from: doi:10.1186/1471-2334-9-79. Biesiada G, Czepiel J, Leśniak MR, Garlicki A, Mach T. Lyme disease: Review. Arch Med Sci. 2012;8(6):978-82. Available from: doi:10.5114/aoms.2012.30948. Hubálek Z, Halouzka J. Distribution of Borrelia burgdorferi sensu lato genomic groups in Europe, a review. Eur J Epidemiol. 1997;13(8):951-7. Available from: doi:10.1023/a:1007426304900. Wormser GP, Dattwyler RJ, Shapiro ED, Halperin JJ, Steere AC, Klempner MS, et al. The clinical assessment, treatment, and prevention of Lyme disease, human granulocytic anaplasmosis, and babesiosis: Clinical practice guidelines by the Infectious Disease Society of America. Clin Infect Dis. 2006;43(9):1089-134. Available from: doi:10.1086/508667. Marques A. Chronic Lyme disease: A review. Infect Dis Clin North Am. 2008;22(2):341-60. Available from: doi:10.1016/j.idc.2007.12.011. Wright WF, Riedel DJ, Talwani R, Gilliam BL. Diagnosis and management of Lyme disease. Am Fam Physician. 2012;85(11):1086-93. Available from: https://www.aafp.org/afp/2012/0601/ p1086.html [cited 2019 Nov 6]. Centers for Disease Control and Prevention. Recommendations for test performance and interpretation from the second national conference on serologic diagnosis of Lyme disease. MMWR Morb Mortal Wkly Rep. 1995;44(31):590-1. Available from: https://www.ncbi.nlm.nih.gov/ pubmed/7623762 [cited 2019 Nov 6]. McLean DM, Donohue WL. Powassan virus: Isolation of virus from a fatal case of encephalitis. Can Med Assoc J. 1959;80:708-11. Available from: https://www.ncbi.nlm.nih.gov/ pubmed/13652010 [cited 2019 Nov 6]. Gholam BIA, Puksa S, Provias JP. Powassan encephalitis: A case report with neuropathology and literature review. Can Med Assoc J. 1999;161(11):1419-22. Available from: http://ncbi.nlm.nih.gov/ pmc/articles/PMC1230834/ [cited 2019 Nov 6]. Piantadosi A, Rubin DB, McQuillen DP, et al. Emerging cases of Powassan virus encephalitis in New England: Clinical presentation, imaging, and review of the literature. Clin Infect Dis. 2016;62(6):707-13. Available from: doi:10.1093/cid/civ1005. Centers for Disease Control and Prevention. Tick removal and testing [Internet]. 2019 Apr 22. Available from: http:// www.cdc.gov/lyme/removal/index.html [cited 2019 Nov 6].

POWASSAN ENCEPHALITIS

While Lyme disease is the most common tick-borne illness, Powassan encephalitis, transmitted by the Powassan virus, is the most severe. This virus was first discovered in 1958 when a 5-year-old boy from Powassan, Ontario was admitted to the Hospital for Sick Children. Three weeks prior, the child had fallen from a couch and suffered a swelling over his left eyebrow. He complained of dizziness, although his gait was regular. In the day prior to admission, his parents noticed tremor and unsteadiness of his left arm, as well as rhythmic movement of his eyes to the left. On admission, the child had a right-sided headache and was slightly drowsy. Tests returned normal results and an EEG showed a moderately diffusely abnormal pattern. However, two days later the patient became progressively drowsier and developed neck stiffness, eventually becoming unconscious. By the next morning, he was only responsive to painful stimuli. Four days post-admission to the hospital, the child was deeply comatose. An EEG this time showed a highly abnormal pattern indicative of encephalitis. That same day, the child suddenly stopped breathing, dying two days later.13 Since then, Powassan virus has been identified in five different tick species: Ixodes cookei, I. marxi, I. spinipalpus, I. scapularis, and Dermacentor andersoni. Since 2017, Ixodes species have expanded their range extensively through central and eastern Canada; the disease’s incidence is expected to grow.1 Diagnosing Powassan encephalitis is difficult. Tick bites can be easily overlooked without careful examination, since Ixodes ticks are small and can bite areas that are hard to check, such as the scalp, groin, and armpits. Out of the 27 reported cases of Powassan viral infection from 1958 to 1998, only 26% of patients reported noticing tick bites.14 The disease also causes nonspecific symptoms ranging from headaches, nausea, vomiting, and lethargy, to cranial nerve palsies, aphasia, dysphasia, seizure, confusion, and coma.15 Results from CT, MRI, and laboratory tests —including blood and cerebrospinal fluid investigations— may be non-specific or negative. After

several weeks, serial serum specimens may be required to accurately identify the antibodies for Powassan virus infection. Even once diagnosed, supportive treatment is the mainstay choice of management. Currently, there is no specific medication available to treat Powassan encephalitis. Often, patients who recover may have already suffered permanent neurological damage. For these reasons, it is important to raise awareness and understanding of the disease so that it can be addressed more effectively.

PREVENTION

Prevention of tick bites is one of the most important defenses against tickborne diseases. Ticks can be found in grassy or wooded areas, as well as on animals, particularly deer, making camping, gardening, and hunting highrisk activities for tick exposure. Treating clothing or gear with 0.5% permethrin or using EPA-registered insect repellents can help ward off ticks. Finally, avoidance and removal of plant litter and woodpiles, as well as mowing tall grass and fencing off yards, can be useful in reducing tick exposure.11 During and after outdoor activities, it is important to frequently check for ticks and their bites to prevent tick-borne diseases. If a tick is found and removed within 24 hours of attachment, Lyme disease can typically be prevented. Finetipped forceps should be used with care to remove the tick as close to the skin as possible without pressing on the tick body, to prevent bacteria being squeezed out of the tick.16

CONCLUSION

Tick-borne diseases can be most effectively managed with adequate awareness and prevention. Strategies to prevent and treat tick bites are vital in reducing the incidence of both common tick-borne infections such as Lyme disease, as well as rare and disabling conditions such as Powassan encephalitis.

