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WELCOME

Dear CRINA Research Day Attendee:

Thank you for joining us at the fourth annual CRINA Research Day. Last year, at our third event we welcomed over 250 attendees and featured more than 100 posters from many departments and faculties across campus. We are happy to announce that many of those attendees signed up to be members of CRINA, forming the core of our cancer research community. One year later, we continue to build our cancer research community by hosting a cancer-themed Research Day yet again. This year, we have continued to provide trainees with an opportunity to organize the program and present their work orally to our cancer research community at the University of Alberta. We hope that you continue to explore what the University of Alberta has to offer in the cancer research sphere and grow your network of collaborators through future CRINA Research Days.

CRINA as an institute has a well-established reporting structure with operations committees and advisory boards. At our core, we continue to strengthen connections within our cancer research community by hosting events throughout the year such as seminars and symposia. Our leadership team is working on defining University of Alberta cancer research strengths in terms of research excellence and available infrastructure and platforms, with plans to build on these strengths to accelerate discovery and innovation. CRINA also represents the interests of its members as a unified voice on the provincial stage, working with AHS, AIHS and the ACF. Our ultimate goal is to establish our Institute as a national leader in cancer research and patient care, wherein clinical outcomes are addressed with scientific inquiry and where research drives innovations in cancer prevention, treatment and survivorship.

We hope you will enjoy today’s program and find time to strengthen or make new collaborations with the CRINA community.

Sincerely,

Dr. Lynne-Marie Postovit

Director, Basic Research, Cancer Research Institute of Northern Alberta

MESSAGE FROM THE DEAN

Cancers remain as the second leading cause of death globally, responsible for 8.8 million deaths in 2015. During the 2017 World Health Assembly, representatives from 40 States and 11 NGOs reached broad consensus that cancer is a growing public health concern in urgent need for attention and prioritizing. Almost half of the North American population will develop cancer in their lifetimes and 25 per cent will die from this disease. Furthermore, Canada’s new cancer case rates are projected to increase by more than 40 per cent by 2030.

In Alberta, new cancer cases will reach an appalling number of more than 18,000, and death rates are more than 6,500 per year. Lung cancer is the leading cause of cancer death in both Alberta men and women, and the most common cancers for males and females are prostate cancer and breast cancer, respectively. These increasing numbers can only be changed through partnerships between health systems and academia to bolster the research that will bring us closer to a cure.

In April 2014, the University of Alberta established the Cancer Research Institute of Northern Alberta (CRINA), as a hub for talented clinicians and cancer researchers from all affiliated teaching hospitals, departments and faculties at the University of Alberta. CRINA’s research programs, aligned with clinical trials provided at the Cross Cancer Institute and the University of Alberta Hospital, as well as pediatric cancer clinical trials based at the Stollery Children’s Hospital, are setting a new course for cancer centres across Canada.

We are committed to leading the fight against cancer by seeking the improvement of cancer diagnosis, primary and secondary cancer prevention, and developing novel treatment approaches through clinical trials. Through the work of CRINA and its members, we aim to make Alberta a global leader in cancer research, fostering an environment of innovation that translates into tangible solutions for patients everywhere.

On behalf of the Faculty of Medicine & Dentistry, we wish you a productive day to celebrate your achievements in the past year and share ideas that will inspire future research ventures.

Richard N. Fedorak, MD, FRCPC, FRCP (London), FRCP (Edinburgh), FRSC Dean (on leave), Faculty of Medicine & Dentistry University of Alberta

Dennis Kunimoto, MD, FRCPC Acting Dean, Faculty of Medicine & Dentistry University of Alberta

MESSAGE FROM THE VICE PRESIDENT (RESEARCH)

More than 80,000 Canadians will lose their battles with cancer this year and, of these, 6500 will be Albertans. To improve outcomes for patients and families with cancer, the University of Alberta created the Cancer Research Institute of Northern Alberta (CRINA), one of the multidisciplinary Translational Science Institutes (TSIs) dedicated to fostering collaboration between researchers to move the latest discoveries from the laboratory to the clinic. CRINA has had two successful research days with over 250 attendees each year to highlight their achievements. They have engaged world-class scientists in several ways, including holding a cancer-focused Gairdner Symposia in 2016 and the hosting of Sir Paul Nurse in 2016, a Nobel laureate for his discoveries on cell cycle control.

These and other events illustrate the exciting momentum that CRINA has initiated, and Research Day 2017 will continue to demonstrate its tremendous potential. CRINA will redefine the standard of cancer care in Alberta by improving our understanding of cancer biology, cancer imaging, development of new therapeutics and biomarkers to diagnose patients with greater accuracy that will improve relapse rates and aid in cancer prevention.

I congratulate the life sciences faculties at the University of Alberta on this important interdisciplinary initiative and I wish you continued success.

Walter T Dixon, PhD Interim Vice-President (Research) University of Alberta

CRINA RESEARCH DAY 2017 PROGRAM

7:30 AM REGISTRATION CCIS PCL Lounge

7:30 AM – 8:15 AM BREAKFAST CCIS PCL Lounge

8:15 AM – 8:40 AM WELCOME ADDRESS CCIS 1-430 Dr. Paula Robson, Scientific Director, Cancer Research, CancerControl Alberta and the Cancer Strategic Clinical Network

Dr. Declan Ali, Acting Associate Vice-President (Research), University of Alberta Dr. Dennis Kunimoto Acting Dean, Faculty of Medicine and Dentistry, University of Alberta Dr. Lynne Postovit, Co-Director CRINA Associate Professor, Oncology

8:40 AM – 9:10 AM Keynote Address, Dr. David Huntsman

Professor, Departments of Pathology & Laboratory Medicine and Obstetrics & Gynaecology, University of British Columbia Director, Ovarian Cancer Research Program (OVCARE) Medical Director, Centre for Translational and Applied Genomics, BC Cancer Agency

Title: Cell Context Mutation and the Origins of Ovarian Cancer

______

SESSION I: NOVEL THERAPEUTICS AND BIOMARKERS CCIS 1-430 Co-Chairs: Dr. Lena Bilyk (Post-Doctoral Fellow, Department of Oncology) and Dr. Ing Swie Goping (Department of Biochemistry) 9:10 AM – 9:25 AM Dr. Hanne Ostergaard, Department of Medical Microbiology and Immunology Title: CD8+ T Cell Mediated Killing of Cancer Cells

9:25 AM – 9:40 AM Dr. Roger Zemp, Department of Electrical & Computer Engineering Title: Towards Label-Free Virtual Histopathology

9:40 AM – 9:55 AM Dr. Ratmir Derda, Department of Chemistry Principal Investigator in Alberta Glycomics Centre

Title: Genetically Encoded Chemistry: Platform for Development of New Therapeutics and Bio Active Materials

9:55 AM – 10:10 AM Igor Paiva, PhD Student, Faculty of Pharmacy and Pharmaceutical Sciences

Title: Biodistribution of EGFR Targeted Polymeric Micelles in Orthotopic Colorectal Cancer Xenografts in Mice

10:10 AM – 10:30 AM CRINA 1st Annual Speed Poster Session CCIS 1-430 Co-Chairs: Dr. Mary Hitt (Department of Oncology) and Dr. Shairaz Baksh (Department of Pediatrics)

Poster #2, Dr. Guanmin Meng, Post-Doctoral Fellow, Department of Biochemistry Poster #7, Laura Lee, MD/PhD Student, Department of Oncology

Poster #10, Dr. Mohammad A.M. Ali, Post-Doctoral Fellow, Department of Oncology Poster #19, Mackenzie Coatham, PhD Student, Department of Obstetrics and Gynecology Poster #24, Bindu Thapa, PhD Student, Department of Pharmaceutical Sciences Poster #29, Dr. Kyle G. Potts, Post-Doctoral Fellow, Department of Medical Microbiology and Immunology Poster #30, Dr. Deepak Dinakaran, Resident, Department of Oncology

Poster #31, Francesco Gentile, PhD Student, Department of Physics Poster #37, Michael J. Taylor, MD Student, Department of Medicine Poster #40, Dr. Amy Kirkham, Post-Doctoral Fellow, Department of Biomedical Engineering ______

10:30 AM – 11:50 AM POSTER SESSION 1 CCIS PCL Lounge ODD NUMBERED POSTERS

11:50 PM – 12:30 PM LUNCH /NETWORKING CCIS PCL Lounge

SESSION 1I: PREVENTION, LIFESTYLE AND SURVIVORSHIP CCIS 1-430

Co-chairs: Sarah Purcell (PhD Student, Department of Agricultural, Food & Nutritional Science) And Dr. Amit Bhavsar (Department of Medical Microbiology and Immunology)

12:35 PM – 12:50 PM Dr. Carla Prado, Department of Agricultural, Food & Nutritional Science Director of the Human Nutrition Research Unit Title: Body Composition and Energy Metabolism: Applications to Cancer Care 12:50 PM – 1:05 PM Dr. Jennifer Vena, Research Lead, Alberta's Tomorrow Project CancerControl, Alberta Health Services

Title: Alberta’s Tomorrow Project: A Health Research Resource Available Now ______

SESSION 1II: CANCER BIOLOGY CCIS 1-430 Co-Chairs: Zelei Yang (PhD Student, Department of Biochemistry) and Dr. Ismail Ismail (Department of Oncology)

1:05 PM – 1:20 PM Dr. Shelagh Campbell, Department of Biological Sciences Title: Developmental Regulation of the Cell Cycle by Wee1 and Myt1 Kinases

1:20 PM – 1:35 PM Dr. Armin Gamper, Department of Oncology Title: When 1 + 1 Does not Equal 2: Conditional Synthetic Lethality as Strategy for Cancer Therapy

1:35 PM – 1:50 PM Dr. Gordon Chan, Department of Oncology Title: Prolonged Mitotic Arrest induced by Wee1 Inhibition Sensitizes Breast Cancer Cells to Paclitaxel

1:50 PM – 2:05 PM Fahed A Elian, PhD Student, Department of Medical Genetics Title: Molecular Analysis of FOXC1 in Triple Negative/Basal- Like Breast Cancer (TN/BL BC)

2:05 PM – 3:35 PM POSTER SESSION 2 CCIS PCL Lounge

EVEN NUMBERED POSTERS

SESSION IV: THE NEXT GENERATION CCIS 1-430 Co-Chairs: Brittany Umer (PhD student, Department of Medical Microbiology and Immunology and Dr. Michael Hendzel (Department of Oncology)

3:40 PM – 3:55 PM Dr. Gabrielle Siegers, Research Associate, Department of Oncology

Title: Functional Plasticity of Gamma Delta T cells and Breast Tumour Targets in Hypoxia

3:55 PM – 4:10 PM Jingjie Xiao, PhD Student, Department of Agricultural, Food & Nutritional Science

Title: Associations of Pre-existing Comorbidities with Skeletal Muscle Mass and Radiodensity in Early Stage Colorectal Cancer Patients

4:10 PM – 4:25 PM CLOSING REMARKS AND AWARDS CCIS 1-430

4:30 PM – 6:30 PM RECEPTION & CASH BAR CCIS PCL Lounge

SPEAKER ABSTRACTS

Pages 11-20

Dr. Roger Zemp Electrical & Computer Engineering

Towards Label-Free Virtual Histopathology Abstract: A new all-optical non-contact technology called photoacoutsic remote sensing (PARS) microscopy will be introduced for visualizing capillary-level angiogenesis and label- free histopathology. PARS is capable of visualizing tumor vasculature and blood oxygen saturation. The technology is under development for incorporation into a surgical microscope for intra-operative tumor margin assessment.

Igor Paiva, PhD Student Pharmacy and Pharmaceutical Sciences Biodistribution of EGFR Targeted Polymeric Micelles in Orthotopic Colorectal Cancer Xenografts in Mice Igor Paiva(1), Amir Soleimani(1), Sams Sadat(1), Zahra Shrine(2), Mohammad Vakili(1), Gary Martin(3), Frank Jirik(3), Michael Weinfeld(2), Afsaneh Lavasanifar(1) Igor Paiva(1), Amir Soleimani(1), Sams Sadat(1), Zahra Shrine(2), Mohammad Vakili(1), Gary Martin(3), Frank Jirik(3), Michael Weinfeld(2), Afsaneh Lavasanifar(1) Introduction: Nanocarrier surface modification using ligands interacting with receptors overexpressed on cancer cells is hypothesized to enhance the homing of nano-carrier in tumor. The objective of this study was to test this hypothesis in an orthotopic mice model of colorectal cancer (CC) for polymeric micelles modified on their surface with GE11 peptide that targets epidermal growth factor receptor (EGFR) on CC cells. Methods: Traceable GE11 modified poly(ethylene oxide)-poly(caprolactone) (PEO-PCL) or poly(ethylene oxide)- poly(abenzyl carboxylate-caprolactone) (PEO-PBCL) were developed through peptide attachment to the acetal-PEO and Cy5.5 to PCL or PBCL end (Garg et al., 2017). The uptake of

polymeric micelles by HCT-116 cells overexpressing EGFR was measured using flow cytometry. An orthotopic colorectal mice model was developed and used to assess the biodistribution of plain versus GE11 modified micelles following intravenous administration using live animal or ex vivo organ imaging by an IVIS imager. Results: The results showed micelles with GE11 modification to have a higher in vitro uptake than plain micelles in HCT- 116 cells. The cell uptake enhancement seemed to be more prominent for GE11-PEO-PBCL micelles than for GE11-PEO-PCL micelles. The fluorescence intensity in the tumor site from the mice was higher for micelles having PCL core at early time points, whereas micelles with PBCL core accumulated more into the tumor site at later time points (i.e. ~24h). There was a trend both in vivo and ex vivo for a higher micelle tumor accumulation when GE11 peptide was present on micellar surface. The results also showed faster clearance of PEO-PCL based micelles compared to PEO-PBCL ones mostly through mice kidneys leading to lower accumulation of the former nano-carriers in non-target tissues as well as tumor. Conclusion: Surface decoration of polymeric micelles with the GE11 peptides positively impacted their in vitro CC cell uptake and in vivo accumulation into orthotopic CC tumors.

Dr. Carla Prado Agricultural, Food & Nutritional Science Body Composition and Energy Metabolism: Applications to Cancer Care Abstract: Abnormal body composition in cancer is an independent predictor of poor outcomes and occurs across cancer types and stages. This presentation will highlight the diagnosis of abnormal body composition and energy metabolism using simple and sophisticated tools. We will additionally discuss potential approaches to countermeasure abnormal body composition with particular focus on targeted nutrition interventions.

Dr. Jennifer Vena Alberta's Tomorrow Project CancerControl, Alberta Health Services Alberta’s Tomorrow Project: A Health Research Resource Available Now Alberta’s Tomorrow Project (ATP) is a longitudinal prospective cohort study. The largest of its kind in the province, ATP was established to facilitate research into the causes and prevention of cancer and chronic disease. Between 2000 and 2015, 55,000 adults (age 35- 69y) were enrolled and completed baseline health and lifestyle questionnaires. As a result of the lengthy recruitment period, ATP has amassed both cross-sectional and growing longitudinal data: some late-entry participants finished only a baseline questionnaire, while others who joined earlier have completed up to 4 follow-up surveys. Additional data collection is ongoing, creating an ever-growing and robust dataset capable of supporting a variety of health research investigations. ATP also conducts twice-yearly linkages with the gold-certified Alberta Cancer Registry; the most recent linkage found that more than 3300 ATP participants have been diagnosed with some type of cancer (other than non-melanoma skin cancer). This talk will provide an overview of ATP’s vast dataset, examples of research projects currently underway using data and biosamples, incidence and prevalence of cancer and chronic diseases in the cohort, and exciting areas of dataset enrichment on the horizon.

Dr. Shelagh Campbell Biological Sciences Developmental Regulation of the Cell Cycle by Wee1 and Myt1 Kinases Precisely timed progression through the cell cycle is essential for producing appropriate numbers of properly specified cell types during animal development. Many different types of cell cycles exist, serving specialized developmental functions. For example, many early embryos undergo simplified cleavage cycles consisting of rapid oscillations between S and M phases of the cycle by dispensing of gap phases. In spite of this diversity, regulation of Cdk1 activity is a universal mechanism that coordinates mitosis with other stages of the cell cycle in all eukaryotic cells. Defects in Cdk1 regulation are frequently associated with cancer and so new therapies targeting these conserved Cdk1 mechanisms are being actively sought. During interphase, Cdk1 activity is inhibited by two related kinases: Wee1 and Myt1. In my laboratory we are studying the developmental functions of these partially redundant inhibitory kinases in Drosophila. For this purpose we have created null wee1 and myt1 mutants and found unique developmental functions for each of these Cdk1 inhibitors. Drosophila Wee1 mutants are early embryonic lethal due to specific S/M checkpoint defects that result in mitotic catastrophe. Wee1 inhibitors being developed for cancer therapy may trigger similar effects. In contrast, Myt1 mutants exhibit a completely different spectrum of cellular defects, affecting stem cell proliferation and sensory organ development as well as male meiosis. Insights gained from studies of these mutants may thus be relevant to understanding the recent proposal that human PKMyt1 plays an important role in glioblastoma. Our studies of these genetically tractable animal models provide opportunities for learning about how these specialized cell cycle regulators function in physiologically meaningful settings. Such information could help us to better understand how manipulating Cdk1 inhibitory phosphorylation can be used to help treat cancer without compromising patient viability.

Dr. Armin Gamper Oncology When 1 + 1 Does not Equal 2: Conditional Synthetic Lethality as Strategy for Cancer Therapy Synthetic lethality refers to an interaction between two genes when the perturbation of either gene alone is viable but the simultaneous perturbation of both genes leads to cell death. The discovery that BRCA1 or BRCA2 mutant cancers are hypersensitive to PARP inhibitors, led to therapeutic approaches targeting cancer cells with deficiencies in one DNA damage response (DDR) pathway by inhibition of an alternative DDR pathway. As this approach only targets cells with a defective DNA damage response, it is bound to only affect a subset of cancers or populations within a tumour. Resistance can arise by reactivation of the defective pathway. Conditional synthetic lethality refers to synthetic lethality observed only under certain circumstances, such as genetic background or metabolic state of cells or cellular environment and exposure to therapeutic agents. In view of the inter- and intratumour heterogeneity commonly observed, to achieve the necessary therapeutic window for a wide range of tumours it is necessary to identify a common “condition” upon which to base cancer-selective conditional synthetic lethality. Oncogene-induced DNA damage is a common feature of cancer cells leading to high levels of replication stress as well as mitotic stress in cancer cells compared to normal proliferating cells. Based on this tumour- specific property, we tested in an orthotopic breast cancer xenograft model whether increasing genotoxic stress and simultaneously inhibiting an important rescue pathway would lead to cancer cell-specific death. We identified a drug combination that both inhibited tumour growth and metastasis with minimal side effects.

Dr. Gordon Chan Oncology Prolonged Mitotic Arrest induced by Wee1 Inhibition Sensitizes Breast Cancer Cells to Paclitaxel Wee1 kinase is a crucial negative regulator of Cdk1/cyclin B1 activity and is required for normal entry into and exit from mitosis. Wee1 activity can be chemically inhibited by the small molecule MK-1775, which is currently being tested in phase I/II clinical trials in combination with other anti-cancer drugs. MK-1775 promotes cancer cells to bypass the cell- cycle checkpoints and prematurely enter mitosis. In our study, we show premature mitotic cells that arise from MK-1775 treatment exhibited centromere fragmentation, a morphological feature of mitotic catastrophe that is characterized by centromeres and kinetochore proteins that co-cluster away from the condensed chromosomes. In addition to stimulating early mitotic entry, MK- 1775 treatment also delayed mitotic exit. Specifically, cells treated with MK-1775 following release from G1/S or prometaphase arrested in mitosis. MK-1775 induced arrest occurred at metaphase and thus, cells required 12 times longer to transition into anaphase compared to controls. Consistent with an arrest in mitosis, MK-1775 treated prometaphase cells maintained high cyclin B1 and low phospho-tyrosine 15 Cdk1. Importantly, MK-1775 induced mitotic arrest resulted in cell death regardless the of cell-cycle phase prior to treatment suggesting that Wee1 inhibitors are also antimitotic agents. We found that paclitaxel enhances MK-1775 mediated cell killing. HeLa and different breast cancer cell lines (T-47D, MCF7, MDA-MB-468 and MDA-MB-231) treated with different concentrations of MK-1775 and low dose paclitaxel exhibited reduced cell survival compared to mono-treatments. Our data highlight a new potential strategy for enhancing MK-1775 mediated cell killing in breast cancer cells.

Fahed Elian, PhD Student Medicine and Dentistry Medical Genetics Molecular Analysis of FOXC1 in Triple Negative/Basal-Like Breast Cancer (TN/BL BC) Fahed A Elian, David N Brindley, Michael A Walter Rapidly accumulating evidence implicates forkhead box C1 (FOXC1) in Triple Negative Breast Cancer (TNBC), particularly in Basal-Like Breast cancer (BLBC). Recently additional studies have demonstrating that FOXC1 is also a major player in hepatocellular carcinoma (HCC), endometrial cancer, Hodgkin’s lymphoma (HL), non-Hodgkin’s lymphoma (NHL), and others. The FOXC1 gene encodes a transcription factor that is crucial to mesodermal, neural crest, and ocular development. Loss of function mutations in FOXC1 have been shown to cause autosomal dominantly inherited Axenfeld-Rieger’s Syndrome (ARS), a developmental disorder associated in eye anomalies and glaucoma. Interestingly, while FOXC1 missense mutations that cause ARS reduce FOXC1 activity, increased FOXC1 function appears now often linked to more aggressive cancer phenotypes in TN/BL-BC, HCC, HL, and NHL.