REVIEWED BY: DANIEL SHI Daniel Shi is a second-year medical student at Queen’s University. He graduated from the BHSc program at McMaster in 2018. Daniel’s research interests focus on clinical trial methodology. He has published a review in the use of the crossover design in evaluating infertility treatments under Dr. Stephen Walter. EDITED BY: SHADI SADEGHIAN & ANITA SHAH

critical review M E D U C AT O R | DECEMBER 2019

ARTIST JEFF ZHANG

CRITICAL REVIEW Pyruvate Kinase Deficiency and Developments In its Treatment

doi:10.35493/medu.36.18

KASHYAP PATEL 1

Bachelor of Health Sciences Class of 2020, McMaster University Correspondence: patelk69@mcmaster.ca 1

OMAIKE SIKDER 2

Bachelors of Science (Honours Biochemistry) Class of 2020, McMaster University Correspondence: sikdero@mcmaster.ca 2

NIKHIL NAIR HARIHARAN 2

Correspondence: hariharn@mcmaster.ca

ABSTRACT

INTRODUCTION

Pyruvate kinase (PK) deficiency results in impaired PK activity in erythrocytes, and is caused by mutations in the PKLR gene. It presents with hemolytic anemia, which occurs when erythrocytes are lysed at a faster rate than they are produced, reducing erythrocyte numbers.1 Due to hemolytic anemia, an individual with PK deficiency can experience lethargy, pallor and jaundice. Other complications of this condition can include splenomegaly, gallstones, and pulmonary hypertension.2,3

This review discusses the nature of PK deficiency, current treatments and mitapivat, a novel therapy that aims to address the symptoms of PK deficiency.

PATHOPHYSIOLOGY

PK is responsible for transferring a phosphate group from phosphoenolpyruvate (PEP) to ADP, ultimately yielding 2 pyruvates and 2 ATPs per glucose molecule.8 The enzyme is expressed by either the PKLR or PKM genes in a tissue-specific manner. The PKLR gene codes for the PK enzyme in the liver and red blood cells (R-enzyme), whereas PKM codes for PK in muscle tissues and during fetal development (M2-enzyme). The M2-PK enzyme eventually persists in white blood cells and platelets.3

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Although the true prevalence of PK deficiency remains

Initial work-up for hemolysis usually involves complete blood count, peripheral blood smear, and reticulocyte count, along with measurement of bilirubin, haptoglobin and a direct antiglobulin test. This helps to assess the severity of hemolytic anemia and aids in the elimination of other acquired and inherited disorders with similar presentations.2,3,7 The differential diagnosis of PK deficiency includes acquired hemolysis and congenital anemias.

M E D U C AT O R

As PK deficiency is rare, its true prevalence is currently unknown.4 In addition, current prevalence estimates of PK deficiency are skewed due to errors in diagnosis. Diagnostic accuracy is an important consideration when analyzing prevalence, as PK deficiency is only diagnosed when physicians purposefully evaluate erythrocyte assays. Furthermore, errors in diagnosis persist due to the failure to associate hemolytic anemia with a genetic disorder such as PK deficiency.5 Even with the presence of enzyme assays and genetic testing, the variability and complexity of clinical presentation often result in delayed PK deficiency diagnosis.

The diagnosis of PK deficiency requires confirmation of hemolytic anemia, accompanied by low PK activity and/or positive genetic testing for mutations in the PKLR gene. This usually involves genetic screening or complete blood count assays and peripheral blood smears. These diagnostic tests are performed in individuals with unexplained anemia or hemolysis, particularly if there is a family history of congenital hemolysis coupled with PK deficiency.

critiical review

Pyruvate kinase (PK) is an enzyme involved in ATP production in the glycolytic pathway. PK deficiency is a rare, autosomal recessive disorder that manifests with hemolytic anemia of variable severity, partly due to an insufficiency of ATP production in erythrocytes. Its exact prevalence is unknown due to lack of awareness and challenges in making the diagnosis of PK deficiency. Current treatments for PK deficiency include blood transfusion and splenectomy, which focus on temporarily alleviating anemia and reducing hemolysis, respectively. Due to the rarity of this condition and its variable severity, a systematic treatment protocol for novel therapies is difficult to compile and validate. Oral administration of mitapivat, an allosteric activator of erythrocyte PK, is a novel treatment. Despite its promising effects on the hemoglobin levels in PK deficient patients, mitapivat’s efficacy and safety profiles must be determined through randomized controlled trials.

elusive, it has been established that PK deficiency has a worldwide distribution, with greater occurrence in northern Europe and regions of China. As PK deficiency is an autosomal recessive disorder, it is equally prevalent in male and female populations. However, women between the ages of 21 and 30 have a higher than basal prevalence; this increase is attributed to pregnancy, which is hypothesized to worsen anemia in those with PK deficiency.5 PK deficiency is also more prevalent in areas or social groups with higher rates of consanguinity, which can occur in isolated populations where the “founder effect” significantly reduces genetic variance.5,6,7

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Tanaka KR, Valentine WN, Miwa S. Pyruvate kinase (PK) deficiency hereditary nonspherocytic hemolytic anemia. Blood. 1962;19(3):267–95. Available from: doi:10.1182/blood-2016-02-702365. Grace RF, Bianchi P, van Beers EJ, Eber SW, Glader B, Yaish HM, et al. Clinical spectrum of pyruvate kinase deficiency: Data from the Pyruvate Kinase Deficiency Natural History Study. Blood. 2018;131(20):2183–92. Available from: doi:10.1182/blood-2017-10-810796. Zanella A, Bianchi P. Red cell pyruvate kinase deficiency: From genetics to clinical manifestations. Baillieres Best Pract Res Clin Haematol. 2000;13(1):57-81. Available from: doi:10.1111/j.13652141.2005.05527.x. Beutler E, Gelbart T. Estimating the prevalence of pyruvate kinase deficiency from the gene frequency in the general white population. Blood. 2000;95(11):3585– 8. Available from: doi:10.1182/blood. v95.11.3585.011k39_3585_3588. Hirono A, Forman L, Beutler E. Enzymatic diagnosis in non-spherocytic hemolytic anemia. Medicine. 1988;67(2):110–7. Available from: doi:10.1097/00005792198803000-00004. de Medicis E, Ross P, Friedman R, Hume H, Marceau D, Milot M, et al. Hereditary nonspherocytic hemolytic anemia due to pyruvate kinase deficiency: A prevalence study in Quebec (Canada). Hum Hered. 1992;42(3):179–83. Available from: doi:10.1159/000154063. Carey PJ, Chandler J, Hendrick A, Reid MM, Saunders PW, Tinegate H, et al. Prevalence of pyruvate kinase deficiency in northern European population in the north of England. Blood. 2000;96(12):4005– 6. Available from: https://www.ncbi.nlm. nih.gov/pubmed/11186276. [cited 2019 Nov 14]. Zanella A, Fermo E, Bianchi P, Valentini G. Red cell pyruvate kinase deficiency: Molecular and clinical aspects. Br J Haematol. 2005;130(1):11-25. Available from: doi:10.1111/j.13652141.2005.05527.x. Canu G, De Bonis M, Minucci A, Capoluongo E. Red blood cell PK deficiency: An update of PK-LR gene mutation database. Blood Cells Mol Dis. 2016;57:1009. Available from: doi:10.1016/j. bcmd.2015.12.009. Zanella A, Bianchi P, Baronciani L, Zappa M, Bredi E, Vercellati C, et al. Molecular characterization of PK-LR gene in pyruvate kinase-deficient Italian patients. Blood. 1997;89(10):3847-52. Available from: doi:10.1046/j.13652141.2001.02711.x. Wang C, Chiarelli LR, Bianchi P, Abraham DJ, Galizzi A, Mattevi A, et al. Human erythrocyte pyruvate kinase: Characterization of the recombinant enzyme and a mutant form (R510Q) causing nonspherocytic hemolytic anemia. Blood. 2001;98(10):3113-20. Available from: doi:10.1182/blood.v98.10.3113. Kung C, Hixon J, Kosinski PA, Cianchetta G, Histen G, Chen Y, et al. AG-348 enhances pyruvate kinase activity in red blood cells from patients with pyruvate kinase deficiency. Blood. 2017;130(11):134756. Available from: doi:10.1182/ blood-2016-11-753525. Oski FA, Marshall BE, Cohen PJ, Sugerman HJ, Miller LD. The role of the left-shifted or right-shifted oxygen-hemoglobin equilibrium curve. Ann Intern Med. 1971;74(1):44-6. Available from: doi:10.7326/0003-4819-74-1-44. Grace RF, Layton DM, Barcellini W. How we manage patients with pyruvate kinase deficiency. Br J Haematol. 2019;184(5):721–34. Available from: doi:10.1111/bjh.15758. Grace RF, Rose C, Layton DM, Galactéros F, Barcellini W, Morton DH, et al. Safety and efficacy of mitapivat in pyruvate kinase deficiency. N Engl J Med. 2019;381(10):933–44. Available from: doi:10.1056/NEJMoa1902678. National Institutes of Health. Extension Study of AG-348 in Adult Participants With Pyruvate Kinase Deficiency Previously Enrolled in AG-348-006 or AG348-C-007 [Internet]. Available from: https://clinicaltrials.gov/ct2/show/ NCT03853798 [cited 2019 Nov 14].

PK deficiency is an autosomal recessive disorder, meaning a child must inherit one mutated allele from each parent to display clinical features. While there are patients with homozygous mutations, most patients show compound heterozygosity, where each inherited mutation is different from the other. Clinical PK deficiency can be caused by over 250 mutations on the PKLR gene.3,9 These include missense, nonsense, insertion, and deletion mutations. Many of these mutations are geographically specific, such as the R486W and R510Q mutations in the PKLR gene, which are highly prevalent in Southern and Northern European populations, respectively.10,11 R510Q, R486W, and other mutations on the PKLR gene may result in impaired formation of the enzyme’s tetrameric structure and stability. Furthermore, these deleterious mutations can alter PK’s affinity for PEP and allosteric modulators such as fructose 1,6-bisphosphate, resulting in PK deficiency.11,12 Impaired PK function leads to a decrease in glycolytic ATP production, as well as an accumulation of glycolytic intermediates. As erythrocytes primarily utilize ATP created by glycolysis, a decrease in the available energy is a contributing factor to hemolysis and the removal of erythrocytes from circulation.3 The effects of hemolytic anemia are partially offset by an accumulation of 2,3-bisphosphoglycerate (2,3-BPG), which binds to hemoglobin to improve oxygen delivery to tissues. 2,3BPG levels are increased in patients with PK deficiency due to an accumulation of glycolytic intermediates, allowing for patients to better tolerate anemia-related symptoms.13