We have investigated the mechanism(s) by which FOXC1 activity is increased in TNBC/BLBC. Samples were obtained from TNBC tumors, Triple Positive Breast Cancer (TPBC) tumors or from normal patient breast tissue. Using quantitative PCR, we found that FOXC1 is significantly over-expressed in TNBC patients as compared to controls. In contrast, FOXC1 mRNA was significantly less expressed in TPBC samples as compared to either control or TNBC samples. FOXC1 protein half-life was found to be significantly longer in the TNBC/BLBC cell line (BT 549) compared to FOXC1 protein’s half-life in HeLa cells. Studies of FOXC1 Copy- number variation (CNV) in TNBC/BLBC cell lines reveal that cell lines that have higher levels of FOXC1 protein (HS 587T, BT-549) have an extra copies of FOXC1, in contrast with cell lines with lower expression of FOXC1 protein (MDA-MB-231) may have a deletion of FOXC1. Our

results suggest that increased FOXC1 function in TNBC/BLBC results from over-expression of FOXC1 in TNBC patients that appears to be the result of changes to FOXC1 stability and amplification of FOXC1 copy number. We anticipate that understanding the mechanism(s) underlying FOXC1’s activation in cancer may aid in designing improved therapies for cancer patients.

Dr. Gabrielle Siegers, Research Associate Medicine & Dentistry Oncology Functional plasticity of gamma delta T cells and breast tumour targets in hypoxia Gabrielle M. Siegers, Indrani Dutta, Raymond Lai and Lynne-Marie Postovit The interaction of immune and tumour cells in the tumour microenvironment (TME) largely dictates patient outcome, yet remains poorly understood. In addition, the TME is characterized by biophysical features such as hypoxia (low oxygen). Gamma delta T cells (GDTc) naturally kill transformed cells and are therefore under development as immunotherapy for various cancers. Clinical trials have proven the safety of GDTc immunotherapy, and increased circulating GDTc levels correlate with improved patient outcome. Yet, the function of GDTc tumor infiltrating lymphocytes (TIL) in human breast cancer remains controversial.

We have studied the impact of hypoxia on expanded primary human GDTc. In vitro, GDTc viability under hypoxia (2% oxygen) increased compared to normoxia (20% oxygen). Surface marker analysis by flow cytometry suggested enhanced activation of GDTc exposed to hypoxia; in particular, CD56 was significantly upregulated under low oxygen. Cytokine arrays revealed significant increases in secretion of MIP1α/MIP1β, CCL5/RANTES and CD40L/TNFSF5 under hypoxia compared to normoxia. Hypoxia exposure for ≥48 hours prior to co-incubation with breast tumour targets increased cytotoxicity of most donor GDTc against targets maintained in normoxia; conversely, breast cancer lines incubated in hypoxia for 48 hours prior to the assay were astonishingly resistant to GDTc cytotoxicity. Blocking assays

conducted under normoxia confirmed the importance of NKG2D on GDTc and the corresponding NKG2D ligand MICA/B on tumour targets.

We are now investigating the impact of hypoxia on tumour antigens recognized by GDTc on targets, to determine whether global downregulation or antigen shedding accounts for their remarkable resistance to GDTc killing. Furthermore, we are assessing whether GDTc are present in hypoxic regions of breast tumour tissues and attempting to model this in patient- derived xenograft models of breast cancer. Understanding GDTc TIL function in breast cancer will be of utmost importance to develop effective GDTc immunotherapies.

Jiangjie Xiao, PhD Student ALES AFNS Associations of Pre-existing Comorbidities with Skeletal Muscle Mass and Radiodensity in Early Stage Colorectal Cancer Patients Jingjie Xiao, MSc, Bette J. Caan, DrPH, Erin Weltzien, Elizabeth M. Cespedes Feliciano, ScD, MSc, Candyce H. Kroenke, ScD, MPH, Jeffrey A. Meyerhardt, MD, MPH, Vickie E. Baracos, PhD, Marilyn L. Kwan, PhD, Adrienne L. Castillo, RD, Carla M. Prado, PhD, RD Background: Comorbidities and computerized tomography (CT)-measured muscle abnormalities are both common in cancer patients and evident in diseases such as diabetes and obesity. This is the first study to examine the association between comorbidities and muscle abnormalities in patients with colorectal cancer (CRC). Methods: This cross-sectional study included 3051 patients with stage I-III CRC. Muscle abnormalities were defined as low skeletal muscle mass index (SMI) or low skeletal muscle radiodensity (SMD) quantified using diagnostic CT images. Charlson comorbidities were ascertained. Chi-square tests were used to compare the prevalence of comorbidities by the presence/absence of each muscle abnormality. Logistic regression analyses were performed

to evaluate which comorbidities predicted muscle abnormalities adjusting for age, sex, body mass index, weight change, stage, race/ethnicity, and smoking. Results: Mean age was 63 years; 50% of patients were male. The prevalence of low SMI and low SMD were 43.1% and 30.2%, respectively. Comorbidities examined were more prevalent in patients with low SMD than in those with normal SMD (58.6% vs. 34.4%, p<0.001), and most remained significant predictors of low SMD, including myocardial infarction (odds ratio [OR]=1.82, p=0.017), congestive heart failure (OR=3.34, p<0.001), peripheral vascular disease (OR=2.20, p<0.001), diabetes with or without complications (OR=1.61, p=0.008; OR=1.47, p=0.003, respectively) and renal disease (OR=2.23, p<0.001). No comorbidities were associated with low SMI except for diabetes with complication which was associated with a lower likelihood of low SMI (OR=0.64, p=0.007). Conclusion: Prevalence of muscle abnormalities was high in early stage CRC. Pre-existing comorbidities were most commonly associated with low SMD, suggestive of a potential shared mechanism between fat infiltration into muscle and each of these comorbidities.

SPEED POSTER ABSTRACTS

Pages 22-124

2 Guanmin Meng Medicine & Dentistry Biochemistry An inflammatory response involving increased autotaxin production in human adipose tissue after radiotherapy: implications for breast cancer treatment Guanmin Meng, Xiaoyun Tang, Zelei Yang, Matthew G.K. Benesch, Alison Marshall, David Murray, Denise G. Hemmings, Frank Wuest, Todd P.W. McMullen and David N. Brindley Background: Our group developed a new paradigm for understanding breast tumor progression and treatment in which adipose tissue adjacent to breast tumors becomes inflamed by inflammatory cytokines from the tumor. This stimulates autotaxin (ATX) secretion from adipocytes, whereas breast cancer cells produce insignificant ATX. Lysophosphatidate (LPA) produced by autotaxin promotes a vicious cycle of inflammation by stimulating the secretion of more inflammatory mediators, which increases further ATX production. This LPA-induced inflammation promotes tumor growth, metastasis and resistance to chemotherapy. LPA signaling also protects cells from radiation-induced cell death. Neo-adjuvant or post-adjuvant radiation treatment for breast cancer patients normally involves irradiating the whole breast with 25 fractions of ~2 Gy.

Hypothesis: Irradiation-induced damage to breast adipose tissue will increase the secretion of ATX and other inflammatory mediators that should protect residual cancer cells against further fractions of radiotherapy.

Experiments: Cultured rat abdominal adipose tissue and human breast and neck adipose tissue were exposed to 0.25 to 5 Gy of γ-radiation. This increased mRNA concentrations for ATX, cyclooxygenase-2, IL-1β, Il-6, IL-10, TNFα and LPA1/2 receptors. There was also increased secretion of ATX and 14 inflammatory mediators from human adipose tissue. Blocking signaling downstream of radiation-induced damage by inhibiting the activities of NFκB, cyclooxygenase-2, poly[ADP-ribose]polymerase-1(PARP-1) or ataxia telangiectasia and

Rad3-related protein (ATR) prevented the inflammatory responses to γ-radiation. Furthermore, higher ATX activity and levels of IL-2, CXCL5 and TNFα, together with lower levels of anti-inflammatory adiponectin, were detected in the plasma of mice where only part of the breast adipose tissue was irradiated with 7.5 Gy.

Conclusions: Radiation-induced damage of breast adipose tissue establishes a comprehensive inflammatory response, which involves increased signaling by LPA, COX-2 and other inflammatory mediators. We will now test if blocking this inflammatory response with an ATX inhibitor will increase the efficacy of focused radiotherapy in destroying breast tumors.

7 Laura Lee Medicine and Dentistry Oncology Hypoxia: epigenetic modulator in the induction of cancer cell plasticity Lee LJ, Postovit LM. Phenotypic plasticity affords cancer cells with the ability to metastasize and resist therapies, leading to reduced survival in patients. A tumor micro-environmental factor that can lead to an induction of plasticity is hypoxia, a condition of low oxygen. Tumor hypoxia can lead to an induction of various stem cell and epithelial-to-mesenchymal transition markers, surrogates for measuring cancer cell plasticity. Hypoxic reprogramming of the cancer cell can be achieved through changes in the epigenome, with global alterations in histone modifications. Here we propose to address how the epigenomes of breast cancer cells respond to hypoxia by examining the alterations in histone modifications in concert with the resulting transcriptional response. T47D and MDA-MB-231 breast cancer cell lines, epithelial and mesenchymal-like respectively, were used. Using ChIP-seq, we examined the global changes in repressive (H3K27me3) and active (H3K4me3) histone marks, which were chosen for their association with hypoxia and regulation of the stem cell phenotype. Structural changes in

chromatin was interrogated using electron spectroscopic imaging. RNA-seq was employed for characterization of the resulting transcriptome. We identified striking differences in the hypoxic responses of T47D and MBA-MB-231 cells, encompassing epigenetic and transcriptional alterations. T47D cells responded more robustly to chronic hypoxia (0.5% O2, 48 hours), in both histone modification alterations, overall chromatin condensation, and differentially expressed genes. Our findings indicate that the epigenetic and transcriptional responses to hypoxia are highly context dependent and heterogeneous. Elucidating the nuanced cellular response to hypoxia will better our understanding of hypoxia induced cancer cell plasticity, leading to improved therapeutic targets.

10 Mohammad Ali Medicine & Dentistry Oncology Matrix metalloproteinase-2 facilitates ribosomal RNA transcription by cleaving histones in the nucleoli Mohammad A.M. Ali*, Javier A. Garcia*, Ramses Ilarraza, Michael Hendzel, and Richard Schulz. Background: Cellular proliferation requires synchronization of multiple cellular organelles including the nucleolus. The nucleolus, a specialized region of the nucleus, encodes genes for ribosomal RNA (rRNA). Epigenetic regulation, also important in cellular replication, includes post-translational modifications of the N-terminal tail of histones. Matrix metalloproteinase- 2 (MMP-2) is localized to the nucleus but its role there remains elusive. We report here the localization of MMP-2 to sites within the nucleolus where rRNA is transcribed and provide evidence that it stimulates rRNA transcription and cell proliferation. Methods: Using biochemical and microscopy techniques we determined the presence of MMP-2 in nucleoli of human bone osteosarcoma (U-2 OS) cells. We used in vitro cleavage

assays testing active MMP-2 on different histones to determine if they could be regulatory targets. ChIP-qPCR was used to map MMP-2 on the rDNA gene promoter and the effects on rRNA transcription were assessed by qRT-PCR. Results: Confocal imaging revealed that MMP-2 accumulates at active rRNA transcription sites within the nucleolus. Nucleolar ChIP-qPCR showed that MMP-2 is enriched on the rDNA gene promoter suggesting a role in regulating rRNA transcription. Indeed, knocking down MMP-2 or inhibiting its activity in cells decreased pre-rRNA levels and inhibited cell proliferation similarly to actinomycin D. MMP-2 proteolyzed different histones in vitro, including cleaving the N-terminal tail of histone H3 which facilitates the transcription of RNA polymerase II-transcribed genes. Notably, cleaved histone H3 was found in histone preparations isolated from purified nucleoli. Our results reveal that MMP-2-mediated proteolysis of the histone H3 N-terminus in the nucleolus facilitates rRNA transcription and thus proliferation. Conclusions: Nucleolar MMP-2 positively regulates rRNA gene expression through cleavage of histones. This novel intranucleolar mechanism of MMP-2, which previously was only thought to act extracellularly in facilitating tumour cell proliferation, adds to the variety of roles MMPs play in cancer.

19 Mackenzie Coatham Medicine and Dentistry Obstetrics and Gynecology Utilizing Cell Line Models with Chromatin Remodelling Protein Deficiencies to Study Aggressive Dedifferentiated Endometrial Cancer Mackenzie Coatham, Cheng-Han Lee, Lynne-Marie Postovit An estimated 6600 Canadian women were diagnosed with uterine cancer in 2016, with 20% of women succumbing to the disease. One of the most lethal subsets of uterine cancer is dedifferentiated endometrial carcinoma (DDEC). DDEC tumors possess both well-

differentiated and undifferentiated regions. The majority of metastatic disease is made up of cells from the undifferentiated component of DDEC yet it is unclear how these poorly differentiated regions are initiated and sustained within the carcinoma.

Examining the well-differentiated and undifferentiated components of DDEC tumors in terms of mutation profiles, we demonstrated that 80% of the undifferentiated regions in DDEC lesions lack the expression of core chromatin remodeling proteins, BRG1 or ARID1A and ARID1B. We hypothesize that the loss of these proteins, which are known regulators of transcription, may lead to the induction and/or maintenance of stem cell-like gene expression programs that drive dedifferentiation, metastasis and therapy resistance.

BRG1-deficient or ARID1A/B co-deficient cell line models were generated by CRISPR and validated using immunofluorescence and immunohistochemistry. qRT-PCR and immunofluorescence were used to assess the level of expression and localization of markers of epithelial-to-mesenchymal transition (EMT), stemness and endometrial lineage. The ability of the generated knockouts to proliferate and form spheres was evaluated, as well as an examination of the response of these BRG1-deficient or ARID1A/B co-deficient cell line models to current clinically relevant epigenetic inhibitors (vorinstat and tazemetostat). Tumor formation in immune-compromised mice will be monitored to ascertain any histological differences between wildtype and BRG1 or ARID1A/B knockout endometrial cancer cells.

BRG1 deficient endometrial cancer cell lines have been shown to partially undergo EMT and recapitulate the clinical DDEC phenotype. Determining the extent to which loss of BRG1 or ARID1A/B contributes to the acquisition of DDEC is a critical step towards improving diagnostic and treatment practices for aggressive stem-like forms of gynecological cancers.

24 Bindu Thapa Faculty of Pharmacy and Pharmaceutical Sciences Pharmaceutical Sciences Identification of complementary therapeutic targets to sensitize TRAIL-induced apoptosis in breast cancer cells via RNAi screening Bindu Thapa, Remant Bahadur KC, Hasan Uludag Identification of complementary therapeutic targets to sensitize TRAIL-induced apoptosis in breast cancer cells via RNAi screening

Bindu Thapa1, Remant Bahadur KC2, Hasan Uludağ1,2,3 1 Faculty of Pharmacy and Pharmaceutical Sciences, 2 Department of Chemical and Material Engineering, Faculty of Engineering, 3 Department of Biomedical Engineering, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta, Canada Inducing apoptosis has become a central therapeutic strategy employed in cancer management, which bears a strong potential to eradicate cancer cells. Tumor necrosis factor- related apoptosis-inducing ligand (TRAIL) induces apoptosis in variety of cancer cells without affecting most normal cells, making it a promising agent for cancer therapy. However, TRAIL therapy is clinically ineffective due to resistance induction in malignant cells. To identify novel protein targets whose silencing sensitize breast cancer cells against TRAIL, a siRNA library against 446 human apoptosis-related proteins were screened in breast cancer cells; MDA-231 in presence or absence of TRAIL. A library of small cationic lipopolymers were synthesized, screened for siRNA delivery and most effect one was used as siRNA delivery agent. Based on the inhibition of cell growth of MDA-231 cells, sixteen siRNAs were found to sensitize TRAIL- induced cell death. Among them, novel and the most promising targets BCL2L12 and SOD1 were further evaluated. Silencing both targets sensitized TRAIL-induced death in MDA-231 cells and TRAIL-resistant MCF-7 cells. Importantly, TRAIL and siRNA silencing BCL2L12 had no effect in normal cells; human umbilical vein cells (HUVEC) and human bone marrow stem cell (hBMSC). Hence, siRNAs targeting BCL2L12 and SOD1 were found to be novel regulators of

TRAIL-induced cell death in breast cancer cells, providing a new approach for enhancing TRAIL therapy. The combination of TRAIL and siRNA targeting BCL2L12 can be the most effective synergistic pair for breast cancer therapy without affecting non-transformed cells.

29 Kyle Potts Medicine and Dentistry Medical Microbiology and Immunology Novel oncolytic vaccinia virus as a treatment for Bacillus Calmette-Guérin refractory bladder cancer Kyle G. Potts, Megan A. Desaulniers, Chad R. Irwin, Ronald B. Moore, David H. Evans, and Mary M. Hitt Bladder cancer (BCa) has a recurrence rate of 80% and many patients require multiple treatments that often fail, leading to disease progression. Standard of care for high-grade disease, Bacillus Calmette–Guérin (BCG), fails in 30% of patients. Recently, it was found that different abilities of BCa cells to internalize BCG is determined by Pak1 mediated macropinocytosis. It is hypothesized that internalization of BCG is a critical step in activating the anti-tumor response. Interestingly vaccinia virus (VACV) utilizes multiple modes of entry, including fusion with the cellular membrane or through macropinocytosis.

We have reported on a VACV with deletions in F4L (ribonucleotide reductase small subunit) and J2R (thymidine kinase) that exhibits oncolytic properties and promotes anti-tumor immunity with superior safety in BCa. In a screen of BCa cells, we found significant differences in BCG internalization while, delF4LdelJ2R VACV could replicate in and killed both BCG susceptible and resistant cell lines to similar levels. Comparison of delF4LdelJ2R VACV and BCG in the orthotopic AY-27 immunocompetent rat model shows that the virus induced superior tumor clearance and long term survival. Pretreating cells with a Pak1 inhibitor, or

inducing BCG-resistance with Pak1 shRNA, showed decreased BCG uptake with little effect on VACV replication. Finally, we have developed an AY-27-shPak1 cell line that is resistant to BCG uptake that we will use to generate an BCG-resistant BCa model.

Given the high rate of recurrence and the lack of treatment options for BCG-resistant BCa, our delF4LdelJ2R VACV could provide a safe and urgently needed therapy.

30 Deepak Dinakaran Medicine & Dentistry Oncology Novel X-ray Activated Photodynamic (radioPDT) Theranostic Nanoparticles for Deep Seated Tumors Dr Deepak Dinakaran, Dr Jayeeta Sengupta, Dr Hua Chen, Dr Nawaid Usmani, Dr Piyush Kumar, Dr Ravin Narain, Dr John Lewis, Dr Ron Moore Background: Radiotherapy uses precision technology to treat prostate cancer and minimize side effects, but damage to surrounding tissues limit the therapeutic dose delivered. Photodynamic therapy (PDT) uses photosensitizers that, when exposed to light, generate reactive singlet oxygen species to cause remarkable tumor cytotoxicity. The dependence on delivering the activating light, which has restricted tissue penetrance, limits PDT’s use in deep seated cancers. Radiotherapy’s X-rays have unlimited depth penetrance. Therefore, X-ray activated PDT (radioPDT) could be advantageous in treating deep seated tumors. This is done with scintillating nanoparticles that convert X-rays to visible light to activate PDT. Currently, there is no data on how radioPDT is affected by tumor hypoxia, which is relevant because both radiotherapy and PDT are oxygen consumptive processes. Therefore, our objective is to characterize radioPDT in hypoxia, and its impact on singlet oxygen yield and cancer cell death.

Method: We developed a novel radioPDT nanoparticle by encapsulating nanoscintillators (lanthanum fluoride) and photosensitizers (protoporphyrin) into nanocarriers (PEG-PLGA). Therapeutic potential was evaluated via singlet oxygen yield and cell viability assay in normoxic and hypoxic conditions under radiation and light PDT. As a secondary objective, the nanoparticle’s diagnostic properties were evaluated via CT and MRI, as well as X-ray imaging of in vivo distribution characteristics using a chorioallantoic membrane (CAM) chick embryo model. Results: Preliminary data demonstrates radioPDT effectively augments radiotherapy in hypoxia via singlet oxygen production. In vitro viability assays shows minimal intrinsic toxicity in prostate cancer and fibroblast cell lines when not activated, and significant increase in cytotoxicity when activated. CT and MRI studies show appreciable signal generation, and CAM model studies show preferential tumor-targeting capabilities. Future studies are aimed at exploring their diagnostic and therapeutic potential in vivo in tumor-bearing mice models. Conclusion: radioPDT may offer a novel method of augmenting traditional radiotherapy and minimizing toxicity.

4Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, AB, Canada The ERCC1-XPF heterodimer is a 5ʹ-3ʹ structure-specific endonuclease which is essential for nucleotide excision repair (NER) and inter-strand crosslink (ICL) DNA repair pathways. Although its action is critical to maintain genome integrity and stability and prevent against damage-induced mutations, it can counteract the effect of DNA damaging therapies such as platinum-based chemotherapy and radiotherapy. A promising approach to enhance the effect of such therapies is to inhibit the action of XPF-ERCC1 in cancer cells using small molecules. In this study, we used a computational drug design workflow to rationally design novel analogues of the F06 molecule, a lead inhibitor which targets the dimerization between XPF and ERCC1, required for the functional endonuclease. We synthesized seven compounds, which show in silico show interesting binding affinities and ligand efficiency towards an XPF site on the dimerization interface. This approach yielded compound 3 and 4 as potent inhibitors of ERCC1-XPF activity. An iIn vitro ERCC1-XPF endonuclease assay identified compound 4 as the best ERCC1-XPF inhibitor with an IC50 value of 0.3 µM. Also, the Kd value of this compound was measured as ~100 nM with by fluorescence spectroscopy. Compound 4 also showed a significant inhibition of the removal of cyclobutane pyrimidine dimers (CPDs) compared with control cells after exposure of HCT 116 cells to UV radiation. Compound 4 can potentially be used in combination with existing DNA-damaging therapies to amplify their effect on cancer cells.

37 Michael Taylor Medicine & Dentistry Medicine Alberta Rectal Cancer Initiative: Implementation of a Provincial Rectal Cancer Clinical Pathway Quality Improvement Project Michael Taylor, W. Donald Buie, Todd McMullen BACKGROUND: A multi-disciplinary clinical pathway with discipline-specific goals was created. Following baseline data collection, stakeholders from radiology, oncology, surgery, and pathology were engaged to standardize care and inform reporting schemas. Education days with international experts were held to reinforce best practice, and synoptic reporting templates were developed.

METHODOLOGY: Quality indicators determining best practice adherence and oncologic outcomes were collated and reported. Pathology reports were used as quality assurance for surgical technique and MRI-staging. The appropriate use of neoadjuvant therapy was correlated with collaborative staging. These measures were then used to provide ongoing individualized audit/feedback reports to practitioners through a secure web-based portal; reports contain individual physician data and aggregate provincial data for each indicator to inform and improve practice.

‘complete’ or ‘near-complete’ and a margin positivity of 7% on pathology. Nearly all (94%) pathology reports were completed synoptically, with 90% reporting all mandatory data elements.

CONCLUSION: Implementation of a clinical pathway for rectal cancer has improved uptake of best practice across the clinical continuum; this sustainable multifaceted approach includes education, engagement, feedback reporting, and is easily adaptable to other tumor groups.

single 30-minute aerobic exercise session performed 24 hours prior to each treatment; 2) a 50% reduction in caloric intake for 48 hours prior to each treatment; or 3) usual care. Outcome measures will include reserve capacity (peak capacity – rest) of left ventricular ejection fraction, aortic distensibility, isolated skeletal muscle oxygen consumption, and whole body oxygen consumption (i.e. aerobic fitness). Peak capacity will be measured using maximal incremental exercise tests. Cardiac, aortic, and skeletal muscle function will be measured using novel magnetic resonance imaging (MRI) techniques. Outcomes will be assessed prior to chemotherapy, end of treatment, and one year after starting treatment. Exploratory outcomes will include patient-reported quality of life and fatigue, and clinically measured tumor size at the end of treatment in patients receiving neoadjuvant treatment. These interventions will be immediately accessible by all patients at all institutions and have the potential to improve the health of breast cancer survivors.

POSTER ABSTRACTS

Pages 36-157

1 Rachel Maki Science Biological Sciences “O-GlcNAcylation” as a Potential Myt1 kinase Regulator in Drosophila melanogaster Kylie M. Parent, JoAnn J. Thai, Rachel J. Maki & Shelagh D. Campbell Regulation of the cell cycle and its checkpoints are essential for proper proliferation, beginning with chromosomal events during mitosis. Chromosomal aberrations such as aneuploidy are often linked to human cancer, with aneuploidy classified as a distinct chromosomal mutation involving malignant transformation and proliferation. O- GlcNAcylation, a type of post-transcriptional modification involving O-GlucNAc transferase (OGT), is a form of cell regulation activated by nutrient level changes in the cellular environment. We are interested in O-GlcNAcylation as a potential regulator of Myt1 kinase, a cell cycle “brake” that inhibits the activity of cyclin dependent kinase 1 (CDK1) by phosphorylation.

Previous studies in our laboratory showed that loss of Drosophila Myt1 activity resulted in multipolar meiotic divisions, leading to aneuploid spermatids. In mammalian cells, elevation of OGT activity was reported to cause abnormal cell cycle arrest due to Myt1 inhibition of Cdk1, suggesting this as a new molecular mechanism linking nutritional cues with cell cycle control.

To study how OGT might affect cell cycle control in Drosophila we are using the UAS-Gal4 system to generate transgenic lines that over- or under express OGT in the germline, wing tissue or sensory organ precursors. We observed mutant wing phenotypes in strains depleted for OGT, indicating that abnormal levels of OGT cause developmental defects. In contrast, F1 progeny depleted for OGT in the germline were fertile. We will be performing further experiments to verify these results and investigate the impact of OGT on Myt1 protein modification using a Myt1 reporter for quantitative analysis by western blotting.

3 Scott Findlay Medicine and Dentistry Oncology A digital PCR-based method for rapid and sensitive mutation validation in genome edited cancer cells Scott D Findlay, Krista M Vincent, Jennifer Berman, Lynne-Marie Postovit The rapid adoption of precision gene editing tools such as CRISPRs for cancer research and eventually therapeutics necessitates assays that can rapidly detect and quantitate desired alterations. While validation by deep sequencing serves as a gold standard, associated technologies are not practical for many projects and are not yet cost-effective for many researchers. Thus, the most commonly used assay to validate genome alterations today employs a mismatch nuclease such as T7E1 that recognizes and cleaves heteroduplexed DNA amplicons. While useful, this assay has many shortcomings: It is prone to false positives due to cancer-associated mutations and/or single nucleotide polymorphisms (SNPs), requires large amounts of starting material, and cannot discriminate compound heterozygote clones from those with both altered and wild type alleles. Here we describe a powerful alternative wherein droplet digital PCR (ddPCR) can be used to

validate desired mutations using duplexed primer-probe assays. These assays allow single molecule-based simultaneous quantification of indels resulting from non-homologous end joining (NHEJ) as well as unaltered wild type alleles at target loci. We generated human breast cancer and melanoma clonal cell lines containing functional knockout-inducing indel mutations for the NODAL or SFRP1 genes to compare the performance of ddPCR assays to mismatch nuclease assays at these loci. The ddPCR assays offered extraordinary sensitivity and readily discriminated compound heterozygotes from clones with both wild type and altered alleles. Collectively, this work highlights the ability of ddPCR-based screening to contribute to a sensitive, specific, and streamlined approach for the validation of desired alterations in genome editing workflows.

4 Dong-Woo Kang Physical Education and Recreation NA A Phase II Randomized Controlled Trial of Exercise in Prostate Cancer Patients Undergoing Active Surveillance: A Study Protocol Dong-Woo Kang, MSc., Normand G. Boulé, PhD., Adrian S. Fairey, MD., Catherine J. Field, PhD., Kerry S. Courneya, PhD RATIONALE: Active surveillance (AS) is the preferred primary treatment strategy for men with low risk clinically localized prostate cancer (PCa), however, approximately 50% of men require radical treatment within 10 years. Interventions designed to suppress tumour growth, mitigate fear of cancer progression, and precondition men for impending radical treatments presents an unmet clinical need. Exercise has been shown to delay the progression of tumors in animal models through alterations in immune function; however, these findings have not been confirmed in clinical settings. OBJECTIVE: The primary aim of this study is to examine the effects of exercise on anti-cancer

immune activity related to tumour progression, physical fitness, and fear of cancer progression in PCa patients on AS. METHODS: This phase II randomized controlled trial will randomize 66 men undergoing AS to either an exercise group or usual care group. The exercise group will perform a 12-week, supervised, high-intensity interval aerobic exercise program, consisting of 3 sessions/week and 30 minutes/session. The primary outcome will be peak oxygen consumption (VO2peak) as a measure of physical fitness. Secondary outcomes will include systemic immune function and tumour-related biomarkers, fear of cancer progression, and cancer-related symptoms. Exploratory outcomes will include tumour progression variables using PSA and endo-rectal mp-MRI. The trial has 80% power to detect a significant between-group difference in VO2peak of 3.5 ml/kg/min with a two-tailed alpha level of less than 0.05 and a 10% drop-out rate. SIGNIFICANCE: This trial may establish the feasibility and preliminary efficacy of exercise in PCa patients receiving AS and inform larger phase II and III trials on exercise in this setting. If positive, this line of research may support exercise as a low cost, low toxicity intervention that may help PCa patients on AS to remain on AS longer and better prepare for the eventuality of radical treatments.

5 Noureen Ali Medicine and Dentistry Pediatrics Active Beta-Catenin (ABC) as a prognostic marker for Osteosarcoma progression Noureen Ali, Geetha Venkateswaran, Elizabeth Garcia, Sujata Persad Introduction: Osteosarcoma (OS) are aggressive primary bone malignancies having peak incidences in children and young adolescents. Outcome remains poor for most patients with metastatic disease which develops in 1 of every 5 cases. Presently, there are no widely recommended screening tests for early diagnosis of OS and no reliable prognostic marker for

aggressive/metastatic disease. We investigated the putative role of the Wnt/Beta-catenin pathway, specifically the transcriptionally active form of Beta-catenin, Activated Beta-Catenin (ABC), in OS progression.

Methods: We used two pairs of cell lines that simulate OS progression: Saos2/Saos2-LM7 and HOS/HOS-143B. MMP2 and MMP9 activity were used to confirm the greater metastatic potential of Saos2-LM7 & HOS-143B compared to SaOS2 and HOS respectively. Total cellular/nuclear levels/localization of ABC/Beta-catenin were evaluated by Western blot and immunofluorescence (IF) analysis. High Content analysis of IF was used for quantification of cellular/nuclear levels of ABC/Beta-catenin. Transcriptional activity of ABC/Beta-catenin was evaluated by RT-qPCR of target genes (MMP2, MMP9, Cyclin D1, VEGFA) and TopFlash activity.

Results: Results show significantly higher cellular levels of ABC in the SaOS2-LM7 and HOS- 143B cell lines compared to the respective parent cell lines (SaOS2 & HOS). Additionally, ABC exhibited a more prominent nuclear localization in the SaOS2-LM7 and HOS-143B cell lines compared to SaOS2 & HOS, respectively. No significant differences in cellular levels and localization of Beta-catenin were observed. SaOS2-LM7/HOS-143B exhibited significantly greater transcriptional activity.

Conclusion: The strong correlation between cellular/nuclear ABC levels/activity and OS progression supports the potential for ABC to serve as a prognostic marker for OS progression.

6 Huachen Chen Medicine & Dentistry Oncology ZIC2 regulates tumorigenic phenotypes in epithelial ovarian cancer Huachen Chen*, Krista Vincent, Zhihua Xu, Lynne-Marie Postovit, YangXin Fu Objectives Epithelial ovarian cancer (EOC) is the most lethal gynecological malignancy. Current regimens against advanced EOCs are inefficient owing to disease relapses and acquired chemoresistance driven by cancer stem cells (CSCs). The transcription factor ZIC2 has been discovered as a potential factor regulating self-renewal of CSCs in several cancer types. This project focuses on the study of the relationship between transcription factor ZIC2 and tumorigenic phenotypes in EOC.

Methods The EOC patient survival and ZIC2 mRNA expression from The Cancer Genome Atlas (TCGA) were analyzed for correlation. The mRNA levels of ZIC2 and Oct4 (a major pluripotency factor) was examined in a panel of EOC cell lines. ZIC2 knockout models established by CRISPR/cas9 were subjected to immunoblotting and quantitative (q)RT-PCR analysis for expressions of OCT4 and ALDH1A1 (the most commonly used ovarian CSC marker). ALDEFLUORTM assay was used to determine the CSC population in these models. Biological functions of ZIC2 in EOC were examined by clonogenic assay, soft agar assay, and sphere formation assay.

Results The Kaplan-Meier survival plot indicates that higher ZIC2 mRNA level is associated with shorter survival of EOC patients. ZIC2 expression is positively correlated with pluripotency factor Oct4 expression in the panel of EOC cell lines. Functionally, ZIC2 knockout impaired the capacity of EOC cells to grow and form colonies and spheres. Furthermore, ZIC2 knockout

decreased expression of stem cell-associated genes and ALDH1+ populations in EOC cells, suggesting a role for ZIC2 in regulating CSCs in EOC.

Conclusions Our initial work provides insight into the biological functions of ZIC2 in regulating tumorigenic phenotypes in EOC.

encompassing epigenetic and transcriptional alterations. T47D cells responded more robustly to chronic hypoxia (0.5% O2, 48 hours), in both histone modification alterations, overall chromatin condensation, and differentially expressed genes. Our findings indicate that the epigenetic and transcriptional responses to hypoxia are highly context dependent and heterogeneous. Elucidating the nuanced cellular response to hypoxia will better our understanding of hypoxia induced cancer cell plasticity, leading to improved therapeutic targets.

8 Rory McDonald Science Chemistry Vacquinol-1 Analogs as a Potential Treatment for Glioblastoma Multiform by Exploiting Methuosis, a Novel Form of Nonapoptotic Cell Death Rory M. McDonald, Christian L. Jankovic, Samantha Kwok, Saket Jain, Roseline Godbout*, and Dennis G. Hall* Glioblastoma multiform (GBM) is a malignant form of glioma that is resistant to standard surgical and chemotherapy regimens. New therapeutics are desperately needed for improved treatment and prognosis of this devastating cancer. Vacquinols are a class of alpha- hydroxyalkyl piperidines that were discovered by a phenotypic screen against GBM cell lines. Testing these compounds in vitro revealed that they induced cell death of GBM cells via methuosis — a nonapoptotic pathway characterized by macropinocytosis and catastrophic vacuolization. Optimization of these compounds led to Vacquinol-1, which has promising biological activity (IC50 = 3.14 μm), and potential as a drug candidate for GBM. However, there is still opportunity to streamline its lengthy chemical synthesis, and improve its potency against GBM cells by studying its Structure-Activity Relationship (SAR) via the synthesis of enantioenriched analogs.

The Hall Group has developed and optimized a borylative migration reaction, where a piperidinyl alkenyl perfluorosulfonate is transformed into a enantioenriched allylic boronate ester. These products can then be further functionalized by Suzuki-Miyaura cross-coupling, and allylboration reactions. By using a unique stereoselective allylboration methodology with a chiral allylic piperidine and quinoline aldehyde, followed by functional group transformations, all erythro and threo stereoisomers of Vacquinol-1 at the C16 and C17 chiral centres can be synthesized. Vacquinol-1 analogs with an unsaturated piperidine ring, and substitutions on the 2(4-chlorophenyl) moiety on the quinoline ring can also be synthesized stereoselectively via this sequence. The Godbout lab can then test these Vacquinol-1 analogs against GBM cells using the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay and the Transwell migration assay to examine the analogs’ potency and effect on migration. SAR studies and in vitro testing of the analogs are ongoing.

9 Sahar Arbabimoghadam Physics Physics Amino Acid Frequency in Different Species Sahar Arbabimoghadam Proteins play critical roles in all biological processes. They are made of building blocks called amino acids. There are twenty standard amino acids which have varying properties depending on their side-chains. Amino acids are encoded by nucleotides. In fact, the genetic code has 64 possible permutations/combinations of three nucleotide sequences made from the four nitrogenous nucleotide bases: adenine, guanine, cytosine and thymine. Sixty-one of these combinations encode the twenty amino acids while the remaining three encode the stop codons. In this project, we studied both the amino acids Gibbs free energy of formation as well as the

Gibbs free energy of the encoding codons in double-stranded DNA. The energy of the latter was calculated as a sum of the individual nucleotide’s energy, the interaction energy with their nearest neighbor and the energy of the melting stability of the base pair stacking. We studied both these energies in relation to the frequencies of the individual amino acids. In addition, we also analyzed the amino acids frequencies in different species in the phylogenetic tree ranging from the simplest model, E. coli, to the most complex one, humans. Our results show that, in almost all species, the amino acid leucine is the most frequent while tryptophan is the least probable. Although both of these amino acids have a high energetic cost of formation, there are more biological pathways for leucine synthesis, which could explain its higher frequency. Surprisingly, cysteine was found to have the second lowest frequency although it has a low energetic cost of formation. Glycine, on the other hand, has the least energetic cost of formation and has a relatively high level of abundance. We are now working on correlating Adinkra symbols and neural network models to encode the genetic code.

2 (MMP-2) is localized to the nucleus but its role there remains elusive. We report here the localization of MMP-2 to sites within the nucleolus where rRNA is transcribed and provide evidence that it stimulates rRNA transcription and cell proliferation. Methods: Using biochemical and microscopy techniques we determined the presence of MMP-2 in nucleoli of human bone osteosarcoma (U-2 OS) cells. We used in vitro cleavage assays testing active MMP-2 on different histones to determine if they could be regulatory targets. ChIP-qPCR was used to map MMP-2 on the rDNA gene promoter and the effects on rRNA transcription were assessed by qRT-PCR. Results: Confocal imaging revealed that MMP-2 accumulates at active rRNA transcription sites within the nucleolus. Nucleolar ChIP-qPCR showed that MMP-2 is enriched on the rDNA gene promoter suggesting a role in regulating rRNA transcription. Indeed, knocking down MMP-2 or inhibiting its activity in cells decreased pre-rRNA levels and inhibited cell proliferation similarly to actinomycin D. MMP-2 proteolyzed different histones in vitro, including cleaving the N-terminal tail of histone H3 which facilitates the transcription of RNA polymerase II-transcribed genes. Notably, cleaved histone H3 was found in histone preparations isolated from purified nucleoli. Our results reveal that MMP-2-mediated proteolysis of the histone H3 N-terminus in the nucleolus facilitates rRNA transcription and thus proliferation. Conclusions: Nucleolar MMP-2 positively regulates rRNA gene expression through cleavage of histones. This novel intranucleolar mechanism of MMP-2, which previously was only thought to act extracellularly in facilitating tumour cell proliferation, adds to the variety of roles MMPs play in cancer.

11 Kirby Ziegler Medicine & Dentistry Oncology An integrative roadmap to PAX3 target gene networks in melanocytes and melanoma Kirby A. Ziegler and D. Alan Underhill Melanoma accounts for 70% of skin cancer-associated deaths, despite representing a small fraction of all skin cancers. The developmental origin of melanocytes is thought to be a driver of this aggressive behaviour. In this context, melanocyte identity is determined during embryogenesis by the hierarchical action of transcription factors. These proteins have key roles in melanoma, reflecting their capacity to control pathogenic gene expression. Within this scheme, PAX3 controls cell division and differentiation by recognizing specific target sequences in the genome and altering expression of associated genes. To this end, PAX3 contains two DNA-binding domains, the homeodomain and the paired domain, which itself comprises two subdomains. In theory, the inherent modularity in this architecture permits the recognition of multiple DNA-binding patterns, yet we have not had a comprehensive view of PAX3 DNA-binding specificity or associated targets. We hypothesize that PAX3 utilizes multiple modes of DNA recognition that can contribute to distinct functional outputs during melanoma progression. To address this, we have repurposed data derived from in vitro binding assays to statistically model PAX3 DNA-binding specificities. Significantly, these profiles represent the first set of optimal motifs described for full-length PAX3. This library was validated in situ by calculating its enrichment across published ChIP-seq datasets for exogenous PAX3 variants. This provided a foundation for predicting PAX3 occupancy in cell- specific regulatory regions defined using epigenomic signatures, across a range of PAX3- expressing cell types. To identify downstream targets of PAX3, we used RNA-sequencing to profile differential gene expression following PAX3 attenuation across melanocyte and melanoma cell lines. Putative targets were subsequently integrated with predicted PAX3 occupancy to connect distinct DNA-binding profiles to transcriptional pathways. Collectively, these analyses provide novel insight into the discrete target gene networks associated with

the differential use of PAX3 DNA-binding modules and how these programs may be altered during melanoma progression.

12 Aishwarya Iyer Medicine & Dentistry Medicine TRANSLATING GENOMIC INTRATUMOR HETEROGENEITY INTO PROGNOSTIC BIOMARKERS IN T-CELL LYMPHOMA Aishwarya Iyer, Weiwei Wong, Dr. Gane Wong, Dr. Robert Gniadecki, Mycosis fungoides (MF) is a common extranodal T-cell lymphoma primarily arising in the skin. In early disease stages, MF presents as skin patches and plaques that in some cases may progress to tumor and disperse to lymph nodes and other internal organs. Early diagnosis is difficult as the histology overlaps with features of inflammatory skin diseases. Even when the diagnosis is established there are no prognostic markers that predict whether the disease will be aggressive or indolent. Lastly, there are no curative treatments and MF will invariably relapse even after aggressive chemotherapy. The disease is a diagnostic, prognostic and therapeutic challenge. The main objective of this study is to address the question of tumor heterogeneity in MF. To date, MF is considered to be monoclonal, derived from a transformed, mature memory T- cell. However, clinical observations and preliminary data suggest that MF comprises multiple subclones, which may be of importance for understanding of disease evolution and resistance to therapy. We plan to address this objective using Whole Exome Sequencing (WES) of MF tissue prepared by laser microdissection (LMD). Following are our study objectives: Description of mutational landscape of early disease (plaques) and late stages (tumors). The emphasis of this objective is to find genetic differences between early and late stage disease and to identify key signaling pathways likely to be affected by these mutations. Deciphering the genetic evolution of MF.

Patients with MF usually develop multiple lesions and it is not clear whether advanced lesions (tumors) develop by evolution from plaques or rather emerge from lymphoma precursor cells. We will compare mutational landscapes between plaques and a tumors sampled from the same patient to describe the developmental tree of the tumor. Achieving these objectives will assist in the precision diagnosis and prognostication of the disease.