CURRENT TREATMENTS

Since PK deficiency is rare and prognosis varies greatly between individuals, there are no evidence-based management guidelines. Depending on the symptoms and severity presented by patients, healthcare providers can prescribe blood transfusions, surgical procedures, or investigational therapies including gene therapy and hematopoietic cell transplants. The central feature of PK deficiency is hemolytic anemia.2 A common approach to alleviate the symptoms of hemolytic anemia and maintain hemoglobin levels is blood transfusion, the frequency of which depends on the severity of the symptoms. Children with severe PK deficiency may undergo frequent blood transfusions until

splenectomy, which is typically deferred until the age of five when they can receive full immunization against Streptococcus pneumoniae. In a study with 250 PK deficient patients who underwent transfusions, 48% of patients aged five or younger received regular transfusions. In comparison, only 26% of patients between ages 5 to 12 received regular transfusions.2 According to a medical record review, the most common reasons for splenectomy in PK deficiency were: to improve baseline anemia, decrease the burden of transfusions, and reduce jaundice.2 A review by Grace et al. suggests that PK deficiency treatment plans are often individualized on a case-by-case basis.14

FUTURE DIRECTIONS

Mitapivat is an orally administered allosteric activator of erythrocyte PK.15 A recent phase II multicenter study evaluated the efficacy and safety of mitapivat in 52 PK deficient patients who were not undergoing transfusions. This study randomly assigned patients to daily mitapivat doses of either 50mg or 300mg for a 24-week period.15 The results indicated that mild adverse events such as headaches (92%) and insomnia (47%) occurred at drug initiation and were resolved within a week.15 In the 29-month follow-up period, 16 out of the 52 patients experienced a grade 3 adverse event, while 3 patients experienced a grade 4 event. Furthermore, pharyngitis and hemolytic anemia occurred in only two patients. The efficacy of this intervention was evaluated through monitoring changes in hemoglobin levels. 50% of patients had an increase of over 1.0 g/dL in their hemoglobin levels.15 Interestingly, this hemoglobin response was only found in patients who had at least one missense mutation. The 10 patients who had two non-missense mutations and 5 patients who were homozygous for R479H mutations did not have this response. This indicates that the response to mitapivat might depend on which mutations are causing the disease. Even though the mean maximum increase in hemoglobin was 3.4 g/dL, the study could not demonstrate a definitive relationship between mitapivat and hemoglobin levels.15 Although these results are worth consideration, mitapivat does have side effects; Grace et al. did not depict the severity of the adverse outcomes. In the 29-month follow-up period, some patients had adverse

outcomes including osteoporosis, ovarian cyst, renal-cell carcinoma, gastroenteritis, hernia, and thrombosis.15 While it is unclear which of these adverse events are a result of mitapivat as oppose to unrelated medical concerns, the effects are not transient and further study is required to evaluate the safety profile of the drug. Moreover, this uncontrolled phase II study does not provide sufficient quality of evidence to guide clinical treatment at this time. It is imperative that more rigorous studies, such as randomized controlled trials, be conducted to gain a better understanding of the efficacy and safety of mitapivat, as well as which patients are most likely to benefit from the treatment. There is an ongoing, multicentre, open label study that is evaluating the long-term efficacy and safety of mitapivat. The primary outcomes of this study are adverse events while secondary outcomes include the change in concentrations of various relevant biomarkers (hemoglobin, bilirubin and erythrocyte count, etc.).16

CONCLUSION

PK deficiency is a rare autosomal recessive disease that causes lifelong hemolytic anemia. The impaired functioning of PK in erythrocytes leads to a decrease in ATP production,

contributing to hemolytic anemia. Current treatment options are limited to blood transfusion and splenectomy. Recent research suggests that mitapivat, an allosteric activator of PK, holds promise in improving anemia in some patients with PK deficiency. However, both its efficacy and safety require further study. Future research should focus on the safety of this drug and its efficacy for patients with different mutations in the PKLR gene. An example is the aforementioned extension study, which aims to evaluate adverse events of mitapivat administration, that is currently in the recruitment stage.16 EDITED BY: PARNIKA GODKHINDI & NICK TELLER

REVIEWED BY: DR. MADELINE VERHOVSEK Dr. Madeleine Verhovsek has been an assistant professor in the department of Medicine at McMaster University since October 2010. She is the director of the adult hemoglobinopathy program at the McMaster Hemoglobinopathy Clinic and works as a Consultant Hematologist at St. Josephâ&#x20AC;&#x2122;s Healthcare Hamilton. Her research interests center on sickle-cell disease, red cell disorders and thalessemia.

critical review M E D U C AT O R | DECEMBER 2019

ARTIST ESRA RAKAB

RESEARCH INSIGHT

Developing a qualitative chemotaxis-sensitive assay for bacteriophage-host interactions on E. coli BW2511

doi:10.35493/medu.36.22

AARON WEN

Bachelor of Health Sciences (Honours), Class of 2022, McMaster University Correspondence: wena@mcmaster.ca

ABSTRACT To better understand the interactions between bacteriophages and host bacteria, one factor to observe is chemotaxis. Chemotaxis describes the biased directional movement of bacteria either towards or away from a substance. This phenomenon is created by the inhibition or potentiation of tumbling events, which randomize the direction of the host every few seconds. The presence of chemoattractants and chemorepellents can modify the number of tumbling events that occur. In order to elucidate the role of bacteriophages in bacterial chemotaxis, this project aimed to optimize an assay to qualitatively observe chemotaxis within E. coli BW2511 (WT) in comparison to a mutant strain with cheA deletion. Three main assay methods were tested: Tris HCl plate assays, Tris HCl microcapillary assays, and TTC Eppendorf assays. Of all the methods tested, only TTC Eppendorf assays with TTC indicator consistently showed chemotaxis. This study continued to use the Eppendorf assay and determined if various bacteriophages could attract E. coli BW2511 to chemotactically move up a layer of 0.4% PBS-based agar and against gravitational pull.