13 John Githaka Medicine & Dentistry Biochemistry Bad phosphorylation regulates mammary gland morphogenesis John Maringa Githaka, Namita Tripathi, Raven Kirschenman, Namrata Patel, Rachel Montpetit, Nika Danial, Ing Swie Goping During the post-natal development of the mouse, the mammary gland progresses from a microscopic anlage to a visible ductal network with the capacity to differentiate into a milk- producing organ. This process is dependent on tightly controlled cycles of proliferation, migration, differentiation and apoptosis. These signaling pathways driving normal organ development are often hijacked during tumor development. BH3-only protein Bad is a prognostic marker for survival of breast cancer patients. However, whether Bad has a physiological role in the mammary gland is not known. Bad is extensively regulated through reversible phosphorylation of three key serine (S) residues 75, 99 and 118 (S112, 136 and 155 in the mouse). Using a mouse model wherein the three S residues are replaced by alanine (3SA), we observed a delay in ductal elongation and decreased primary branching of the mammary epithelial tree during puberty. Transplant experiments between wild-type (WT) and 3SA mice demonstrated a requirement for Bad phosphorylation in both the epithelial and stromal compartments of the mammary gland. 3D culture of purified mammary epithelial organoids phenocopied the in-vivo observations, confirming the cell autonomous- effect of Bad phosphorylation. Furthermore, Bad 3SA mutants reconstituted in the

untransformed human mammary epithelial cell line, MCF-10A, had significantly delayed tubulogenesis compared to wild type Bad, reminiscent of the observed in-vivo mammary gland whole mounts and ex-vivo 3D organoid phenotypes. Taken all together, these findings provide a framework with which to query both normal mammary gland development and cancer pathophysiology.

14 Asmaa Basonbul Medicine and Dentistry Medicine BCL-2 Inhibitor Venetoclax Enhances Temozolomide Sensitivity in AML Asmaa Basonbul Introduction: Temozolomide (TMZ) is an alkylating agent, which adds a methyl group to O6 position of guanine, resulting in mismatch pairing with double strand breaks leading to apoptosis. The DNA repair enzyme O6-methylguanine methyltransferase (MGMT) enhances tumor cell resistance to TMZ. BCL-2 is an anti-apoptotic protein preventing cell to death. Venetoclax (Vent) is a small molecule, which promotes cell apoptosis through inhibition of BCL-2 protein. Objective: To evaluate the ability of the BCL-2 inhibitor Vent to enhance TMZ sensitivity in acute myeloid leukemia (AML) cells. Methods: Western blot and ELISA were used to measure MGMT and BCL-2 expression in KG1, MV4-11 and MOLM13 AML cell lines. Cell lines were incubated with TMZ 5, 10, 15, 20, 50 and 100 uM in combination with a fixed concentration of Vent (1 uM for KG1 and 2.5 nM for other cell lines). After 3 days, PrestoBlue cell viability assay was performed using spectrophotometry at 560 ex and 590 em. Synergy was evaluated by the Chou-Talalay method. Results: MGMT was expressed only in KG1 and was absent in MV4-11 and MOLM13. BCL-2 was highly

expressed in MV4-11, with lower expression in the other cell lines. KG1 demonstrated strong resistance to TMZ; however, co-incubation with 1 uM Vent resulted in a marked enhancement of sensitivity to TMZ. Vent 2.5 nM alone inhibited cell growth by approx. 50% in MV4-11 and MOLM13. This dose in combination with TMZ markedly increased the cytotoxicity with nearly 100% inhibition at 100uM TMZ. Synergistic effect was demonstrated in all cell lines with combination index (CI) < 1. Conclusion: Venetoclax enhances TMZ sensitivity in all cell lines and induces cytotoxicity, including MGMT overexpressing cells. Future direction: Evaluate the drug combination design on AML patient cells, followed by testing in an animal model. Moreover, explore the relationship mechanism between the two inhibitors.

15 Vrajesh Pandya Medicine and Dentistry Biochemistry Bik mediated genomic instability as a potential mechanism underlying the poor prognosis in Bik-high breast cancers Mr. Vrajesh Pandya, Raven Kirschenman, Namrata Patel, Darryl Glubrecht, Dr. Sambasivarao Damaraju, Dr. John Mackey, Dr. Judith Hugh, Dr. Ing Swie Goping Biomarkers give information on disease progression and important biological pathways. We identified that the pro-apoptotic Bcl-2 interacting killer (Bik) was associated with poor breast cancer prognosis. We postulated that Bik contributes to poor outcomes through a nonapoptotic mechanism. To test this, we created Dox-inducible breast cancer cell lines expressing titratable amounts of Bik. Bik-expression altered cell line growth characteristics. This effect was dependent on Bik, was heritable and did not require persistent Bik expression. Specifically, transient Bik expression generated cells with increased anchorage- independent growth and clonogenic potential. This aggressive phenotype was independent

of anti-apoptotic Bcl-2, Bcl-xL, MCL-1 and GRP78, supporting the involvement of a novel mechanism. Since genomic instability drives cancer evolution, we tested the effect of Bik on DNA damage. Bik induced DNA damage as measured by comet assay and the DNA-damage marker γH2AX. This genomic instability was dependent on the Bik BH3-domain, caspase activation and caspase activated DNase (CAD). Of clinical interest, anti-estrogens that are the standard of care for ER positive tumors increased Bik levels and induced γH2AX levels in cell culture, suggesting a negative effect of therapy. When we analyzed Bik protein levels in a cohort of breast cancer patients, ER positive- anti-estrogen treated patients with high Bik levels showed significantly reduced survival compared to ER negative patients. Thus Bik may contribute to poor outcomes of breast cancer patients by inducing CAD activity, genomic damage and development of aggressive tumors. Further work to understand the interactions between Bik, DNA instability and clinical anti-estrogen treatments are warranted. 16 Subha Kalyaanamoorthy Pharmacy and Pharmaceutical Sciences Pharmacy Binding site and interaction analysis of Kv7 modulators on KCNQ1 and KCNQ3 channels Subha Kalyaanamoorthy and Khaled H Barakat Ion channel proteins and their modulators are being recognized to play a significant role in several diseases, including cancer. Kv7 is a voltage-gated potassium channel family that comprises of five subtypes, KCNQ1 to KCNQ5. Suppression of different Kv7 (KCNQ/M) voltage-gated potassium channels has been reported in various cancers and their physical effects. For example, suppression of KCNQ1 was found in gastrointestinal cancer[1], while low-level expression of KCNQ2 and KCNQ3 were identified to be the cause of bone cancer pain in animal models[2]. Therefore, small molecule modulators that activate these channels have been of interests for therapeutic applications. Diverse classes of channel activators have been identified, but the knowledge on their sites of action is still limited[3]. A clinically tested drug, retigabine (RTG) is known to activate all Kv7 channels from KCNQ2 to KCNQ5 except KCNQ1. On the other hand, Zinc pyrithione (ZnPy) another Kv7 activator, activates all KCNQ

channels except KCNQ3. The knowledge of molecular determinants and isoform-specific potency of RTG and ZnPy would help in the development of new structures with isoform specificity[3]. Therefore, we generated three-dimensional structural models of KCNQ1 and KCNQ3 channels using molecular modelling methods. Potential binding sites were predicted from these models, and the interaction of RTG and ZnPy in these sites were analyzed using molecular docking and binding free energy methods.

References:

1. Than, B.L.N et al., (2014). The role of KCNQ1 in mouse and human gastrointestinal cancers, Oncogene, 33, 3861-3868. 2. Cai, J., et al., (2015). Suppression of KCNQ/M (Kv7) potassium channels in the spinal cord contributes to the sensitization of dorsal horn WDR neurons and pain hypersensitivity in a rat model of bone cancer pain. Oncology Reports, 33, 1540-1550. 3. Xiong Q et al., (2015). Combinatorial augmentation of voltage-gated KCNQ potassium channels by chemical openers. PNAS, 195(8), 3128-3133

17 Amira Fitieh Faculty of Medicine and Dentistry Oncology-Division of Exp. Oncology BMI-1 Promotes DNA End Resection through CtIP Ubiquitylation Amira Fitieh 1, Lubna Yasmin1, and Ismail Ismail1 (1) Department of Oncology, University of Alberta, Edmonton, Alberta, Canada. The BMI-1–containing Polycomb Repressive Complex 1, PRC1, plays an important role in aberrant cancer gene silencing and is a clinical target for cancer therapy. BMI-1, a major component of PRC1, is an oncogene that is notably upregulated in a variety of cancer types, including lymphomas, prostate cancer, non-small cell lung cancer, colon cancer, breast cancer, and nasopharyngeal carcinoma.

In recent years, BMI-1 has been known to be involved in DNA double-strand break (DSB) repair, yet, to date its downstream mechanism of action is still unclear and needs to be elucidated. Here, we have found that BMI-1 is required for the Homologous Recombination (HR)-mediated repair of DNA DSBs. More specifically, results from our study have shown that BMI-1 acts at the DNA end-resection level, which is an essential step for the early events of the HR repair pathway. We have demonstrated that BMI-1 regulates end resection through its interaction with CtIP, a key DNA endonuclease controlling DNA end resection. Co- immunoprecipitation experiments revealed that BMI-1 interacts with CtIP, as well as, mediates its ubiquitylation. Additionally, pharmacological inhibition of BMI-1 by the small molecule inhibitor, PTC-209, drastically reduced the accumulation of DNA damage-induced foci for key players of the HR repair pathway, including RAD51, RPA, and CtIP. In summary, BMI-1–mediated ubiquitylation of CtIP is required for the key step of DNA end resection that is essential in the HR pathway of DNA repair. This unravels a novel role for BMI-1 in the HR repair pathway of DSBs, which could be further exploited as a potential target and prognostic marker for better cancer therapeutics.

18 Xia Xu Medicine & Dentistry Oncology Brain Fatty Acid-binding Protein and Polyunsaturated Fatty Acids: Effect on Phospholipid Bilayer of Malignant Glioma Cell Membrane Xia Xu, Yixiong Wang, Xuejun Sun, Roseline Godbout Malignant glioma (MG) is the most common primary malignancy in brain. Brain fatty acid binding protein (FABP7) is associated with increased MG cell migration and a poor clinical prognosis. Long chain polyunsaturated fatty acids such as omega-3 docosahexaenoic acid (DHA) and omega-6 arachidonic acid (AA) are abundant in brain cell membranes. When long chain polyunsaturated fatty acids are incorporated into cell membranes, this significantly alters membrane properties, particularly as related to order and fluidity.

We used the fluorescent dye Laurdan combined with quantitative fluorescence microscopy to study the effect of DHA on dipolar relaxation of malignant glioma membranes. Dipolar relaxation is a measurement of membrane order and fluidity. In particular, we were interested in determining the effect of FABP7 on membranes of cells cultured in medium supplemented with AA or DHA. We have tested four MG cell lines to date: FABP7-positive U251 and M049, and FABP7-negative T98 and A172. Cells were treated with bovine serum albumin (BSA) vehicle, 60 μM AA or 60 μM DHA for 24 hours. Raw images of liquid- disordered and liquid-ordered phases were processed with ImageJ. Ratiometric measurement of the fluorescence intensity in two spectral channels were recorded as generalized polarization (GP) value. Our preliminary data indicate increased membrane disorder, suggesting increased fluidity, in FABP7-positive cell lines (U251 and M049) with 60 μM DHA treatment. However, FABP7-negative cell lines (T98 and A172) did not show this GP value shift. FABP7/DHA-dependent alteration of membrane fluidity could underlie previously observed FABP7/DHA-dependent inhibition of migration. 19 Mackenzie Coatham Medicine and Dentistry Obstetrics and Gynecology Utilizing Cell Line Models with Chromatin Remodelling Protein Deficiencies to Study Aggressive Dedifferentiated Endometrial Cancer Mackenzie Coatham, Cheng-Han Lee, Lynne-Marie Postovit An estimated 6600 Canadian women were diagnosed with uterine cancer in 2016, with 20% of women succumbing to the disease. One of the most lethal subsets of uterine cancer is dedifferentiated endometrial carcinoma (DDEC). DDEC tumors possess both well- differentiated and undifferentiated regions. The majority of metastatic disease is made up of cells from the undifferentiated component of DDEC yet it is unclear how these poorly differentiated regions are initiated and sustained within the carcinoma.

Examining the well-differentiated and undifferentiated components of DDEC tumors in terms

of mutation profiles, we demonstrated that 80% of the undifferentiated regions in DDEC lesions lack the expression of core chromatin remodeling proteins, BRG1 or ARID1A and ARID1B. We hypothesize that the loss of these proteins, which are known regulators of transcription, may lead to the induction and/or maintenance of stem cell-like gene expression programs that drive dedifferentiation, metastasis and therapy resistance.

21 Indrani Dutta Medicine and Dentistry Experimental Oncology Can Gamma Delta T cells Target Breast Cancer Stem Cells? ndrani Dutta, Lynne-Marie Postovit and Gabrielle M. Siegers Gamma Delta T cells (GDTc) comprise 2-5% of circulating lymphocytes in humans. Immunosurveillance cells garnering great interest for their anti-tumoral activity, GDTc recognize antigens directly without antigen presentation, enabling rapid response. GDTc respond to self-molecules signaling cellular stress; recognition maybe mediated by the natural killer receptor NKG2D, whose ligands are upregulated on transformed cells. Tumour lysis is mediated in part by FasL and TRAIL released by GDTc that bind target cells bearing Fas and TRAIL receptors inducing apoptosis. Clinical trials have confirmed the safety of GDTc in cancer immunotherapy. However, few studies have investigated the ability of GDTc to target cancer stem cells (CSC), a small population responsible for resistance to cancer therapies, leading to cancer recurrence. We are investigating whether GDTc can target breast cancer CSC (BCSC). In breast cancer, the CD24lowCD44high signature is considered one identifying feature of BCSC. We sort SUM 149 cells into BCSC (CD24low) and non-BCSC (CD24high) targets. CD24low cells are 10-20% less susceptible to GDTc killing than CD24high targets. MCF-7 and SUM149 mammospheres, which are enriched for CD24low cells, are also 20-40% less susceptible than adherent targets. CD24low SUM149 fractions and SUM149 mammospheres have two to three-fold lower expression of NKG2D ligands compared to CD24high and adherent SUM149 respectively, suggesting that this may be one mechanism whereby these cells evade detection and subsequent killing by GDTc. Serial dilution and mammosphere assays have shown that the target cells resistant to GDTc targeting are enriched for BCSCs and have higher sphere forming ability. We find that the Fas-FasL pathway does not perform its function in CD24lowCD44high cells and mammospheres when targeted by GDTc. We are now validating these results with blocking and loss-of-function assays. Determining how CSCs

evade GDTc killing and devising strategies to overcome this resistance will improve GDTc immunotherapy for breast cancer.

22 Francisca Cristi Munoz Medicine & Dentistry Medical Microbiology & Immunology Combination of mutations to produce a reovirus variant with improved oncolytic potency Cristi-Munoz Francisca, Shmulevitz Maya*, Hitt Mary* (*:co-supervisors) Reovirus is a nonpathogenic virus that naturally inhabits the enteric tract of humans. Reovirus can also selectively infect and replicate in tumor cells, and is therefore a candidate for cancer therapy undergoing clinical trials. Given that wild-type reovirus is naturally adapted to enteric environments (rather than tumors), our laboratory used positive selection to isolate 13 reovirus variants that replicate more efficiently in tumor cells relative to wild- type reovirus (T3wt). Since these 13 reovirus variants have 1-4 mutations each, we did not know which mutations were important or dispensable. To identify the mutations critical for enhanced oncolysis, I introduced 20 of the mutations into the reovirus reverse genetics plasmids using site-directed mutagenesis. Then, I generated viruses with either single mutations, or various combinations of the 1-4 mutations found in the original variants. Using plaque size to reflect the proficiency of reovirus replication in tumor cells, I found that one mutation in each variant could fully restore the enhanced oncolytic potential. The oncolysis- promoting mutations predominated in reovirus structural protein lambda2 and the sigma1 cell attachment protein. Concurrently I began to establish a mouse xenograft model of breast cancer in our lab to compare the reovirus variants in their ability to reduce tumor growth and increase animal survival. I have determined the best reovirus dose and the time frame to observe these effects. As expected, T3wt reduced tumor growth and improved survival. In future animal experiments, I will compare the oncolytic activity of reovirus variants to identify the most-oncolytic reovirus platform. My experiments will provide a better understanding of reovirus biology and how to improve reovirus as a cancer therapeutic.

23 Quinn Storozynsky Medicine and Dentistry Oncology Combining an Oncolytic Vaccinia Virus with Image-Guided Radiotherapy for Treatment of Malignant Glioma Quinn Storozynsky, David Evans, Kyle Potts, Kate Agopsowicz, Roseline Godbout, and Mary Hitt Malignant gliomas (MG) are highly invasive and aggressive brain tumors. Despite the current standard of care and decades of research the prognosis for patients with MG is abysmal– with five-year survival rates as low as 5.5%. Novel more effective treatment options are desperately needed to combat this aggressive disease. Oncolytic virus (OV) therapy is an advancing treatment option that harnesses tumor-selective viruses to kill cancer cells while also jarring the host immune system into action– further wiping out cancer cells by facilitating a systemic anti-tumor immune response. However, OV’s alone are unlikely to clear most tumors completely, but could enhance other cancer therapies. Image-guided radiotherapy (IGRT) uses computer-modulated imaging techniques to precisely deliver ionizing radiation to treat cancer. Despite the precision IGRT offers, tumor microextensions and radioresistant cell populations such as glioma stem cells can still be ‘missed’ and may contribute to recurrence or progression. Many studies have noted synergistic effects when OV’s are combined with radiotherapy to treat cancer in preclinical models. Here we propose to combine our mCherry-tagged mutant vaccinia virus (∆F4L∆J2RmCh), which is tumor- selective due to mutations in viral nucleotide biosynthesis genes, with IGRT executed using state-of-the-art Small Animal Radiation Research Platform (SARRP) technology. We hypothesize that combining ∆F4L∆J2RmCh with IGRT will produce better tumor control by generating additive or synergistic effects, in which IGRT destroys the majority of the tumor mass while our OV seeks out and targets any remaining cancer cells that have been missed or are resistant to radiotherapy.

25 Clayton Bell Medicine & Dentistry Oncology CR42-24, a Novel Colchicine Derivative, as a therapy for Bladder Cancer Clayton Bell, Kyle Potts, Mary Hitt, John Lewis, and Jack Tuszynski Colchicine is an anti-mitotic drug that targets microtubules, and blocks microtubule dynamics. Colchicine is primarily used to treat gout but has been examined in many clinical trials to treat conditions such as prostate cancer, Behcet’s disease, and many others. However, due it’s narrow therapeutic window colchicine has had limited clinical translation. A possible method of overcoming the toxicity is to produce a derivative that has increased cancer specificity, and therefore reduced toxicity. Previously our lab identified a novel colchicine derivative (CR42-24) that exhibits increased cancer-killing capabilities and decreased toxicity compared to colchicine. This was accomplished by designing CR42-24 to have increased affinity for βIII tubulin. βIII tubulin is a β-tubulin isotype that is incorporated into tubulin dimers that make up microtubules. βIII is an excellent target for novel therapies as it has low expression in healthy tissue, is overexpressed in metastatic cancers, and is a clinical marker of poor prognosis. Using cell line screening we demonstrate that CR42-24 is highly toxic to a variety of cancer types with effective killing doses at nanomolar concentrations. More specifically CR42-24 is shown to be highly effective against bladder cancer. Current chemotherapy for bladder cancer is a combination of Gemcitbaine and Cisplatin (Gem/Cis). Although marginally successful many patients develop resistance to Gem/Cis leaving them with limited options for a second line therapy, thus development of alternative therapies is highly desired. Using in-vitro methods we demonstrate that CR42-24

kills aggressive bladder cancer cell lines, and is more effective than Gemcitabine and Cisplatin. Additionally, CR42-24 doubles survival of mice with bladder cancer xenografts. Finally, we show that CR42-24 is highly synergistic with other chemotherapies, thereby increasing its therapeutic potential. Through our studies we have show that CR42-24 is effective in treating aggressive bladder cancer and thus may serve as an effective alternative therapy.

26 Andrew Locke Medicine & Dentistry Oncology CtIP SUMOylation Mediates its Function in DNA End Resection and Homologous Recombination Repair Andrew Locke and Ismail Hassan Ismail DNA double-strand breaks (DSBs) are among the most lethal lesions in the genome if they are not repaired or if their repair is executed incorrectly. They are mainly repaired by non- homologous end joining (NHEJ) and homologous recombination (HR); proper choice of the repair pathway is critical for genome stability. Initiation of DNA end resection by the CtIP/Mre11-Rad50-Nbs1 complex generates single-stranded DNA overhangs that are refractory to canonical NHEJ. This decision to resect is fundamental to DSB repair pathway choice, as end resection is necessary for homology searching, which is central to HR. To coordinate the appropriate and timely execution of end resection, CtIP’s function is tightly controlled by multiple post-translational modifications, including phosphorylation, acetylation, and ubiquitylation. Analysis of CtIP’s primary sequence reveals multiple potential SUMOylation sites, but little is known about which of the sites are SUMOylated in vivo, by which SUMO ligase, and the biological relevance of these SUMOylation events. We have found that CtIP is SUMOylated with SUMO1 and SUMO2, and mutational analysis has identified lysine 896 (K896) as CtIP’s main SUMOylation site. SUMOylation of CtIP was attenuated by short interfering RNA-mediated depletion of the SUMO ligase PIAS1 (protein

inhibitor of activated STAT1), while co-immunoprecipitation experiments revealed DNA damage stimulates an interaction between CtIP and PIAS1. Consistent with PIAS1 being involved in CtIP SUMOylation, depletion of PIAS1 reduced the frequency of HR in a cell-based gene conversion assay. Taken together, our findings reveal a role for SUMOylation in the regulation of HR, particularly at the level of CtIP-mediated end resection.