PREMISE In learning more about how phages interact with bacteria, this project developed an assay to clearly differentiate between motility (i.e. random, independent movement) and chemotaxis in E. coli BW2511. The development and optimization of this assay would later be used to better understand whether phages can alter bacterial chemotaxis to improve viral fitness. It was hypothesized that the lysis of a bacterium due to phage infection releases host amino acids and nutrients, which attract other bacterial hosts within the environment. This provides the phage with more opportunities for infection.

Within E. coli and other microorganisms, the chemotaxis pathway starts with the binding

In order to determine if bacteriophages play any role in the chemotactic behaviour of bacteria, an accurate and reliable assay had to be developed to differentiate chemotaxis from random motility. Three main assay types were explored: Tris HCl plate assays, Tris HCl capillary assays, and triphenyl tetrazolium chloride (TTC) - based Eppendorf assays.

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Chemotaxis can be explained as a series of simple switching events between two modes of cellular movement: moving and tumbling.3 Movement by the bacteria is encouraged by the long flagella, which whips around to create a propulsion motor.3 In the absence of any chemical gradient, a swimming bacterium moves erratically due to the switching process of tumbling, where the bacterium briefly stops and reorients itself in any random direction before moving again.3 If the cell is swimming towards an increasing concentration of an attractant, it will suppress the proteins responsible for tumbling, subsequently decreasing the probability of a tumbling event occurring.3 Alternatively, if a concentration gradient of a repellent is sensed to be near, the bacterium will increase its probability of tumbling as a form of escape.4 The net effect of this temporal sensing and regulation is the foundation for the biased movement known as chemotaxis.3

M E D U C AT O R

of ligands (e.g. serine, aspartate, or citrate) to transmembrane receptors.5 Depending on the type of ligand bound (i.e. chemoattractant or chemorepellent), tumbling will be either encouraged or inhibited.5 Ligand docking generates an intramolecular conformational change to the receptor that regulates a bound cytoplasmic histidine kinase, also known as chemotaxis protein CheA (CheA).5 Ligand binding causes CheA to be either upregulated or downregulated, allowing it to phosphorylate itself on a side chain and initiate a signalling cascade that generates the bacterium tumbling state. If an attractant is bound, CheA phosphorylation is downregulated to limit the sudden randomization of movement; the opposite occurs for the binding of a repellent.6

research insight

BACKGROUND With alarming reports of antimicrobial resistance and few new antibiotics on the market, there has been an increased interest in bacteriophage research over the past decade.1 Bacteriophages, also known as phages, are viruses that can infect and destroy specific bacterial hosts, giving them potential as anti-bacterial interventions.2 The biased movement of a bacterium towards a certain target is known as chemotaxis.3 Experiments and techniques previously used to investigate chemotaxis have been unconvincing, with little evidence to suggest unidirectional movement. As a result, there is a need to optimize a known chemotactic assay to distinctly differentiate chemotaxis from random motility of bacteria.

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research insight

METHODS AND RESULTS For every assay type, the bacterial strain E. coli BW2511 (WT) was used along with its nonchemotactic CheA knockout mutant (∆CheA). In each assay, bacteria were added with either a chemoattractant or chemorepellent, and directional movement was qualitatively observed.

FIGURE 1: Schematic of procedure for layered agar chemotaxis assay. An artificial serine gradient is made (as an example) by allowing the 1% agar to cool at an angle prior to adding the 0.35% agar overlay.

was not appropriate for chemotaxis detection. In parallel with plate assay experimentation, capillary assays (Tris HCl microcapillary assays) were also attempted. Previously, the capillary assay has shown increased attraction of WT to glucose than ∆CheA with this design. The principle behind this assay was to have WT or ∆CheA move up a glass capillary filled with either a chemoattractant, chemorepellent, or Tris HCl (negative control). Using a glass capillary would create a concentration gradient the moment that the capillary came in contact with the liquid within a 96 well plate. Chemotaxis was measured by the concentration of bacteria within the capillary after 30 minutes, which was quantified via a serial dilution in Tris HCl. Serial dilutions of bacteria were pipetted onto 1% standard agar plates and incubated overnight at 37oC. Colony forming units (CFUs) were counted the next day and plotted to create a violin graph (Figure 2). The thickness of each “violin” (purple or gold) represents the degree of consensus of data at that CFU/mL. The thicker the graph, the more data from the capillary experiments was pooled at that CFU/mL value. Although capillary assays were done with comparisons of serine, leucine, and a negative control, the majority of experiments were done with high concentrations of serine to hopefully induce a greater measure of chemotaxis. Ultimately, the majority of results did not reflect any chemotactic behaviour. None of the E. coli BW2511 strains exhibited any directed motility in response to either chemoattractants or repellants. Other changes such as time of incubation and type of chemoattractant also showed inconsistent results. Similar to plate assays, capillary assays were concluded to be an unfeasible methodology for our purposes.

For plate assays, E. coli BW2511 was inoculated in 10 mL of Lysogeny Broth (LB) overnight and then pipetted onto 0.4% 10 mM Tris HCl agar plates (pH 6.8) to stop bacterial growth but allow for chemotaxis. Then, a concentration of either 10 mM, 100 mM, or 1 M of chemoattractant (glucose), repellent (leucine), or nothing (as a negative control) was added at a distance of 1.5 cm away. Plates were left overnight at 37oC and observed for chemotactic movement. Initial results suggested that chemotaxis was not detected, as the diameters of the bacterial circles illustrated between WT and ∆CheA against all conditions were nearly identical. Adjustments made to the concentration, density of agar, and distance of placement all showed no difference in bacterial movement. Glucose was then swapped with serine, which has been demonstrated to be a definitive chemoattractant.7 It was also suspected that the ineffectiveness of the plate assays was due to the glucose and leucine being too diffuse through the 0.35% agar, causing the concentration gradient to be too weak for chemotaxis. To address this, two layers of 10 mM Tris HCl agar were used, one at a density of 1% and the other at 0.35%. Plates were poured over a period of two days per experiment, with either serine, leucine, or negative control mixed into the 1% agar prior to pouring at a slant. This design yielded no novel findings or affirmations to bacterial chemotaxis. The 1% agar layer, when poured at an angle, generated After failures in both plate and capillary an artificial concentration gradient for assays, triphenyl tetrazolium chloride bacteria to swim towards after adding a 0.35% (TTC) assays were tested to determine if agar overlay (Figure 1). Another set of plates bacterial chemotaxis could be observed up were also built with dual 1% agar gradients of an Eppendorf tube. This assay was pioneered serine and leucine and a 0.4% agar overlay, but in chemotactic research for the bacterium C. they were also ineffective. These plates were jejuni.8 TTC is a compound that develops a the final iteration of the plate assay, as these pink stain in the presence of mitochondrial results suggested that the entire assay design enzymes from the electron-transport chain,