27 Aysha Ansari Engineering Chemical & Materials Engineering Customizing Lipopolymers for Efficient siRNA Delivery to Different Leukemia Cells Aysha Ansari, Remant Bahadur KC, Cezary Kucharski, Hasan Uludağ Purpose: Chemical functionalization of low molecular weight (MW) polyethylenimine (PEI) with aliphatic lipids enhances their siRNA transfer efficiency in leukemia cells owing to increased cell membrane interaction facilitated by the hydrophobic lipid substituents. However, the response of different leukemia cell lines and primary patient cells varies depending not only on biological differences but also on several factors related to the chemistry of the lipid-substituted PEI lipopolymers. The aim of this study is to analyze the cell-type specific observations in the context of differential chemistries of the lipopolymers. Methods: Caprylic acid (CA), palmitic acid (PA), and linoleic acid (LA) substituted polymers were synthesized by N-acylation of low MW PEI with lipid chlorides. THP-1, KG-1, KG-1a, HL- 60, and K562 cell lines were utilized for cell studies. Primary cells were obtained from the peripheral blood or bone marrow of AML patients. Gene silencing and cellular uptake of particles was performed by flow cytometry. Results: Modification with short carbon chain CA yields higher substitution compared to long chain LA at a given lipid:PEI feed ratio. LA lipopolymers resulted in higher siRNA complex uptake across different AML cell lines, K562 CML cell line, and primary AML patient cells. While LA lipopolymer showed higher gene silencing activity than CA lipopolymer in the least differentiated KG-1 AML cell line, the latter was more effective in the more differentiated

THP-1 cell line. In contrast, PA lipopolymer exhibited higher silencing efficiency in K562 cells than LA lipopolymer. In primary AML patient cells, CA lipopolymer exhibited relatively more significant silencing activity than LA lipopolymer. Conclusion: Lipid structure and composition, amount of lipid substitution, and MW of PEI backbone play a critical role in dictating the siRNA complex uptake and subsequent gene silencing efficiency of these lipopolymers in different types of leukemia cells, including primary patient cells.

28 Brennen Dobberthien Medicine & Dentistry Oncology Detecting Glutamate, Glutamine, and GABA Simultaneously at 9.4 T with Improved Magnetic Resonance Spectroscopy PRESS Sequence Timings Brennen J. Dobberthien, Anthony G. Tessier, Atiyah Yahya Glutamate (Glu), glutamine (Gln), and γ-aminobutyric acid (GABA) are brain metabolites whose levels are relevant to the study of cancer. Proton Magnetic Resonance Spectroscopy (MRS) is a non-invasive technique that can be used to quantify these levels. The high field strength of 9.4 T offers increased spectral resolution; however, there is spectral overlap among the signals of Glu (≈2.35 ppm), Gln (≈2.45 ppm), GABA (≈2.28 ppm), and N- acetylaspartate (NAA; ≈2.49 ppm). By optimizing the echo times (TE1 and TE2) of Point RESolved Spectroscopy (PRESS), a commonly employed technique in in-vivo MRS, the J- coupling interactions of these metabolites can be exploited to minimize the NAA signal while retaining and resolving the signals of Glu, Gln, and GABA, so that all three can be detected simultaneously.

The spectra of Glu, Gln, GABA, and NAA were simulated, and an objective function was created by subtracting the normalized peak area of NAA from that of Gln. A contour plot was created based on this function, and the PRESS {TE1, TE2} combination that maximized the

objective function while retaining sufficient peak areas for Glu and GABA was considered optimal. This timing set was verified with a 9.4 T animal MRI scanner on phantom solutions and in vivo on five Sprague Dawley rat brains. In-vivo spectra were analyzed with LCModel.

An optimal {TE1, TE2} combination was determined to be {106 ms, 16 ms}, which resulted in an objective function value of 0.65. For individual phantom solutions of Glu, Gln, GABA, and NAA, the optimal TE gave peak areas of 0.59, 0.44, 0.70, and ≤0.05, respectively, relative to the corresponding short-TE values. Spectra from rat brain showed individual, well resolved peaks for Glu, Gln, and GABA. Reported LCModel Cramér-Rao Lower Bound (CRLB) values for all rats were ≤6%, ≤17%, and ≤19% for Glu, Gln, and GABA, respectively.

We have reported on a VACV with deletions in F4L (ribonucleotide reductase small subunit)

and J2R (thymidine kinase) that exhibits oncolytic properties and promotes anti-tumor immunity with superior safety in BCa. In a screen of BCa cells, we found significant differences in BCG internalization while, delF4LdelJ2R VACV could replicate in and killed both BCG susceptible and resistant cell lines to similar levels. Comparison of delF4LdelJ2R VACV and BCG in the orthotopic AY-27 immunocompetent rat model shows that the virus induced superior tumor clearance and long term survival. Pretreating cells with a Pak1 inhibitor, or inducing BCG-resistance with Pak1 shRNA, showed decreased BCG uptake with little effect on VACV replication. Finally, we have developed an AY-27-shPak1 cell line that is resistant to BCG uptake that we will use to generate an BCG-resistant BCa model.

Given the high rate of recurrence and the lack of treatment options for BCG-resistant BCa, our delF4LdelJ2R VACV could provide a safe and urgently needed therapy.

scintillating nanoparticles that convert X-rays to visible light to activate PDT. Currently, there is no data on how radioPDT is affected by tumor hypoxia, which is relevant because both radiotherapy and PDT are oxygen consumptive processes. Therefore, our objective is to characterize radioPDT in hypoxia, and its impact on singlet oxygen yield and cancer cell death.

Results: Preliminary data demonstrates radioPDT effectively augments radiotherapy in hypoxia via singlet oxygen production. In vitro viability assays shows minimal intrinsic toxicity in prostate cancer and fibroblast cell lines when not activated, and significant increase in cytotoxicity when activated. CT and MRI studies show appreciable signal generation, and CAM model studies show preferential tumor-targeting capabilities. Future studies are aimed at exploring their diagnostic and therapeutic potential in vivo in tumor-bearing mice models.

Conclusion: radioPDT may offer a novel method of augmenting traditional radiotherapy and minimizing toxicity.

31 Francesco Gentile Science Physics Rational design of inhibitors of the XPF-ERCC1 heterodimerization for combination cancer therapy Francesco Gentile1, Ahmed H. Elmenoufy2, Feridoun Karimi-Busheri3, Olivier M. Soueidan2, David Jay3, Rajam S. Mani3, Xiaoyan Yang3, Khaled H. Barakat4, Jack A. Tuszynski1,3, Michael Weinfeld3 and Frederick G. West2 1Department of Physics, University of Alberta, Edmonton, Alberta, Canada 2Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada 3Department of Oncology, University of Alberta, Edmonton, Alberta, Canada 4Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, AB, Canada The ERCC1-XPF heterodimer is a 5ʹ-3ʹ structure-specific endonuclease which is essential for nucleotide excision repair (NER) and inter-strand crosslink (ICL) DNA repair pathways. Although its action is critical to maintain genome integrity and stability and prevent against damage-induced mutations, it can counteract the effect of DNA damaging therapies such as platinum-based chemotherapy and radiotherapy. A promising approach to enhance the effect of such therapies is to inhibit the action of XPF-ERCC1 in cancer cells using small molecules. In this study, we used a computational drug design workflow to rationally design novel analogues of the F06 molecule, a lead inhibitor which targets the dimerization between XPF and ERCC1, required for the functional endonuclease. We synthesized seven compounds, which show in silico show interesting binding affinities and ligand efficiency towards an XPF site on the dimerization interface. This approach yielded compound 3 and 4 as potent inhibitors of ERCC1-XPF activity. An iIn vitro ERCC1-XPF endonuclease assay identified compound 4 as the best ERCC1-XPF inhibitor with an IC50 value of 0.3 µM. Also, the Kd value of this compound was measured as ~100 nM with by fluorescence spectroscopy. Compound 4 also showed a significant inhibition of the removal of cyclobutane pyrimidine dimers (CPDs) compared with control cells after exposure of HCT 116 cells to UV radiation. Compound 4 can

potentially be used in combination with existing DNA-damaging therapies to amplify their effect on cancer cells.

32 Michael Chu Medicine Oncology Detection of Minimal Residual Disease in Autograft is Prognostic of Survival Following Stem Cell Transplant in Multiple Myeloma Michael P. Chu, Eva Baigorri, Jitra Kriangkum, Joanne Hewitt, Andrew R. Belch, Christopher P. Venner, Irwindeep Sandhu, Linda M. Pilarski Introduction In multiple myeloma (MM) patients capable of handling aggressive treatment, high dose chemotherapy (HDT) and autologous stem cell transplant (ASCT) are used in first remission to improve progression free (PFS) and overall survival (OS). MM still recurs following HDT/ASCT partially due contaminated autografts. This study hypothesized that a threshold in the autograft exists to predict risk of earlier relapse.

Methods MM patients in this single institution study were prospectively observed following initial diagnosis from 1998 to 2014. Patient demographics and myeloma details were collected. Patients were observed for time to progression following HDT/ASCT. At diagnosis, a bone marrow aspirate aliquot was used to create patient-specific primers for their variable region sequence (VDJ) and used during quantitative PCR on each patient’s autograft at time of collection and reinfusion (i.e. post thawing). Cut-point analysis was used to determine a threshold for which detection of minimal residual disease in the autograft may translate into poorer survival. Cox proportional hazards multivariate analysis took into consideration collected co-variables.

Results Between 1998 and 2014, 89 patients were accrued to study. 77 patients are included here for analysis. Basic demographics included median age at diagnosis of 58 years, 61 male, and a majority of patients having ISS stage 3. Induction therapy was highly variable. A threshold of detection at ≥ 1 in 100,000 circulating myeloma cells in the autograft was associated with poorer PFS following HDT/ASCT (median 1.7 vs 2.6 years, hazard ratio [HR] 1.78, p=0.03). A trend to poorer OS was also seen in this group (median 5.6 vs. 15.8 years, HR 2.65, p=0.08). This detriment remained significant when taking into consideration age, sex and ISS staging.

Conclusions This is the first study of which we are aware that demonstrates an association between the degree of autograft contamination with MM cells and survival.

33 Abdulsalam Alharbi Pharmacy and Pharmaceutical science Pharmaceutical science Development of Novel Polymeric Micellar DACHPt for Enhanced Platinum Based Chemotherapy in Colorectal Cancer Abdulsalam Alharbi, Mohammed Vakili and Afsaneh Lavasanifar Development of Novel Polymeric Micellar DACHPt for Enhanced Platinum Based Chemotherapy in Colorectal Cancer

Abdulsalam Alharbi, Mohammed Vakili and Afsaneh Lavasanifar. Faculty of Pharmacy and Pharmaceutical Science, University of Alberta, Edmonton, Alberta, Canada

Purpose: The parent compound of Oxaliplatin, dichloro(1,2-diaminocyclohexane)platinum(II) (DACHPt) is a potent chemotherapeutic agent with wide spectrum of anticancer activity, lower side effects and no cross-resistance with cisplatin in many cisplatin-resistant cancers.

The objective of this project is to develop polymeric micelles of DACHPt with potential for enhanced platinum based chemotherapy of colorectal cancer. Method: Poly(ethylene oxide)-b-poly-(α-carboxylate-€-caprolactone) (PEO-b-PCCL) diblock copolymer was synthesized. Then, DACHPt was reacted with PEO-b-PCCL to form polymer- metal complexes. The complex was dialyzed in water to prepare DACHPt loaded micelles. Prepared polymeric micelles were then characterized for their average diameter by dynamic light scattering, level of complexed DACHPt by ICP-MS and in vitro release of Pt compared to free drug by ICP-MS. Cytotoxicity of DACHpt micelles against human colorectal cancer cell lines, HCT-116 and SW-620 was evaluated following 24-72 h incubation and compared to free drug cytotoxicity by MTT assay. Results: Drug loading in polymeric micelles reached high levels of 50.2% w/w on average. Polymeric micellar complexes of DACHPt were found to have a mean diameter of 56 nm. The release of Pt from micelles was sustained. Only 53.6% of Pt was released from micelles after 120 h compared to free drug that released 96.5 % of Pt after 7.5 h. The IC50 of polymeric micellar DACHPt was higher than that of free drug in HCT116 cells in all incubation times (12.86 to 288.7 ug/ml for micelles versus 1.9 to 92.4 ug/ml for free drug at 24-72h incubation). In Sw-620 cells an opposite trend was observed (15.11 to 36.4 ug/ml for micelles versus 94.7 to 120.4 ug/ml for free drug at 24-72 h incubation). Conclusion: The results point to a great potential for PEO-b-PCCL/DACHPt polyplex micelles for enhanced Pt based chemotherapy in colorectal cancer. Acknowledgement: AA was funded by Umm Al-Qura University, Mecca, Saudi Arabia. This research was supported by grants from NSERC.

34 Md Touhidul (Apu) Islam Medicine & Dentistry Biochemistry Development of systems for histone expression and purification Md Touhidul Islam, Rashmi Panigrahi, Morenike Ajidagba, Mark Glover The eukaryotic genome is compacted in the cell forming nucleoprotein complexes known as nucleosomes. Each nucleosome is an octameric core composed of four histone subunits (H2A, H2B, H3 and H4) wrapped with 146 bp of DNA. This DNA packaging is crucial for the regulation of gene expression and the mending of damaged DNA. Upon DNA damage, phosphorylation of H2Ax (H2A variant) provides a signal for the recruitment of other DNA damage response (DDR) proteins adjacent to the DNA lesions.

Previously, histones have been extracted from the cell lysate using urea denaturant with subsequent extensive dialysis for refolding the heteromeric histone complexes. This method is time and cost intensive. In the Glover lab, we have established a rapid purification protocol using the co-expression system for the human nucleosome. Additionally, we have developed a system for the in vitro phosphorylation of H2AX that will facilitate the structural analysis of Gamma-H2AX nucleosomes.

35 Shelagh Campbell Science Biological Sciences Developmental regulation of the cell cycle by Wee1 and Myt1 kinases S. D. Campbell Precisely timed progression through the cell cycle is essential for producing appropriate numbers of properly specified cell types during animal development. Many different types of cell cycles exist, serving specialized developmental functions. For example, many early embryos undergo simplified cleavage cycles consisting of rapid oscillations between S and M phases of the cycle by dispensing of gap phases. In spite of this diversity, regulation of Cdk1 activity is used to coordinate mitosis with other stages of the cell cycle in all eukaryotic cells. Defects in Cdk1 regulation are also frequently associated with cancer, consequently new therapies targeting these conserved Cdk1 mechanisms are being actively sought.

During interphase, Cdk1 activity is inhibited by two related inhibitory kinases: Wee1 and Myt1. In my laboratory we are studying the developmental functions of these partially redundant Cdk1 inhibitory kinases in Drosophila. For this purpose we have created null wee1 and myt1 mutants and characterized unique developmental functions for each of these Cdk1 inhibitors. Drosophila Wee1 mutants are early embryonic lethal due to specific S/M checkpoint defects that result in mitotic catastrophe. Wee1 inhibitors being developed for cancer therapy may trigger similar effects. In contrast, Myt1 mutants exhibit a completely different spectrum of cellular defects, affecting stem cell proliferation and sensory organ development as well as male meiosis. Insights gained from studies of these mutants may thus be relevant to understanding the recent proposal that human PKMyt1 may be important for the development of glioblastoma. Our studies of these genetically tractable animal models therefore provide opportunities for learning more about how these specialized cell cycle regulators function in physiologically meaningful settings. Such information could help us to

better understand how manipulating Cdk1 inhibitory phosphorylation could be used to help treat cancer without compromising patient viability.

36 Mikaela Nevin Medicine and Dentistry Medical Genetics DLX2 gain-of- function in murine brainstem glioma cells restores histone H3K27 trimethylation and reduces expression of the H3K27M mutant isoform Authors and Affiliations: Mikaela Nevin 1,4 , Bridget Mulvany-Robbins 4 , Xiaohua Song 1,4 , Roseline Godbout 2,4 , David D. Eisenstat 1,2,3,4 . 1 Department of Medical Genetics, 2 Department of Oncology, 3 Department of Pediatrics, 4 Faculty of Medicine & Dentistry, University of Alberta Diffuse intrinsic pontine glioma (DIPG) is an aggressive pediatric brainstem tumour. Heterozygous missense mutations in the histone H3 variant H3.3 or H3.1, resulting in a lysine 27-to- methionine substitution (H3K27M) are frequent in DIPG. H3K27M mutant histones have been shown to inhibit di- and tri methylation of H3K27 in a dominant negative manner by inhibiting the histone methyltransferase EZH2. The resulting global reduction in the repressive epigenetic marks H3K27Me2/3 is thought to reprogram gene expression to help drive tumourigenesis. The Distal-less homeobox gene 2 (Dlx2) is a homeobox transcription factor involved in neuronal-glial cell fate decisions and differentiation in the vertebrate forebrain. Decreased expression of Dlx2 in H3.3K27M-harbouring tumours has been observed compared to tumours with an alternate H3 mutation (G34R/V) or wild-type H3 only. We hypothesized that Dlx2 overexpression might alter H3K27 methylation in this disease. We stably transfected a H3.3K27M containing murine DIPG cell line with either GFP or Dlx2-GFP and assessed levels of H3K27M and H3K27Me3 by western blot. Dlx2- overexpressing DIPG cells showed a substantial increase in H3K27Me3 levels compared to untransfected and GFP-alone DIPG cells, in which H3K27Me3 was absent. In addition, we observed a reduction in H3K27M levels in Dlx2-overexpressing DIPG cells. We assessed

expression of DLX2 transcriptional targets by qRT-PCR and found significant upregulation of the DLX targets Gad1 and Gad2 in transiently transfected Dlx2-overexpressing DIPG cells compared to controls, indicating competency to respond to DLX-mediated gene regulation. While further work is needed to elucidate the mechanism(s), our preliminary data suggest a role for Dlx2 in modifying the epigenetic landscape of DIPG. Future experiments will include generation of an orthotopic mouse model to investigate the effect of Dlx2 overexpression on H3K27M DIPG tumour behaviour in vivo, and RNA-seq to evaluate gene expression changes in H3K27M DIPG cells and Dlx2-overexpressing H3K27M DIPG cells.

audit/feedback reports to practitioners through a secure web-based portal; reports contain individual physician data and aggregate provincial data for each indicator to inform and improve practice.

RESULTS: Compared to baseline (2010-2013), by 2015 there was a 14% increase in the use of preoperative staging MRI, provincially. Reporting also improved for essential elements on rectal staging MRI, including distance to mesorectal fascia (22 to 81%), extramural venous invasion (17 to 70%), relation to anal sphincter (29 to 78%), and relation to peritoneal reflection (6 to 64%). Surgical technique improved with 91% of rectal specimens graded as ‘complete’ or ‘near-complete’ and a margin positivity of 7% on pathology. Nearly all (94%) pathology reports were completed synoptically, with 90% reporting all mandatory data elements.

38 Laura VanderSluis Agricultural, Life & Environmental Sciences Agricultural, Food & Nutritional Science Eicosapentaenoic acid and docosahexaenoic acid decrease cell viability and are differentially incorporated into triple negative and Her2 positive human breast cancer cells Laura B. VanderSluis, Susan Goruk, Marnie Newell, Sambasivarao Damaraju, and Catherine J. Field Omega-3 long chain polyunsaturated fatty acids (n-3 LCPUFA), docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), decrease cell viability of triple negative (ER-, PR-, Her2-) MDA-MB-231 and Her2+ SK-BR-3 breast cancer (BC) cells and are reported to act via different mechanisms. Studies have yet to systematically investigate if there is a benefit of combining EPA and DHA. The objective of this study was to compare EPA, DHA, and EPA:DHA mixtures on cell viability and membrane phospholipid (PL) incorporation in BC cells. Cells were incubated with a control fatty acid mixture or 100 micromolar of EPA, DHA, or EPA:DHA mixtures (1:1 or 1:2). Cell viability was reduced by all treatments in MDA-MB-231 BC cells relative to control (26-30%, p<0.05). PL incorporation was similar with EPA and DHA when provided alone. As predicted, EPA and DHA were incorporated to the same extent in the 1:1 mixture (48 % vs. 57%), while DHA incorporation was double that of EPA in the 1:2 mixture (33% vs. 67%, p<0.05). In SK-BR-3 BC cells, EPA decreased viability (35%, p<0.05) to a greater extent than DHA yet EPA was incorporated to a lesser extent into PL (6 vs. 31-fold increase p<0.05). EPA:DHA mixtures decreased viability (23-26%, p<0.05) and increased total incorporation (8 and 9-fold increase), similar to EPA alone, but not to the same extent as DHA. DHA incorporation was double that of EPA in the 1:1 mixture (37% vs. 63%, p<0.05) and higher than predicted in the 1:2 mixture (27% vs. 73%, p<0.001). These data confirm that n-3 LCPUFA provided alone or together reduce BC cell viability. In SK-BR-3 BC cells, EPA was more potent than DHA, despite preferential incorporation of DHA. This work will be important for designing targeted treatments for different BC subtypes to reduce BC cell viability and target membrane changes.

39 Kevin Vo Medicine and Dentistry Oncology Establishing and targeting the calpain-NFI signaling pathway in malignant glioma Kevin Vo, Rebecca Burchett, and Roseline Godbout Malignant gliomas (MG) are highly infiltrative tumours with a dismal prognosis. Nuclear factor I (NFI) is a family of 4 transcription factors (NFIA, B, C and X) implicated in the regulation of genes involved in MG cell migration and infiltration, particularly the neural stem cell marker, brain fatty acid binding protein (B-FABP). NFI activity is regulated by its phosphorylation status, with hypophosphorylated NFI being the active form in the context of B-FABP. Our results indicate that the phosphatase calcineurin is able to dephosphorylate NFI. In turn, calcineurin is cleaved and activated by calpain proteases. We have identified CAST, a gene that encodes calpain inhibitor, calpastatin, as a putative target of NFI based on chromatin immunoprecipitation.

Putative NFI binding elements are located in intron 3 of the CAST gene. To determine whether there is a bona fide alternative promoter within intron 3 of CAST, we carried out gel shifts as well as luciferase reporter gene assays using both the canonical and alternative promoter of CAST. These assays confirmed CAST alternative promoter and suggest a preferential usage for this promoter in MG cells with hypophosphorylated (active) NFI. Knockdown of individual NFIs revealed a role for NFIC and NFIX in the repression of CAST gene expression, specifically in cells expressing the hypophosphorylated form of NFI. NFI depletion also altered the subcellular localization of both calpastatin and calineurin protein. Our results suggest a feedback loop for the NFI –calcineurin – calpain – calpastatin pathway in MG cells which may regulate and enhance cell migration.

40 Amy Kirkham Medicine & Dentistry Biomedical Engineering Caloric Restriction and Exercise protection from Anthracycline Toxic Effects: (CREATE) Study Amy Kirkham, Ian Paterson, John Mackey, Carla Prado, Kerry Courneya, Edith Pituskin, Richard Thompson Anthracycline chemotherapy agents play an important role in third generation regimens for early breast cancer treatment. Unfortunately, their use is associated with a dose-related and potentially irreversible cardiotoxicity, as well as less-documented toxicity affecting the vessels and skeletal muscle. Anthracycline-related toxicity is likely a contributor to the increased risk of cardiovascular disease and death, exercise intolerance, experience of fatigue and reduced quality of life among breast cancer survivors. Exercise and caloric restriction are novel potential cardio-protection strategies, but longer-term adherence, especially concurrent to ongoing chemotherapy treatment is not feasible for many patients. The primary purpose of this study is to determine whether short-term administration of these interventions in close proximity to each anthracycline treatment will reduce the detrimental effects of anthracyclines on the heart, aorta, and skeletal muscle. Fifty-six early stage breast cancer patients who will receive anthracyclines will be randomly assigned to either: 1) a single 30-minute aerobic exercise session performed 24 hours prior to each treatment; 2) a 50% reduction in caloric intake for 48 hours prior to each treatment; or 3) usual care. Outcome measures will include reserve capacity (peak capacity – rest) of left ventricular ejection fraction, aortic distensibility, isolated skeletal muscle oxygen consumption, and whole body oxygen consumption (i.e. aerobic fitness). Peak capacity will be measured using maximal incremental exercise tests. Cardiac, aortic, and skeletal muscle function will be measured using novel magnetic resonance imaging (MRI) techniques. Outcomes will be assessed prior to chemotherapy, end of treatment, and one year after starting treatment. Exploratory outcomes will include patient-reported quality of life and fatigue, and clinically measured tumor size at the end of treatment in patients receiving neoadjuvant treatment.

These interventions will be immediately accessible by all patients at all institutions and have the potential to improve the health of breast cancer survivors.

41 Andrei Ghila Medicine & Dentistry Oncology Experimental Verification of EGSnrc Monte Carlo Calculated PDDs in a Parallel Magnetic Field Andrei Ghila, Dr. Stephen Steciw, Dr. B. Gino Fallone, Dr. Satyapal Rathee Integrated linac-MR systems are uniquely suited for real time tumour tracking during radiation treatment. Understanding the magnetic field dose effects and incorporating them in treatment planning is paramount for linac-MR clinical implementation. We experimentally verified the EGSnrc dose calculations in the presence of a realistic magnetic field parallel to the radiation beam travel. Two cylindrical bore electromagnets produced a 0.21 T magnetic field parallel to the central axis of a 6 MV photon beam. A parallel plate ion chamber was used to measure the PDD in a polystyrene phantom, placed inside the bore in two setups: phantom top surface coinciding with the magnet bore center (183 cm SSD), and with the magnet bore’s top surface (170 cm SSD). We measured the field of the magnet at several points and included the exact dimensions of the coils to generate a 3D magnetic field map in a finite element model. BEAMnrc and DOSXYZnrc simulated the PDD experiments in parallel magnetic field (i.e. 3D magnetic field included) and with no magnetic field. With the phantom surface at the top of the electromagnet, the surface dose increased by 10% (compared to no-magnetic field), due to electrons being focused by the smaller fringe fields of the electromagnet. With the phantom surface at the bore center, the surface dose increased by 30% since extra 13 cm of air column was in relatively higher magnetic field (>0.13T) in the magnet bore. EGSnrc Monte Carlo code correctly calculated the radiation dose with and without the magnetic field, and all points passed the 1%, 1 mm Gamma criterion when the ion chamber’s entrance window and air cavity were included in the

simulated phantom. A parallel magnetic field increases the surface and buildup dose during irradiation. The EGSnrc package can model these magnetic field dose effects accurately.

42 Anyeld Ubeda Engineering Biomedical engineering Exploring novel molecular therapeutics for Acute Myeloid Leukemia Anyeld Ubeda, Cezary Kucharski, Binthi Samuel, Remant KC and Hasan Uludag Acute Myeloid Leukemia (AML) is currently the most prevalent and deadliest form of leukemia. Significant progress has been made in the development of new leukemia therapies, yet for AML the standard treatment remains to be chemotherapy and only about 30% of patients obtain long term survival. Hematopoietic stem cell transplantation allows for the prevention of recurrence and increased disease-free survival, however, it has the highest treatment-related morbidity and mortality but it is only recommended for patients with poor-risk disease. As a result, new treatment options that pose lower risks and offer alternatives are of urgent need, especially for patients that relapse after chemotherapy and are not eligible for transplantation. In this study, we explore the use of RNA silencing as a promising candidate to be used in conjunction with traditional treatments as it has the potential to act in synergy to lower doses, length of treatments and side effects. Specifically, we are identifying molecular targets using lipid modified polyethylenimine (PEI) delivery systems that could be used as the basis of therapy during AML progression using the stem-cell like KG1a cell line and primary cells. We identified linoleic acid and lauric acid modified PEI (PEI 1.2 LA 6 and PEI 1.2 Lau 8) and Lipofectamine as effective and versatile carriers. Additionally, we have identified that targeting anti-apoptotic genes, BCR5, and BCL-2 and mitogenic regulator RPS6ka5 with siRNA/PEIs reduce DNA replication in KG1a cells and are currently testing their effects in primary cells. We are now inspecting silencing at the mRNA levels, as well as

apoptotic and proliferative effects in primary cells. By selecting different molecular targets and delivery systems, we aim to develop versatile combination therapies that in the future can be individually tailored and can be better translated to clinical applications.

43 Marnie Newell ALES AFNS Feeding the omega-3 fatty acid, docosahexaenoic acid improves the efficacy of docetaxel in mice bearing breast cancer patient derived xenografts Marnie Newell, Miranda Brun, Lynne Postovit, Catherine J. Field Docosahexaenoic acid (DHA), a polyunsaturated fatty acid found in fatty fish, reduces breast cancer cell growth in vitro and in vivo. Our lab has shown that pre-treatment of MDA-MB-231 cells with DHA prior to chemotherapy results in decreased cell growth and increased apoptosis. Because immortalized cell lines do not represent the heterogenicity seen in human tumours, we sought to confirm this work using patient derived xenografts (PDX). NSG (NOD/SCID/IL2 γ- receptor null) mice (6-week-old female) bearing subcutaneous triple negative PDX tumours (100 mm3) were randomized to one of two nutritionally adequate high fat diets (20% w/w ±5% DHA). Half the animals in each group were treated IP twice weekly with docetaxel (TXT,5mg/kg) for 6 weeks (n=6 in each of the 4 groups). Feeding DHA in conjunction with TXT resulted in smaller tumours (1.7 ± 0.1g vs. 5.2±0.5g) and a higher content of DHA (5.5 ± 0.0 vs 2.6±0.3%) in tumour phospholipids. Immunohistochemical staining of Ki67, CD95 and TUNEL confirmed lower proliferation and increased apoptosis in the DHA+TXT tumours (p<0.05). Protein analysis confirmed increased expression of proteins involved in apoptosis including CD95, RIPK1 and Caspase 10 in DHA+TXT tumours (p<0.05). This study confirms, for the first time in patient derived xenografts, that feeding a diet supplemented with DHA facilitates the anti-cancer effect of TXT on breast cancer cells and

suggested that this may be due to higher incorporation of DHA reduced proliferation and increased apoptosis. (supported by CIHR)

44 Sara Omar Medicine & Dentistry Oncology Finding novel PNKP inhibitors using multiple-technique virtual screening Sara Ibrahim Omar, Jack Tuszynski Polynucleotide kinase/phosphatase (PNKP) is a very versatile and crucial DNA repair protein. It has dual catalytic function: it can both phosphorylate 5'-termini and dephosphorylate 3'- termini of DNA, through its kinase and phosphatase activities, respectively. The phosphatase domain of PNKP plays a major role in repairing DNA strand breaks caused by ionizing radiation (IR). Since IR is still an essential pillar of the treatment regimens for many cancer types, a significant number of patients are resistant to these treatments because their tumors possess robust DNA repair capacity. Our aim is to find a novel PNKP phosphatase inhibitor, which can potentially improve the therapeutic benefits of IR. To do so, we virtually screened the Commercial Compound Collection (CoCoCo) database (3.7M compounds) to find novel inhibitors of PNKP. Since the crystal structure of the human PNKP (hPNKP) has not been experimentally resolved, we used the X-ray crystal structure of mouse PNKP, PDB ID: 3U7G, to create the homology model of hPNKP. We simulated the hPNKP homology model for 250 ns using AMBER molecular dynamics software. We extracted the DNA-protein complex structures every 0.5 ns for a total of 460 representative DNA-protein structures. For each of the 460 complex structures, we automatically generated a pharmacophore model for the PNKP bound DNA. The CoCoCo database was virtually screened against the 460 pharmacophore models. The hits from these 460 screens were then assessed using pharmacophore-assisted docking into 10 representative structures of the protein. The top hits were then identified based on their average calculated binding energies to PNKP and the number of times they scored as hits in

each screen. Further, the compounds were screened based on their calculated synthetic feasibility and predicted ADMET properties. These compounds will be purchased for experimental validation.

45 Diana Diaz-Dusan Engineering Chemical & Materials Engineering Nanotheranostics to manage hypoxic cancerous tumors. Diana Diaz-Dussan, Yi-Yang Peng, Ravin Narain and Piyush Kumar. Hypoxia promotes metastasis, invasion, tumor progression and recurrence through resistance to radiotherapy and (due to impaired drug delivery) to chemotherapy. Tumor hypoxia thus poses a formidable challenge for nanotechnology medicine by improving colocalization and “smart delivery” of pharmaceuticals to control cancer progression and ultimately its remission. This study pursues a novel, translational and beyond state-of- the-art theranostic approach to the management of hypoxic solid tumors. Iodoazomycin arabinoside (IAZA), an O 2 -mimetic nucleoside-based imaging agent, that accumulates selectively in hypoxic tumors have been successfully used to diagnosed hypoxic tumors and has an unparalleled potential for therapeutic purposes; either as a radiosensitizer or for in situ molecular radionuclide therapy. Proof-of- principle studies in our laboratory have confirmed that the encapsulated form of IAZA, in functionally modified carbohydrate-based nanogels (NGs), improved the bioavailability of the drug in hypoxic cancer cells and enhanced its radiosensitization potential. The NGs used are composed of LAEMA (2-lactobionamidoethyl methacrylamide) with an outer shell of thermoresponsive DEGMA (di(ethylene glycol) methyl ethyl methacrylate) and cross-linker inner core of MBAm (N,N´-methylenebis(acrylamide)) offering great biocompatibility and lower toxicity. Further applications and optimization of these nanotheranostics through modification of its hydrodynamic size, composition of the core, hydrophobicity, surface/core charge and cross-linker percentage have

demonstrated high drug-loading capacity (@ 80-88%) and time-controlled release over 50 h. nanoIAZA (encapsulated IAZA complex) demonstrated superior sensitization enhancement ratio (SER) compared to IAZA alone (SER 0.1 mM IAZA ~ 1.40 vs. SER nanoIAZA ~ 1.58 and SER 0.1 mM IAZA ~ 1.09 vs. SER nanoIAZA ~ 1.41 ) in PC3 and FaDu cell lines respectively, in brief, delivers IAZA in therapeutically more effective doses to hypoxic cells compared to IAZA alone. This will enhance therapeutic effects without causing undesired toxicity to healthy (oxygenated) cells (where IAZA will not activate) providing more effective hypoxic cancer management.

46 Cody Lewis Medicine and Dentistry Oncology High Myt1 expression in breast cancer cells facilitates resistance to the Wee1 inhibitor MK- 1775 Cody W Lewis, Won Shik Choi, and Gordon Chan Mitosis is negatively regulated by Wee1 (nuclear) and Myt1 (membrane) kinases, which add inhibitory phosphates to Cdk1 on Thr14 and Tyr15. The small molecule Wee1 inhibitor MK- 1775 induces premature mitosis resulting in centromere fragmentation. In addition, MK-1775 increases the mitotic duration and induces cell death independent of centromere fragmentation; prometaphase synchronized cells also arrest in mitosis following MK-1775 treatment. MK-1775 is currently undergoing phase I/II clinical trials against different cancers, including those that affect the breast. There are no small molecule inhibitors of Myt1. Analysis of patient breast cancer tissue has shown that Myt1 is upregulated compared to normal breast tissue. Moreover, high Myt1 expression is correlated with a worse clinical outcome (higher disease relapse and reduced overall survival). We treated different breast cancer cells with MK-1775 and found that they exhibited a differential response; cell line response was strongly associated with the expression of Myt1 (mRNA and protein). We find that knocking down Myt1 sensitizes breast cancer cell lines to MK-1775 by as much as 9-fold.

We measured the mitotic duration (NEBD to anaphase) in HeLa and MDA-MB-231 cells and found that Myt1 knockdown (KD) in the presence of 250 nM MK-1775 prolongs mitosis 2-5 fold compared to mono-treatments (MK-1775 or siMyt1 alone). We observe a cell death morphology that is consistent with that of apoptosis and extracts of cells treated with MK- 1775 and siMyt1 contain cleaved caspase 3 and PARP. Myt1 inhibits Cdk1 by phosphorylation and sequestration outside the nucleus. We confirmed that Myt1 can still phosphorylate Cdk1 on Thr14 in the presence of MK-1775, but it is not clear if Myt1 kinase activity alone is required for promoting resistance. Understanding how Myt1 is mediating MK-1775 resistance in breast cancer cells will have important implications for the future development of small molecules that target Myt1.

47 Aravindhan Ganesan Pharmacy Pharmacy How do the co-stimulatory CD28/B7-1 interactions differ from the co-inhibitory interactions of CTLA/B7-1 for orchestrating T-cell immune responses? Aravindhan Ganesan, Tae Chul Moon, Khaled Barakat Activation of T lymphocytes (or T-cells) plays a central role in anti-cancer immune responses, which controls tumor initiation and progression. CD28 and CTLA-4 are transmembrane receptors that are responsible for the regulation of the second signal that is crucial for T cell activation. These T-cell receptors share some extents of sequence and structural conservation; and they also bind to the same B7 ligands expressed on the antigen-presenting cells. Nevertheless, CD28/B7 interactions trigger a co-stimulatory signal for T-cell activation; while, CTLA-4/B7 interactions generate an inhibitory signal that inactivates the T-cells. Thus, understanding the molecular bases of these key protein-protein interactions at the immunological synapse are important for targeted immune checkpoints-based cancer therapy. Although, the X-ray crystal structure of CTLA-4/B7-1 has been reported earlier, however, the complex structure of CD28/B71 has not been resolved experimentally. In this

work, we employed a combination of advanced molecular modelling and extensive molecular dynamics (MD) simulations to model the CD28/B7-1 complex and characterize the key interactions that stabilize the complex. The ensemble protein-protein docking and MD-based binding-free energy calculations were useful to obtain the structure of the CD28/B7-1 complex, which was validated with various mutation-based experimental data (from the literature) and binding assay experiments performed here. Later, extended MD simulations of CTLA:B7-1 and CD28-B7-1 complexes revealed the similarities and differences in their interactions at molecular level. The results presented in this work will, on a long-run, lead to significant insights for designing novel inhibitors to specifically target CTLA-4 and unleash the immune system against cancer.

48 Ryan Noyce Medicine and Dentistry Medical Microbiology & Immunology Construction of infectious orthopoxviruses from chemically synthesized DNA fragments for use as novel oncolytic viruses Ryan Noyce, Megan Desaulniers, Seth Lederman, and David Evans Molecular analyses show that modern vaccinia virus (VACV) vaccine strains share common ancestry with horsepox virus (HPXV). Given that the toxicity of modern VACV vaccines limits their use, we asked whether a HPXV-based vaccine might still provide a safer alternative. Since HPXV may be extinct and the only specimen of HPXV that has been identified and sequenced is unavailable for investigation, we explored whether a stock of HPXV could be obtained by large-scale gene synthesis. Ten large (10-30 kb) fragments of DNA were synthesized based on the HPXV sequence along with two 157 nt VACV terminal sequences, and were recombined into a live synthetic chimeric HPXV (scHPXV) in cells infected with Shope fibroma virus (SFV). Sequencing of the 212 kbp scHPXV confirmed it was a faithful copy of the input DNA. We believe this is the first complete synthesis of a poxvirus. This scHPXV grew slower, produced smaller plaques, and exhibited less virulence in mice than VACV, but

still provided vaccine protection against a lethal VACV challenge. We are currently using this approach to rapidly construct novel oncolytic orthopoxviruses expressing tumour-specific antigens to induce potent anti-tumour immune reponses in tumour-bearing animals. We anticipate that these next-generation oncolytic poxviruses will eventually lead to personalized oncolytic vectors to treat cancer.

49 Daniel Krys Medicine and Dentistry Oncology Hypoxia-Induced Regulation of Membrane Transporters in Breast Cancer Daniel Krys, Ingrit Hamann, Melinda Wuest, Frank Wuest

Hypoxia-inducible factor (HIF)-1 is the master homeostatic regulator during hypoxia in human cancer cells. Reduced oxygen supply in cancer cells leads to tumor hypoxia, activates HIF-1a, which then controls the expression of multiple target genes, including membrane transporters like GLUT1. Recently we have also demonstrated regulation of fructose transporter GLUT5 via HIF-1a in breast cancer (BC). To determine if and how tumor hypoxia regulates other membrane transporters in BC, we studied protein expression and functionality of nucleoside transporter hENT1 and amino acid transporter LAT1 under normoxic and hypoxic conditions. Cellular uptake experiments were performed with radiolabeled nucleoside [18F]FLT and radiolabeled amino acid [18F]FDOPA in ER(+) MCF7 and triple-negative MDA-MB231 BC cells. Higher [18F]FLT uptake was observed in MDA-MB231 cells (241±10% radioactivity/mg protein) compared to ER(+) MCF7 cells (147±18% radioactivity/mg protein) at 60 min. This also corresponded with detected higher hENT1 protein expression levels in MDA-MB231 versus MCF7 cells. Data also indicated that [18F]FLT uptake and hENT1 expression levels were not influenced significantly by hypoxia. In contrast, LAT1 expression was higher in ER(+) MCF7 versus MDA-MB231 cells. Initial cell uptake experiments with [18F]FDOPA revealed no

difference between both BC cell lines under normoxic conditions, whereas a ~50% increased uptake in MDA-MB231 was observed under hypoxia. We found that nucleoside transport via hENT1 is not regulated by hypoxia, while hypoxic regulation amino acid transporter LAT1 still needs further investigation.

50 Ana Anoveros FGSR AFNS Immune cell characterization in the muscle tissue of patients with cancer Ana Anoveros, Amritpal S. Bhullar, Abha Dunichand-Hoedl, Karen Martins, David Bigam, Vickie Baracos and Vera Mazurak Background: Inflammation is a recognized contributor to muscle wasting. Gene array analysis in human biopsies has revealed that immune cell recruitment is a major event occurring in cancer patients experiencing systemic inflammation. Lymphoid and myeloid cells in the tissue influence muscle atrophy through diverse mechanisms; however, little is known about the muscle’s immunological environment and its relationship to muscle loss in conditions of malignancy. Therefore, we hypothesized that features of muscle atrophy are related to a higher recruitment of immune cells in the muscle of patients with cancer.

Methods: Rectus abdominis biopsies and computed tomography (CT) images were collected from cancer patients (n=22) at the University of Alberta Hospital, Edmonton, Canada, from July 2015 to August 2016. Frozen muscle biopsies cut in transverse sections were stained with immunofluorescence to evaluate immune cells: T cells (CD8+ and CD4+), antigen presenting cells (APCs; CD3-CD4+), and granulocytes (CD11b+). Features of muscle atrophy were explored by evaluating mean muscle fiber area (µm2), and calculating skeletal muscle index (SMI; cm2/m2) from CT images. Spearman’s coefficient was used to assess correlations.

Results: Muscle with higher SMI (cm2/m2) (r=0.66, P=0.001) and mean fiber area (µm2)

(r=0.55, P=0.008) had a larger number of T cells (CD4+ and CD8+ combined). CD8+ T cells (not CD4+ T cells) maintain a significant association with SMI (r=0.54, P=0.008) and mean muscle fiber area (r=0.49, P=0.021). APCs and granulocytes were not related to any muscle feature evaluated; however, APCs were positively correlated to number of CD8+ T cells (r=0.45, P=0.036).