FIGURE 2: Violin graph of capillary assay data, with red dots representing mean values of CFU/mL. It can be seen that the capillary assay has insufficient resolution to distinguish between motility and chemotaxis.

CONCLUSION With more attention being put toward understanding the behaviour and potential of bacteriophages in medicine, there is a rising demand to generate appropriate tools and methodologies to elucidate their complex mechanisms. Here, an assay to detect chemotaxis in E. coli is proven to be accurate and consistent, opening more possibilities to investigate bacteriophage-host interactions.

Furfaro LL, Payne MS, Chang BJ. Bacteriophage therapy: Clinical trials and regulatory hurdles. Front Cell Infect Microbiol. 2018;8:376. Available from: doi:10.3389/fcimb.2018.00376. Vandamme EJ, Mortelmans K. A century of bacteriophage research and applications: Impacts on biotechnology, health, ecology and the economy! J Chem Technol Biotechnol. 2018;94(2):32-42. Available from: doi:10.1002/jctb.5810. Webre DJ, Wolanin PM, Stock JB. Bacterial chemotaxis. Curr Biol. 2003;13(2):R47-9. Available from: doi:10.1016/S0960-9822(02)014240. Barrangou R, Fremaux C, Deveau H, Richards M, Boyaval P, Moineau S, et al. CRISPR provides acquired resistance against viruses in prokaryotes. Science. 2007;315(5819):1709-12. Available from: doi:10.1126/science.1138140. Falke JJ, Bass RB, Butler SL, Chervitz SA, Danielson MA. The two-component signaling pathway of bacterial chemotaxis: A molecular view of signal transduction by receptors, kinases, and adaptation enzymes. Annu Rev Cell Dev Biol. 1997;13(1):457–512. Available from: doi:10.1146/annurev.cellbio.13.1.457. Wang X, Vu A, Lee K, Dahlquist FW. CheA–receptor interaction sites in bacterial chemotaxis. J Mol Biol. 2012;422(2):282-90. Available from: doi:10.1016/j.jmb.2012.05.023. Kalinin Y, Neumann S, Sourjik V, Wu M. Responses of Escherichia coli bacteria to two opposing chemoattractant gradients depend on the chemoreceptor ratio. J Bacteriol. 2010;192(7):1796-800. Available from: doi:10.1128/JB.0150709. Dwivedi R, Nothaft H, Garber J, Kin LX, Stahl M, Flint A, et al. L-fucose influences chemotaxis and biofilm formation in Campylobacter jejuni. Mol Microbiol. 2016;101(4):575-89. Available from: doi:10.1111/mmi.13409. Towill LE, Mazur P. Studies on the reduction of 2,3,5-triphenyltetrazolium chloride as a viability assay for plant tissue cultures. Can J Bot. 1975;53(11):1097-102. Available from: doi:10.1139/b75-129.

research insight

and is often used as an indicator for bacteria within an environment.9 WT and ∆cheA mutant strains were inoculated in 10 mL LB and grown overnight at 37oC. 1.5 mL of bacteria was transferred into a 2 mL Eppendorf tube and spun down at 1200 RPM to pelletize. The supernatant was decanted and the pellet was resuspended in PBS-based 0.4% agar, a variation on Tris HCl agar to limit growth but not motility. At the very top of the Eppendorf tube, a Whatman filter paper soaked in 50 μL of serine or leucine (between 10 mM and 1 M) was placed to generate a concentration gradient for bacterial chemotaxis. These Eppendorfs were incubated for two days at 37oC, with 0.01% TTC added after to develop colour.

When experimental lines were compared to multiple negative controls (no bacteria and no chemotactic substance), it was clear that the TTC assay qualitatively differentiated randomized motility from chemotaxis. In WT, the addition of serine led to purple colors near the top of the PBS-based 0.4% agar layer, and increasing concentrations of serine correlated with a darker shade of purple in the Eppendorf tube. Furthermore, trials with either the leucine repellent (10 mM to 0.14 M) or negative control were either entirely clear or purple at the bottom of the Eppendorf tube, suggesting that the bacteria demonstrated no upward directional movement. These results affirmed the functionality of the TTC assay to differentiate random motility and directional chemotaxis, giving it the potential to elucidate more details on the interactions of virulent bacteriophages on chemotaxis. However, a key limitation is that the degree to which chemotaxis has occured can be difficult to identify qualitatively, and more quantitative assay methods should be investigated.

FIGURE 3: Initial results from 0.01% TTC assay on E. coli BW2511 (WT) with Whatman filter paper soaked in (from left to right) nothing, 50 mL of 0.14 M leucine, and 50 mL of 1 M serine.

M E D U C AT O R

EDITED BY AARON WEN & JERRY DU

Felix Croteau is a PhD candidate in the Department of Biochemistry and Biomedical Sciences at McMaster University, where he has been studying the role of bacteriophages in model bacterial communities since May 2018. He has also studied the CRISPR-Cas adaptive immune system of bacteria in Dr. Sylvain Moineau’s Lab at the Université Laval. He is a recipient of the Canadian Society of Microbiologists Undergraduate Award.

| DECEMBER 2019

REVIEWED BY: FELIX CROTEAU

IN T E R V IE W SPOTLIGHT

DR. GORDON G U YAT T

E V ID E N C E - B A S E D ME D I C INE DANIEL RAYNER1 & HARGUN KAUR2

Bachelor of Health Sciences, Class of 2022, McMaster University Bachelor of Health Sciences, Class of 2023, McMaster University

Dr. Gordon Guyatt is a Distinguished Professor in the Department of Health Research Methods, Evidence, and Impact at McMaster University, and is one of the founders of “evidence-based medicine.” He has played a significant role in over 30 major clinical studies, including large-scale observational and randomized trials and has extensive expertise in study methodology. As the co-founder and co-chair of the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) working group, he has been intimately involved in the development and evolution of the GRADE approach for evaluating research evidence.

AS ONE OF THE FOREFATHERS OF EVIDENCE-BASED MEDICINE, HOW AND WHY DID YOU GET STARTED IN THE FIELD? I was a resident in Internal Medicine at McMaster University and I was very fortunate because [Dr.] David Sackett and other senior people in the department, people like [Dr.] Peter Tugwell and [Dr.] Brian Haynes, were already here and had started a new department— the first in the world—of clinical epidemiology. They were starting to teach people, clinicians, to use the medical literature to optimize their patient care. They called it “critical appraisal” at first, and it started as kind of a classroom activity. You went to a classroom, away from your clinical care, and learned about the principles of understanding and using medical literature. Then, David Sackett came up with the concept of bringing “critical appraisal” to the bedside, which was to take it right where you are delivering care and using the literature in the very process of it. In 1990, I took over the Internal Medicine residency program [as its] director, and my notion was to create the world’s first program that would implement this idea of bringing understanding of the evidence to the direct care of the patients. I needed a name for what I was going to call this, because I wanted to say, “Okay, if you are interested

in doing Internal Medicine residency and you want to come to McMaster, come and you will do ‘something,’ and I needed a name for it. My first name [for it] was ‘scientific medicine’ which was rejected with tremendous hostility by my colleagues in the Department of Internal Medicine. And so, “evidence-based medicine” was my second idea for what to call this, and it turned out to be a very catchy name. THERE HAS BEEN CRITICISM ABOUT EVIDENCEBASED MEDICINE REGARDING INFORMATION OVERLOAD AND VESTED INTERESTS OF PHARMACEUTICAL COMPANIES. WHAT ARE YOUR THOUGHTS ON THIS AND HOW DO YOU THINK THESE PROBLEMS CAN BE BEST COMBATED?

In terms of information overload, few clinicians are going to have both the skills and the time to actually critically read the original literature and use it in their patient care. So what they are going to have to understand [is how to use available results to determine] the benefits and harms of [the] interventions they are thinking of using. But, they need a summary of the information from somebody else and so fortunately, now we have many institutions providing clinical practice guidelines [and] we have many systematic reviews —these are two ways that clinicians can get good evidence summaries. There is another way, and I need to state my conflict of interest here because, for a number of years, I have done a lot of consulting/education work with a world-leading electronic medical textbook, Up -To -Date. I’m biased because my job is to get them to be as evidence-based as possible, so after working with them for a decade, I’d better believe we’ve made some progress. [This] is another source of good pre-processed information that clinicians can use that ideally will direct them to the best evidence and then evidence summaries that allow them to understand the benefits and harms that the studies portray.

What we did was to look at all the evidence on the health effects of red meat. There were four systematic reviews that we published. One focused on observational, non-randomized studies of people who ate more or less red meat and its effects on [developing] cancer and cardiovascular disease, and looked at dietary patterns that included more and less red meat. As well, another review [of ours] looked at all the trials where people were randomized to diets that had more red meat or less red meat. The results of these were very clear, and were quite consistent with prior reviews, although prior reviews did not present the data optimally, which I will explain in a second. So what we found was that the studies do show an association between red meat and cancer, and red meat and cardiovascular disease, but the association is... very weak. To give you an example, we’re convinced appropriately that smoking causes cancer, because in heavy smokers, it increases your risk of cancer ten-fold. Here, the magnitude of association was increases of less than 20%, so very small associations in relative terms. [There are] biases that are possible in observational studies, because people who eat red meat are different than people who don’t eat red meat in ways other than [their red meat consumption]. So the diets that contain red meat are different from the diets that don’t in other ways. People might smoke more, or [have] different socioeconomic statuses, or be exposed to toxins —[there are] lots of differences. These observational studies generally lead to lowquality evidence, unless there are very strong effects, such as in smoking, and that was not there at all [in our study]. Moreover, the most convincing, randomized, large-scale trial available that looked at people who ate more or less red meat did not show any differences in cancer or cardiovascular disease. Now this study has limitations too, probably [involving] low-quality evidence. Bottom line, we only have low-quality evidence [to work with]. Even if we accept that there is a causal relation —which is not at all certain —but even if we accept that the effect is very small for individuals, you would have to cut your meat consumption by three servings a day for the rest of your life to cut your cancer mortality risk, best estimate, by 7 in 1000. Most people would say that is a very small effect. So the bottom lines are, [there is] only low-quality evidence and even if there is an effect —which there may not be —the effect is very small, and the magnitude of gain you would have in cutting your red meat is [also] very small.

| DECEMBER 2019

I think GRADE has been a big step forward for [both] clinical epidemiology and evidence-based medicine. It is crucial to be able to distinguish evidence you can trust from evidence that is untrustworthy, though that’s probably too strict of a dichotomy. There’s evidence that you might call “high-quality,” “moderatequality,” “low-[quality],” and “very low-[quality],” the “very low” being very untrustworthy. And if we are going to get it right, we

YOU RECENTLY PUBLISHED A STUDY INVESTIGATING THE CONSUMPTION OF RED MEAT THAT HAS BEEN SOMEWHAT CONTROVERSIAL. DO YOU MIND ELABORATING A BIT ON THE STUDY’S FINDINGS AND WHY YOU THINK IT HAS RECEIVED THE CRITICISMS THAT IT HAS?