Conclusions: Lymphoid and myeloid cells are present in the muscle of cancer patients. In particular. T cells seem to be associated with better muscle condition. Results suggest interaction between CD8+ T cells and APCs. Further exploration will enable understanding the influence of immune cells within the muscle of cancer patients.

51 Ericka Wiebe Medicine & Dentistry Oncology Volumetric preplanning for intracavitary/interstitial cervix brachytherapy: at what cost? Fleur Huang, Deepak Dinakaran, Ericka Wiebe, Wendy Read, Leanne Kellogg , Michele Janoski, Robert Pearcey, Geetha Menon PURPOSE In curative cervix cancer radiotherapy (RT), MR imaging, hybrid intracavitary (IC)/interstitial (IS) applicators, and complex brachytherapy (BT) planning converge for better dose delivery. Optimal IS catheter placement is expected with an initial dry run (MR-guided IC-alone implant, volumetric preplanning (VPP) using virtual catheters) before actual IC/IS implant, MR-based BT planning and delivery. In Canadian settings, the increased resource utilization required of VPP for ideal IC/IS cervix BT has not been defined.

METHODS In this cost exercise, the base case was a formal IC/IS VPP cycle, an add-on to a simple single- fraction MR-based BT workflow. Costs were considered over a typical treatment course, and

mapped to distinct processes and service items within the reference VPP cycle. For better translatability across jurisdictions, costs categorized into time and material (consumable or reusable) amounts were not converted. Frontline staff and managers were surveyed to generate primary local data on 2016 cost estimates (health sector perspective). Opportunity costs were elicited. Variant IC/IS VPP practice scenarios were assessed for cost implications. Costs associated with the lack of VPP were also reviewed. RESULTS Ten experienced providers (Radiation Oncology (RO), Medical Physics (MP), Dosimetry (Do), BT) and 3 resource managers (RT, Operating Room (OR), and Diagnostic Imaging (DI)) estimated time and material costs, anticipating added processes for a formal VPP cycle to run in our setting. Activity-related time (in minutes) ranged from +200 in DI to +650 in OR, with RT-specific time reflecting our multidisciplinary practice: +130 MP, +200 RO, +230 Do, +280 BT, +60 other. Resource utilization was heaviest for the OR-based implant and for BT planning (190 min provider time spent), eclipsing the costs of VPP-related consumables and poorly sensitive to variances such as omitting CTsim or switching anesthesia. Many costs, direct and indirect, could not be counted simply because reference data was unavailable or unknown for our setting. Opportunity costs were cited often, mainly for high-demand shared resources like DI. In our VPP-less IC/IS practice, a median of 3 IS catheters are implanted but approximately 18% not loaded, for calculated waste (unused consumables, processed reusables) that is negligible (< $100/implant) compared to the expenditures needed to generate and reproduce an ideal IC/IS BT plan. Actual rate of IS intrusion into adjacent organs is low (1/43), with no complication-related clinical activity to warrant next-level costing.

CONCLUSIONS Dosimetric benefits are well-described with MR-based cervix IC/IS BT, but clinical effectiveness data have yet to mature. The adoption of optimum IC/IS BT workflows should be guided by pragmatic evaluation of costs and adapted to local resource contexts. Time- driven activity-based costing, where feasible, may be informative.

52 Bryson Dietz Medicine & Dentistry Oncology Integration of PCA with compressed sensing acceleration for rapid real-time imaging and on- the-fly reconstruction Bryson Dietz, Eugene Yip, Jihyun Yun, Gino Fallone, Keith Wachowicz This work presents a real-time dynamic image reconstruction technique for magnetic resonance imaging (MRI), that combines compressed sensing acceleration and principal component analysis (CS-PCA). Six fully sampled dynamic data sets of patients diagnosed with non-small cell lung cancer acquired at 3T were used to investigate the CS-PCA algorithm. To gain insight into as to how the reconstruction might proceed at lower fields, 6x noise was added to the 3T data to investigate how the algorithm handles noisy data. The six data sets were retrospectively undersampled using acceleration factors ranging from 2-10x. Metrics to determine the reconstruction quality included the normalized mean square error (NMSE), as well as the dice coefficients (DC) and centroid displacement of the tumour segmentations. Our results demonstrate that CS-PCA performed superior than CS alone. The CS-PCA patient averaged DC for 3T and the 6x noise added data remained above 0.9 for acceleration factors up to 10x. The patient averaged NMSE gradually increased with increasing acceleration; however, it remained below 0.06 up to an acceleration factor of 10x for both 3T and 6x noise added data. The CS-PCA reconstruction speed ranged from 5-20 ms (Intel i7-4710HQ CPU @ 2.5 GHz), depending on the chosen parameters. A real-time reconstruction technique was developed for the purpose of adaptive radiotherapy using a Linac-MRI system. Our CS-PCA algorithm can achieve tumour contours with DC greater than 0.9 and NMSE less than 0.06 at acceleration factors of up to, and including, 10x. The reconstruction speed for CS-PCA reconstruction speed ranged from 5-20 ms implemented using non-optimized MATLAB code. The CS-PCA technique is currently being implemented for use in prospective real-time imaging at the MRI console.

53 Jianyin Liu Medicine and Dentistry Oncology Interaction between gut microbes and genetic instability in anti-colon cancer immunity Jianyin Liu, Courtney Mowat and Kristi Baker Microsatellite instable (MSI) colorectal cancer (CRC) is caused by deficient DNA mismatch repair and constitutes 15% of CRCs. MSI CRC has a better prognosis and stronger immune response than the more common chromosomal instable (CIN) CRC. The colon contains the largest number of bacteria in the body and increasing evidence suggests that gut microbiota have a strong correlation to CRC. Gut microbes and their metabolites can circulate through the whole body and this may contribute to cancer progression and tumorigenesis at either the microbial site or at a distant location from where the microbes originally reside. Microbes can contribute to genomic instability directly through genotoxins and oncoproteins, which induce changes to cellular DNA and impair DNA repair mechanisms, or indirectly by increasing epithelial cell replication and inducing inflammation. To understand if the interactions with intestinal bacteria that are known to promote or protect against CRC contributes to the different immune responses between MSI CRC and CIN CRC, we are going to compare how CRISPR mutated cells lacking MSI or CIN-associated genes respond to stimulation with whole bacteria that are either probiotic or infectious. We expect to observe different activation patterns of pattern recognition receptor (PRR), which regulate host cell recognition of bacterial components, after stimulation of MSI and CIN CRC cell lines with microbial species. We expect these patterns to reflect the magnitude of anti-tumor immune response seen in patients. Furthermore, we anticipate uncovering novel mechanisms for the immunogenicity of MSI CRC and identifying specific molecules or microbes that could be targets for a therapeutic intervention, which would increase anti-tumor immune responses against CRC.

54 Fleur Huang Medicine & Dentistry Oncology Intracavitary/interstitial MR-based cervix brachytherapy: first look at implant quality Fleur Huang, Deepak Dinakaran, Ericka Wiebe, Brenda Rose, Wendy Read, Robert Pearcey, Geetha Menon PURPOSE Image-guided adaptive brachytherapy (BT) holds promise for improved cervical cancer outcomes, with lesser normal tissue toxicity. Key to this are better tumor visualization (via MRI), defined dose-volume targets (via international recommendations), and plan conformity (via interstitial (IS) sources added to traditional intracavitary (IC) applicators). Hybrid IC/IS implant quality is not well characterized, nor its impact on overall BT plan quality. METHODS Our institutional experience with IC/IS MRI-based cervix BT (no volumetric preplanning; post- implant planning on MRI assisted by CTsim) was reviewed. Tandem-ring (T&R) and tandem- ovoid (T&O) based IC/IS implant quality was evaluated for variability in the IS component, with attention to intent at implant vs post-implant geometry. Descriptive statistics were performed. RESULTS From 07/2013 – 12/2016, IC/IS technique was used in n=43 single-fraction pulsed dose rate BT boost cases, during curative (chemo)radiotherapy for locally advanced cervix cancer. Median HR-CTV D90 (EQD2) was 90.7 Gy [76.4 – 96.6], median D2cc 75.8 Gy [43.2 – 90.4] for bladder, 64.3 Gy [52.3 – 74.2] rectum, 62.2 Gy [46.6 – 73.6] sigmoid, and ICRU rectovaginal point dose, 64.2 Gy [52.0 – 82.1]. Of a median of 3 [1 – 6] IS catheters implanted per case (79% T&R), 2 [1 – 5] were loaded, for an overall load rate of 82%, with no appreciable difference by applicator type. More IS catheters (N=10, in 6 unique implants) remained unused among our first 15 cases, compared to the latest 15 (N=7, in 6 implants) in our evolving experience. Radial IS positions, selected based on pre-BT clinical/radiological tumor

residuum and patient anatomy, were loaded most frequently lateral for T&O, and lateral more than anterior for T&R. Least loaded were posterior (T&R) and medial (T&O) positions. There was no change in IS catheter depth on CTsim vs MRI, suggesting no significant movement. Median IS depth beyond ring/ovoid surface was 26.0 mm [7.5 – 47.1], closely matching intended insertion depths, with mean difference of 1.4 ± 3.0 mm. IS catheter deflection was observed (median 1.6 deg [0 – 34.3] anterior-posterior; 1.0 deg [0 – 35.6] medial-lateral), reaching > 5 deg for median insertion depths of 25.6 mm [11.5 – 43.0]. IS weighting ranged from 0.5% to 70.2%, although the median was 7.7% relative to the IC component, with outliers reflecting individual anatomic and tumor topographies. CONCLUSIONS The technical quality of IC/IS cervix BT implants improves with experience, but small variabilities persist in measured IS catheter positions. To what extent post-implant BT planning processes can safely mitigate for suboptimal IS tracks (assuming IC applicator selection appropriate to the target) remains unknown.

55 Amirali Bukhari Medicine & Dentistry Oncology Investigating Synthetic Lethal Interactions in DNA Damage Signaling for Breast Cancer Therapy Amirali Bukhari, Deandra Luong, Joanna Pearce, Cody Lewis, Gordon Chan, Armin Gamper Breast cancer is the most common cancer amongst women in Canada. 10 – 20 % of patients receiving the present standard treatment regime for early stage breast cancers, breast- conserving surgery with radiation therapy, develop local recurrence. Some patients develop metastatic spread associated with a high lethality rate. The underlying cause is likely the failed eradication of tumour initiating cells or cancer stem cells (CSCs). CSCs show characteristically high plasticity and unfortunately often also increased radioresistance compared to the bulk of the cancer cells. Radiosensitizers, drugs that increase ionizing

radiation (IR)-induced tumour cell killing, promise to improve the efficacy of RT. ATR and Wee1 inhibitors have been found to radiosensitize (bulk) cancer cells and now are in phase I/II clinical trials. Following ionizing radiation (IR), DNA damage is signalled by two apical kinases of partially overlapping pathways, ATR and ATM. Of the two, especially ATR activity is often upregulated in cancer stem cells. We hypothesize that: ATR signalling is fundamental for the observed enhanced radioresistance of breast cancer stem cells (BCSCs) and responsible for breast cancers being refractory to radiation therapy. Our data suggests that in vitro inhibition of ATR radiosensitizes the BCSC population. Further, our preclinical findings indicate that ATR inhibition shows tumour-selective synthetic lethality with inhibition of the checkpoint kinase Wee1. The orthotopic breast tumour bearing mice which received combined treatment of the ATR and Wee1 inhibitor showed synergistic killing of tumour cells leading to tumour shrinkage to impalpable levels and suppression of metastasis. As a result, this treatment regime considerably increased overall survival. We are currently expanding our animal studies and will determine the mechanisms underlying synergism between inhibition of ATR and Wee1.

56 Lei Li Medicine & Dentistry Oncology Investigating the role of DEAD box 1 in cell stress response Lei Li and Roseline Godbout DEAD box proteins are putative RNA helicases that function by modifying RNA secondary structure. DEAD box 1 (DDX1) was first identified in our laboratory and was subsequently found to be overexpressed in a subset of retinoblastoma, neuroblastoma and breast cancer tumours. In breast cancer, DDX1 overexpression is associated with a poor prognosis. DDX1 is found in both the nucleus and cytoplasm. In the nucleus, DDX1 is recruited to DNA double strand breaks (DSBs), where it resolves RNA-DNA duplexes, facilitates DSB repair by

homologous recombination and promotes the survival of cells exposed to DSB-inducing conditions. In the cytoplasm, DDX1 localizes to stress granules (SGs), cytoplasmic organelles induced by various noxious stimuli, such as heat shock, oxidative stress and endoplasmic reticulum stress. SGs are disassembled when unfavorable conditions are removed. Although the exact functions of SGs remain elusive, it is generally believed that tightly controlled SG assembly and disassembly is important for cell survival under stress. We found that while DDX1 is dispensable for SG assembly, depletion of DDX1 leads to impaired SG disassembly. Intriguingly, when cells are exposed to both environmental and genotoxic stress, DDX1 is preferentially recruited to SGs and is absent from DSBs. These results suggest that there is crosstalk in cellular response to different types of stressors. We are pursuing the role of DDX1 in stress response by: (1) identifying RNA substrates of DDX1 and examining the fate of these RNAs under stress in the presence and absence of DDX1; (2) studying post-translational modification of DDX1 and its importance in DDX1 localization and; (3) identifying and characterizing proteins that interact with DDX1 under different stress conditions (DSB- inducing condition vs. SG-inducing stimuli).

57 Abul Kalam Azad Medicine and Dentistry Medicine Investigating the role of phosphoinositide 3-kinase beta (p110β) isoform of PI3K in tumor angiogenesis. AK Azad, A Masoud, GY Oudit, RB Moore, AG Murray Background: Drugs targeting VEGF-pathway have been approved to treat various cancer types. However, tumor may become resistance to anti-VEGF therapies due to involvement of multiple other signaling pathways. p110βb isoform of PI3-kinases is uniquely coupled to both receptor tyrosine kinases (RTKs) and G-protein coupled receptors (GPCRs). Our published data showed that capillary formation is impaired in p110β knockout mice, and knockdown of p110β in endothelial cells (EC) attenuates angiogenic sprouting. These data indicate that

p110β is involved in angiogenic pathway.

Hypothesis: p110β isoform of PI3K is involved in tumor angiogenesis.

Methods: B16F10 cells (1*106) were implanted subcutaneously in p110β/Tie2/flx/flx or control (flx/flx) mice. Vehicle or sustained sunitinib (40mg/kg) was initiated when tumor reached an average size of 200mm3, and mice were euthanized when 50% of mice reached an average tumor volume >1000mm3. 2*105 B16F10 cells were injected intravenously in p110β or control mice, and were treated with vehicle or 40mg/kg sunitinib for 20 days. 60mg/kg pimonidazole was administrated 1hour before euthanization at day 21. Immunohistochemical analyses were performed for CD31-positive vessels and pimonidazole- positive tumor hypoxic areas.

Results: EC-specific p110β decreases the growth of B16F10 melanoma line in syngeneic mice after subcutaneous injection compared to the control mice. Similarly, EC p110β loss decreases tumor metastases in the lung, and the overall tumor area in lung cross-section after tail vein injection. Further, B16F10 metastases had a marked decrease in CD31-positive microvessels in p110β vs controls tumors. Surprisingly, pimonidazole-positive area in the tumors was normalized in p110β/Tie2/flx/flx vs control mice.

Significance: Our findings here demonstrated that EC-specific inactivation of p110β decreases primary tumor growth and tumor metastasis accompanied by decreased tumor vasculature, but normalization of the tumor blood flow. Inhibition of endothelial p110β may be useful as adjuvant therapy, and may facilitate delivery and/or response of the tumor to conventional chemotherapy agents.

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58 Martin Srayko Science Biological Sciences Kinetochore recruitment of the C. elegans Ska complex is regulated by the centrosomal PP2A phosphatase regulatory subunit, RSA-1. Karen I. Lange, Aly Suleman, Martin Srayko During mitosis, the microtubule cytoskeleton is restructured to form the bipolar mitotic spindle. Dynamic microtubules interact with chromosomes through kinetochores to move and orient the chromosomes between the centrosomes. During anaphase, the sister chromatids are separated to ensure each daughter cell receives precisely one complement of the genome. Errors in chromosome segregation can cause aneuploidy, which is a driving force in tumour formation. RSA-1, a regulatory subunit for PP2A phosphatase in C. elegans, targets the enzyme to centrosomes and regulates microtubule outgrowth. We observed a genetic interaction between RSA-1 and the Ska complex. For example, increased levels of the Ska complex enhance the embryonic lethality associated with loss of rsa-1 function, and reduced levels of Ska restore embryonic viability. Furthermore, in rsa-1(or598) embryos, SKA- 1 is prematurely and excessively recruited to kinetochores during spindle assembly. RSA-1 also affects the localization of Aurora A kinase on centrosomal microtubules. We present a model whereby RSA-1 regulates Aurora A at the centrosome, which controls Ska complex recruitment at the kinetochores. Additionally, using an RNAi-based screen, we identified several other components required for appropriate Ska localization in the embryo.

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59 Aaron Purchase Medicine & Dentistry Oncology Magnetic resonance imaging contrast and edge enhancement of the animal breast cancer model using double action core-shell nanoparticles Aaron R. Purchase, Barbara Blasiak, Frank C.J.M. van Veggel, Armita Dash, Boguslaw Tomanek Standard clinical magnetic resonance imaging (MRI) is a valuable tool for cancer detection but it often fails to detect cancer in its early stages and/or provides inconclusive results. Contrast agents containing gadolinium are used to increase MRI contrast, as they provide hyperintense T1-weighted MR images of the tumor or other disease. On the other hand, superparamagnetic nanoparticles, such as iron oxide, reduce T2 and T2* creating void signal, have also been employed to localize the disease. Accumulation of these contrast agents is based on differences in the vasculature of tumors and normal tissues. Recently, however, nanoparticles that change both T1 and T2 relaxation times were produced but their synergetic action in cancer detection has not been yet thoroughly investigated. In this report, NaDyF4 (20 nm)/NaGdF4 (0.5 nm) core-shell nanoparticles that change both T1 and T2 relaxation times were applied to enhance MR images of breast tumor. MDA-MB-231 cell line was injected in nude mice providing an animal model of triple negative breast cancer (TNBC) and imaged using a 9.4 T Bruker MRI system. To utilize the double action of the nanoparticles, we combined T1- and T2-weighted MR images pre- and post- injection of nanoparticles providing increased tumor contrast and edge enhancement.

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60 Clara Fallone Medicine & Dentistry Oncology Magnetic Resonance Spectroscopy Methods to Quantify Relative Omega-3 Fatty Acid Content at 3 T Clara J Fallone, Atiyah Yahya Fat unsaturation is relevant to disease. Magnetic Resonance Spectroscopy (MRS) enables fatty acid quantification. Some models assume negligible omega-3 fatty acid content. However, studies show that lower omega-3 linolenic acid content in breast correlates with breast cancer. In addition, a MRS study revealed that smaller breast tissue methyl resonance linewidths were associated with women with breast cancer and women with a higher risk of breast cancer. The change is potentially due to less varied methyl proton groups, indicating differences in omega-3 fat content. In this work, the correlation of methyl linewidth and omega-3 fatty acid content with echo time (TE) was studied for two common in-vivo MRS pulse sequences: PRESS (Point RESolved Spectroscopy) and STEAM (Stimulated Echo Acquisition Mode), at 3T. Optimal TE values were determined for relative omega-3 fatty acid quantification. Spectra were acquired from nine oils as a function of PRESS and STEAM TE. For each TE, the spectral methyl linewidth was plotted against literature omega-3 content. A linear coefficient of determination (R2) was calculated at each TE and used to assess optimal TE values. It was found that non-optimal TEs yield similar linewidths regardless of differences in omega-3 fat content. In contrast, the linewidths broaden as omega-3 fatty acid content increases when the optimal TE is used (160 ms for both PRESS and STEAM). The changes result from differences in J-coupling evolution. The omega-3 methyl protons resonate at ≈ 0.93 ppm and are more weakly coupled than the methyl protons of the other fatty acids, which resonate at ≈ 0.83 ppm. The J-coupling evolution of the omega-3 methyl protons yields in-phase multiplets at optimal TE, widening the collective methyl linewidth. In conclusion, STEAM and PRESS with a TE of 160 ms can be used to correlate methyl linewidth with relative omega-3 fatty acid content at 3T.

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61 Meghan Ferguson Medicine and Dentistry Cell Biology Mechanisms of Bacterial-Induced Intestinal Neoplasia Meghan Ferguson and Edan Foley The microbiome impacts intestinal proliferation across the animal kingdom. For example, in humans intestinal dysbiosis is associated with colorectal cancer and in mice, carcinogen induced intestinal tumour development depends on the presence of intestinal bacteria. Despite the link between the microbiome and neoplasia, the host and bacterial factors involved remain uncharacterized. To address this question, I established a Drosophila model for bacterial-induced neoplasia. With this model, I showed that the microbiome fuels intestinal neoplasia. To identify the bacteria involved in intestinal neoplasia, I associated germ free flies with two common commensals; Lactobacillus brevis and Lactobacillus plantarum. I found that L. brevis causes extensive neoplasia while L. plantarum is relatively benign. Interestingly, these two bacteria colonize the fly to the same levels, which suggests the bacterial species and not the amount fuels neoplasia. Furthermore, the cell wall of L. brevis is sufficient to induce neoplasia, although other factors may be involved. In the future, I will feed flies secreted bacterial factors and isolated cell wall components to determine the precise bacterial factors involved in neoplasia. To identify the host factors responsible for bacterial-induced neoplasia, I will use RNA- Sequencing to determine the transcriptional profiles of purified intestinal progenitor cells in response to L. brevis. I will identify host pathways involved in proliferation, stress, inflammation and metabolism enriched upon association with L. brevis. In future experiments, I will determine whether enriched pathways are necessary for L. brevis-induced neoplasia. For example, if the Hippo pathway is upregulated in response to L. brevis, I will block Hippo activity and measure neoplasia. This study will provide insight into host-microbe interactions that impact neoplasia.