M E D U C AT O R

HOW CAN YOU ENCOURAGE PEOPLE TO ADOPT THE GRADE SYSTEM AND WOULD THAT HELP COMBAT THE AFOREMENTIONED PROBLEMS WITH EVIDENCEBASED MEDICINE?

The GRADE working group started to meet in 2000, [and] put out its first guidance in 2004. It has put out many papers since then, clarifying and deepening the guidance, and it is now a system that is used by over 110 organizations worldwide, including “Up-ToDate.” It provides a uniform, transparent, and standard way of deciding what is more trustworthy and what is less trustworthy evidence, which is a big step forward.

interview spotlight

With respect to the pharmaceutical industry, what we need to do is to train clinicians and the various sorts of experts who [make] guidelines for clinicians to be aware of the spin that is put on pharmaceutical industry studies. The pharmaceutical industry does its studies very well in terms of concealing randomization, blinding everybody possible, managing complete follow-up, and ensuring accuracy of the data. So the methods and the data that come out of [these] studies [are] generally very high-quality. The problem is how they interpret and use the data and that problem is solved by having [a third-party interpret it] for clinicians to be able to understand the appropriate analysis of that data.

need to know whether our evidence is “high,” “moderate,” “low,” or “very low” quality and we need a system that is well-developed, carefully thought out, transparent, and has rules that allow people to apply the system effectively, and GRADE has done all of that.

table of contents interview spotlight

We then did another systematic review, which looked at people’s feelings about their meat [consumption], what we call their values and preferences. Perhaps not surprising, we found that people who eat meat have lots of reasons for doing it that they think are good, and that they might not be at all keen to reduce their red meat consumption substantially for very uncertain and small benefits. So, our guideline panel said, “Okay, so we now want to make a recommendation,” and the recommendation is based on this: what do we think that most people [would do], if they understood the evidence and they were red meat eaters? Would they choose to reduce, or would they not? And what our panel believed is that the majority of people who were informed of the evidence would say, “...very small effects and uncertain? Thanks for the information, [but] I’ll keep eating my meat.” And so, we made what is a weak recommendation that people continue their current red meat consumption. Why a weak recommendation? A weak recommendation in the GRADE system that we used means that the majority of fully informed people would choose the panel’s recommended action if the panel had gotten it right but a minority would not. We thought that although the majority of fully informed people would choose to continue their red meat consumption, a minority would not. They would say, “Okay, it may be a very small effect, and uncertain, but my priority is my health and I’m going to cut down on my red meat.” Why did people get upset at [our recommendation]? People got upset at it because previous recommendations had all been “cut down on your red meat!” Frankly, if somebody says “Hey, wait a minute, let’s look at the evidence more carefully to see [if] that’s really legitimate,” it’s threatening to people who have come out and said that everybody should cut down on their red meat.

M E D U C AT O R

| DECEMBER 2019

HOW HAS EVIDENCE-BASED MEDICINE CHANGED OVER THE YEARS? WHERE DO YOU SEE EVIDENCE-BASED MEDICINE HEADING IN THE NEXT FEW DECADES?

When we started, we were thinking in a way that turned out to be very unrealistic —that every physician would have the time and skills to read original papers, and the methods and results of those papers. That turns out to be a completely unrealistic idea. We still think there are certain things worth educating physicians about in reading papers and particularly understanding the results. They need to understand what the benefits and downsides of interventions are, for instance. They need to understand the uncertainty of the evidence, because [it could be] low-quality evidence where they really don’t know, or high-quality evidence that you can be confident of. So clinicians have to be taught [how to assess evidence]. In my educational practice, I’ve shifted from assessing risk of bias in studies, which is [still] worth knowing about, [towards what you can understand from] the results: what are the magnitudes of the effects, what are the certainties, and on

what basis do people make decisions on the certainty of evidence. So that’s been one big change that’s happened. The other thing is, at the beginning, if you read [the guidelines] we wrote, values and preferences barely appeared at all. You know, “Oh what’s the evidence,” as if, naively, the evidence told you what to do. But, as has happened in other parts of this conversation, most of the time, evidence doesn’t tell you what to do. The right thing to do is different for different people depending on their values and preferences. So there has been a major shift to acknowledge and build the issue of people’s values and preferences into everything we do. So that is the second major shift since the beginning. As to the future, what we need to do is to build on our understanding that many decisions are value- and preference-dependent. In other words, in the same circumstances, the values and preferences of different people will [cause them to] make different choices. And the challenge now is to make sure the choice is correct for every individual. How do we do that? We do that in what has been called “shared decision-making” which is working with the patient, ensuring they understand what the options are, the evidence behind it, whether it is low-or high-quality, and the best guess as to what the benefits and downsides are. That turns out to be very challenging. To do it well, I think we will need what’s called “point-of-aid decision aid,” electronic decision aids where the physician and the patient are looking at a computer or iPad or some sort of screen that tells them a summary of the benefits and downsides. To do shared decision-making most efficiently and effectively, we’re going to need a lot of such decision aids [that are] well-developed so that they are accurate, give the right information, do it simply [and] clearly, and are actually fun for both the patient and clinician. And that’s the challenge for us in the future.

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