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62 Amit Bhavsar Medicine & Dentistry Medical Microbiology and Immunology Mitigating cisplatin-induced ototoxicity in childhood cancer treatment through applied pharmacogenomics Jong W. Lee, Britt I. Drögemöller, Galen E.B. Wright, Shahrad R. Rassekh, Yuling Li, Erandika P. Gunaretnam, Beth Brooks, Ghazal Babolmorad, David D. Eisenstat, W. Ted Allison, Bruce C. Carleton, Colin J.D. Ross and Amit P. Bhavsar Cisplatin is a highly effective chemotherapy treatment for multiple solid tumour cancers. Cisplatin use is limited by the development of permanent high-frequency sensorineural hearing loss – cisplatin-induced ototoxicity (CIO). CIO can have significant life-long consequences, especially for children, by impairing speech, language and social-emotional development. Severe CIO can also cause discontinuation of this effective chemotherapeutic. CIO is estimated to occur in >50% of treated children with susceptibility influenced by genetic variation.

We recently conducted a case-control genome-wide association study in cisplatin-treated Canadian pediatric cancer patients that analyzed 640,000 variants. The most significant signal was identified on chromosome 9q33.2 (P=8.36x10-7, odds ratio=0.14, 95% CI=0.07-0.34) and was associated with protection from CIO. The hypothesis of this research is that an innate immune gene situated at 9q33.2 contributes to the development of CIO in children. Objective 1 is to validate this discovery and develop a genomic biomarker that increases the accuracy of CIO risk prediction. Objective 2 is to develop cisplatin otoprotectants by conducting proof- of-principle in vitro experiments perturbing specific innate immune pathways before advancing to pre-clinical animal models.

An independent patient cohort has been recruited from across Canada and subset analyses showed preliminary replication of the 9q33.2 associations (P<.01). In vitro cisplatin

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transcriptional response assays demonstrated a putative interaction between cisplatin and the innate immune gene that is altered by genetic variation. Both small molecule and genetic inhibition of Tlr4 reduced cisplatin-induced inflammatory responses in a murine inner ear cell line. These data provide preliminary mechanistic insight into the pharmacogenomic association with ototoxicity that furthers our understanding of CIO development. They also provide preliminary proof-of-principle for targeted inhibition of TLR4 to mitigate this severe drug toxicity.

63 Mohammad Refaei Medicine and Dentistry Hematology Utility of Leukemic Stem Cells Markers in Determining Minimal Residual Disease in Patients with AML Mohammad Refaei 1, Artur Szkotak2, Irwindeep Sandhu1, Joseph Brandwein1, Lalit Saini 1. 1. Division of Hematology, University of Alberta, Edmonton, Canada 2. Division of Laboratory Medicine and Pathology, University of Alberta, Edmonton, Canada. Introduction: Minimal residual disease (MRD) assessments are being explored to define prognosis in patients with acute myeloid leukemia (AML). Relapses are thought to be related to the persistence of leukemia stem cells (LSC) that survive chemotherapy. A number of LSC antigens have been identified such as CLL1, and CD123. We hypothesized that the use of a routine antibody panel in conjunction with LSC markers will allow for more accurate determination of MRD using (MPFC) and patients destined to relapse. Methods: In this single center prospective study we evaluated the diagnostic bone marrow samples of all newly diagnosed patients with AML (excluding APL) for leukemia associated immunophenotypes (LAIP’s) using multiparametric flow cytometry (MPFC) and a panel of antibodies targeted towards myeloid, lymphoid and LSC markers. Samples were considered MRD positive if having >0.1% of a specific LAIP as a function of the bulk blast population. Results: Over the study duration, 88 patients with newly diagnosed AML received intensive

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chemotherapy. Complete MRD data was available for 58 patients had at the end of induction (EOI) and 31 patients at the end consolidation (EOC). Following a single induction chemotherapy cycle 52/58 (89.7%) patients were in CR. At the EOI, 26/58 (44.8%) patients were found to be MRD negative using the standard LAIP. Only 3/30 patients (10%) who had CD123+ LSC and 2/28 (7.1%) patients who had CLL1+ LSC at diagnosis were found to be negative at the EOI. However, 21/24 (87.5%) patients that were negative for CD123+ LSC and 22/24 (85%) patients that were negative for CLL1+ LSC at diagnosis were found to be positive for CD123+ LSC and CLL1+ LSC at the EOI, respectively. Although not statistically significant, we observed that only 1 of 6 (16.7%) patients that was negative for CLL1+ LSC at the EOC relapsed in contrast to 11 of 25 (44%) patients that were positive for CLL1+ LSC (p=0.4). After a median follow-up of 11.3 months, the median relapse free survival for patients that were positive for CLL1+ LSC at the EOC was 13.8 months and was not achieved for patients negative for CLL1+ LSC (p=0.4). There was no significant difference in the OS for patients based upon MRD status at the EOC when using standard MRD markers, CLL1 or CD123. Conclusions: This proof-of-concept study suggests that the CLL1+ and CD123+ LSC can be identified at diagnosis at very low levels relative to the bulk blast population. Surprisingly, patients who were negative for CD123+ and CLL1+ LSC at diagnosis were often found to be positive at EOI. This may be related to a reduction in the overall blast population due to chemotherapy, but persistence of the original LSC population.

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64 Jingjie Xiao ALES AFNS CT-assessed Muscle Abnormalities and Surgical Outcomes among Non-metastatic Colon Cancer Patients Jingjie Xiao, MSc, Bette J. Caan, DrPH, Peter D. Peng, MD, Erin Weltzien, Elizabeth M. Cespedes Feliciano, ScD, MSc, Candyce H. Kroenke, ScD, MPH, Jeffrey A. Meyerhardt, MD, MPH, Vickie E. Baracos, PhD, Marilyn L. Kwan, PhD, Adrienne L. Castillo, RD, Carla M. Prado, PhD, RD Introduction: Despite advanced operative techniques for resection surgery, a large number of patients with colon cancer still suffer from post-operative morbidity. This leads to delayed subsequent therapy, prolonged hospital stay, and potentially worse prognosis. Computerized tomography (CT)-measured muscle abnormalities are of emerging interest to surgeons due to their potential value for post-operative risk stratification. This study investigated the independent prognostic effects of muscle abnormalities on surgical outcomes. Methods: Patients diagnosed with stage I-III colon cancer from 2006 to 2011 were included (n=1,715). Muscle abnormalities were defined as a low skeletal muscle index (SMI) and/or radiodensity (SMD) measured on pre-operative CT images. Differences in demographic, clinical and surgical outcomes were compared by each muscle abnormality using independent t-tests or Chi2 tests. Logistic regression was used to evaluate the associations of muscle abnormalities with post-surgical length of hospitalization, any complication(s) and readmission up to 30 days post-surgery or post-discharge. Cox proportional hazards regression was performed to examine the effect of muscle abnormalities on 30-day mortality. Results: Mean age was 64.0 ±11.2 years and 55.5% were females. Patients with low SMI (OR=1.37, 95% CI 1.10-1.72) or low SMD (OR=1.50, 95% CI 1.15-1.96) were more likely to remain hospitalized ≥7 days after surgery, and had higher risk of 30-day mortality (low SMI: HR=4.65, 95% CI 1.43-15.06; low SMD: HR=3.34, 95% CI 1.08-10.30). Additionally, patients with low SMI were more likely to have at least one post-surgical complication (OR=1.26, 95%

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CI 1.02-1.55). Readmission rate was not associated with any muscle abnormality. Conclusion: Muscle abnormalities were associated with poorer surgical outcomes, including longer hospitalization and a higher risk of short-term mortality. Low SMI was associated with a higher risk of post-surgical complications. Research should evaluate whether targeting potentially modifiable factors preoperatively, such as increasing muscle mass and/or radiodensity may improve post-operative outcomes.

65 Jingjie Xiao ALES AFNS Associations of Pre-existing Comorbidities with Skeletal Muscle Mass and Radiodensity in Early Stage Colorectal Cancer Patients Jingjie Xiao, MSc, Bette J. Caan, DrPH, Erin Weltzien, Elizabeth M. Cespedes Feliciano, ScD, MSc, Candyce H. Kroenke, ScD, MPH, Jeffrey A. Meyerhardt, MD, MPH, Vickie E. Baracos, PhD, Marilyn L. Kwan, PhD, Adrienne L. Castillo, RD, Carla M. Prado, PhD, RD Background: Comorbidities and computerized tomography (CT)-measured muscle abnormalities are both common in cancer patients and evident in diseases such as diabetes and obesity. This is the first study to examine the association between comorbidities and muscle abnormalities in patients with colorectal cancer (CRC). Methods: This cross-sectional study included 3051 patients with stage I-III CRC. Muscle abnormalities were defined as low skeletal muscle mass index (SMI) or low skeletal muscle radiodensity (SMD) quantified using diagnostic CT images. Charlson comorbidities were ascertained. Chi-square tests were used to compare the prevalence of comorbidities by the presence/absence of each muscle abnormality. Logistic regression analyses were performed to evaluate which comorbidities predicted muscle abnormalities adjusting for age, sex, body mass index, weight change, stage, race/ethnicity, and smoking. Results: Mean age was 63 years; 50% of patients were male. The prevalence of low SMI and low SMD were 43.1% and 30.2%, respectively. Comorbidities examined were more prevalent

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in patients with low SMD than in those with normal SMD (58.6% vs. 34.4%, p<0.001), and most remained significant predictors of low SMD, including myocardial infarction (odds ratio [OR]=1.82, p=0.017), congestive heart failure (OR=3.34, p<0.001), peripheral vascular disease (OR=2.20, p<0.001), diabetes with or without complications (OR=1.61, p=0.008; OR=1.47, p=0.003, respectively) and renal disease (OR=2.23, p<0.001). No comorbidities were associated with low SMI except for diabetes with complication which was associated with a lower likelihood of low SMI (OR=0.64, p=0.007). Conclusion: Prevalence of muscle abnormalities was high in early stage CRC. Pre-existing comorbidities were most commonly associated with low SMD, suggestive of a potential shared mechanism between fat infiltration into muscle and each of these comorbidities.

66 Amritpal Bhullar Faculty of Graduate Studies and Research Department of Agricultural, Food and Nutritional Science Muscle radiodensity negatively correlates with triglyceride content in skeletal muscle of cancer patients Amritpal S. Bhullar, Ana Anoveros-Barrera, Abha Dunichand-Hoedl, Karen Martins, David Bigam, Todd McMullen, Charles T. Putman, Vickie Baracos and Vera Mazurak Background- Computed Tomography (CT) cross-sectional imaging has been recently applied to evaluate muscle area and radiodensity in cancer patients. Studies of muscle radiodensity, reported in Hounsfield Units (HU)1, reveals an association with poor survival in a variety of cancer types2-5. In healthy adults, muscle radiodensity inversely associates with triglyceride content6 but no studies to date have reported this association in cancer patients. It is also not known whether lipid is located inside myocytes as lipid droplets or adjacent to the myocytes within adipocytes.

Methods- Rectus abdominis (RA) muscle biopsies were collected during surgery from patients diagnosed with gastrointestinal cancers. Skeletal muscle radiodensity was assessed by

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analyzing CT images at the 3rd lumbar vertebra, as well as RA specifically. Triglyceride content of muscle was analysed quantitatively using gas chromatography. RA muscles, frozen for histology, were stained for neutral lipid content using Oil Red O (ORO) to determine location of neutral lipids. Percent area of pixels with ORO stain was calculated by Volocity6.3 Software. Results- Mean muscle radiodensity and RA radiodensity were inversely associated with triglyceride content (µg/mg) (r= -0.518, p=0.023 and r= -0.481, p=0.032, respectively). Of the total percent area of neutral lipid, mean percent area present outside myocytes was 54% compared to 46% mean percent area inside myocytes. There was no significant difference in percent area of neutral lipids present inside (42% vs 54%, p=0.309) and outside (58% vs 46%, p=0.309) myocytes in muscle with low versus high triglyceride content, respectively. Conclusion- In cancer patients, overall abdominal muscle and rectus abdominis radiodensity is associated with triglyceride content of the muscle indicating that low muscle radiodensity is indicative of fatty infiltration, which appears to occur equally inside and outside of the myocyte. Future studies will enable us to understand effects of high triglyceride content within muscle of cancer patients.

67 Wan Kong Yip Medicine & Dentistry Medical Microbiology & Immunology Mutation in Reovirus mu2 Reduce Specific Infectivity but Promote Replication and Dissemination in Tumorigenic Cells Wan Kong Yip and Maya Shmulevitz The non-pathogenic wild-type serotype 3 mammalian orthoreovirus (Reo-wt) is one of the most evaluated oncolytic virus in clinical trials. We identified a variant (Reo-10M) with a single amino acid change in the mu2 protein that shows improved replication over Reo-wt specific to cancer cells. Our data demonstrates that the mutation in Reo-10M mu2 has both infection-hindering and –promoting effects, with an overall net benefit for replication in

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cancer cells. First, we found that despite similar binding to cells and uncoating, Reo-10M established infection in 50% fewer cells relative to Reo-wt as determined by flow cytometric analysis. Mu2 is a polymerase co-factor, and our in vitro phosphate release and qRT-PCR assays showed that Reo-10M mu2 was less efficient at hydrolyzing ribonucleotide triphosphates (rNTP) and RNA synthesis, likely explaining the reduced establishment of infection. However, when equivalent Reo-wt or Reo-10M infected cells were analyzed by Western blot analysis and qRT-PCR, Reo-10M accumulated up to 4-fold more viral proteins and 3-fold more viral RNAs relative to the Reo-wt virus. In addition to its role in transcription, mu2 also participates in creation of viral factories; localized areas of virus amplification and progeny assembly. Interestingly, immunoprecipitation of mu2 and Western blot analysis for other viral proteins showed that Reo-10M mu2 associated with uNS and other viral proteins 3-fold more efficiently than wt mu2. In conclusion, the mu2 mutation in Reo-10M reduces the efficiency of rNTP hydrolysis and RNA synthesis thereby reducing establishment of infection. However, the mutation increases the association of mu2 with factory components, augments levels of virus RNA, proteins and progeny, and ultimately promotes virus replication in cancer cells.

68 Reegan Willms Science Biological Sciences Myt1 Kinase is Required for Stem Cell Homeostasis in the Drosophila Intestine Reegan J. Willms & Shelagh D. Campbell Cdk1 is the master mitotic kinase responsible for propelling cells into mitosis. To prevent Cdk1 from driving cells into M phase prematurely, the Wee-like kinases Wee1 and Myt1 add inhibitory phosphates during interphase. Though Wee1 is often considered the more relevant of the Wee-like kinases, work in mammalian cell culture and in Drosophila has demonstrated the importance of Myt1 in development and endomembrane remodeling [1-4]. Furthermore,

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a recent paper identifying Myt1 as an essential regulator of glioblastoma stem-like cells points to the necessity of better understanding how Myt1 influences cell cycle regulation [5].

Previous studies from our lab focused on dMyt1 function in sensory organ development and male meiosis. Recently we identified a requirement for Myt1 in maintaining adult intestinal stem cell (ISC) homeostasis. The Drosophila intestine is an excellent model system for studying this process with well-defined similarities to the mammalian intestine regarding cell types, morphology, and signaling pathways. A combination of immunolabeling and microscopy showed that the mitotic index of ISCs in the guts of 7 day old myt1 null mutants was approximately 50-fold that observed in wild-type controls. EdU labeling used to analyze the rate of turnover in the gut indicated that this increased mitotic index corresponded to hyperproliferation leading to increased cellular density. Additionally, myt1 mutant guts exhibit disrupted visceral muscle tissue, the consequences of which are not yet fully understood. RNAi depletion of Myt1 in the adult ISCs via a temperature sensitive UAS-Gal4 system phenocopied the mutant phenotype, indicating that the increased mitotic index of myt1 mutants was not due to disruption of the developmental program or to loss of Myt1 in other cell types. Further work is currently being done to determine the mechanism by which loss of Myt1 causes ISC hyperproliferation and how this affects other cell types within the intestine.

References

1. R. Varadarajan et al., MBoC 27, 2051 (2016). 2. Z. Jin, et al., Development 132, 4075 (2005). 3. Z. Jin et al., Genetics 180, 2123 (2008). 4. H. Nakajima et al., J. Cell Biol 181, 89 (2008). 5. C. M. Toledo et al., Cell Rep 13, 2425 (2015).

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69 Bahareh Hamedi Medicine & Dentistry Oncology n-3 LONG CHAIN POLYUNSATURATED FATTY ACIDS (LCPUFA): THE ANTICANCER EFFECTS ON OVARIAN CANCER CELLS IN VITRO Bahareh Hamedi1*, Olena Bilyk1, Marnie Newell2, Susan Goruk2, Catherine Field2, Lynne- Marie Postovit1,3 1Department of Oncology, University of Alberta 2Department of Agricultural, Food and Nutrition Science, University of Alberta 3Department of Obstetrics and Gynecology, and Women’s and Children’s Health Research Institute, University of Alberta Objective: As most ovarian cancer (OC) patients relapse with chemoresistant disease, clearly further advances in cancer research are needed to improve treatment and extend the lives of the women affected with this disease. There is a strong rationale for studying nutritional interventions in cancer prevention and therapy. In this study, we evaluate the effect of n-3 LCPUFA on OC cell growth and sensitivity to carboplatin.

Methods: In vitro assays designed to assess growth and viability (metabolic MTT assay, flow cytometry) in 6 OC cell lines of different histological type and normal ovarian and fallopian tube cells after treatment with concentrations of DHA (Docosahexaenoic Acid) and OA (Oleic Acid used as control) that would be achievable in blood serum by dietary intervention (10- 320 uM).

Results: The effect of DHA seems to be type-specific to malignant ovarian cells. The inhibitory effect of DHA on the growth and viability was demonstrated in clear-cell carcinoma cell line ES2 while OC cells of other histological types were not sensitive to DHA. Pre- treatment with DHA increased the sensitivity of resistant endometrioid cell line A2780cp as well as ES2 and SKOV3 cells to carboplatin. Conclusion: The in vitro evidence for DHA efficacy on ovarian clear-cell carcinoma cell line growth was demonstrated. DHA might increase the sensitivity of OC cells to platinum-based

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chemotherapy. Further in vitro and in vivo studies are necessary to provide a strong rationale for a targeted n-3 PUFA intervention to be considered as an adjuvant to antineoplastic drug therapy of OC.

70 Sams Sadat Pharmacy and Pharmaceutical Sciences Pharmaceutical Sciences Abstract covered under NDA until November 25th.

71 Rong-Zong Liu Medicine & Dentistry Oncology NFIB promotes cell survival by directly suppressing p21 transcription in triple-negative breast cancer Rong-Zong Liu, The M. Vo, Saket Jain, Elizabeth A. Monckton, Elizabeth Garcia, John R. Mackey and Roseline Godbout Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with limited treatment options and poor prognosis. There is an urgent need to identify and understand the key factors and signaling pathways driving TNBC tumor progression, relapse and treatment resistance. In this study, we report that gene copy numbers and expression levels of Nuclear Factor IB (NFIB), a recently identified oncogene in small cell lung cancer, are preferentially increased in TNBC compared to other breast cancer subtypes. Furthermore, increased levels of NFIB are significantly associated with high tumor grade, poor prognosis and reduced chemotherapy response. Concurrent p53 mutations and NFIB overexpression (z scores>0) were observed in 77.9% of TNBC, in contrast to 28.5% in non-TNBC. Depletion of NFIB in p53-mutated TNBC cell lines promotes cell death, cell cycle arrest and enhances sensitivity to docetaxel, a first line chemotherapeutic drug in breast cancer treatment.

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Importantly, these alterations in growth properties were accompanied by induction of CDKN1A, the gene encoding p21, a downstream effector of p53. We show that NFIB directly interacts with the CDKN1A promoter in TNBC cells. Furthermore, knockdown of both p21 and NFIB reverses the sensitivity of TNBC cells to docetaxel observed upon NFIB knockdown, indicating that NFIB’s effect on chemotherapeutic drug response is through p21. Our results indicate that NFIB is an important TNBC factor that drives tumor cell growth and drug resistance, leading to poor clinical outcomes. Thus, targeting NFIB in p53-mutated TNBC may reverse oncogenic properties associated with mutant p53 by restoring p21 activity.

72 Geetha Menon No volumetric preplan: institutional experience with an intracavitary/interstitial MR-based cervix implant strategy Deepak Dinakaran, Ericka Wiebe, Geetha Menon, Michele Janoski, Janet Zimmer, Robert Pearcey, Fleur Huang PURPOSE Modern imaging and hybrid intracavitary(IC)/interstitial (IS) applicators enable optimal BT relative to target and normal tissue topography in cervix cancer. In well-studied protocols, a dry-run procedure and MRI (IC applicator in situ) with volumetric preplan (VPP) (virtual IS sources) precede actual IC/IS implant, planning, and delivery. We describe our institutional MR-based IC/IS practice: an implant strategy for IS catheters (number, location, depth) without VPP. METHODS An IC/IS non-VPP cervix BT workflow is described, along with evaluation criteria applied to pre-procedure imaging core to our implant strategy. Implant decision-making parameters are reviewed, based on initial clinical/technical experience. RESULTS A dedicated pelvic 1.5T MRI (no BT applicator) is obtained 1–10 days prior to ultrasound-

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guided IC/IS implant under anesthesia. An implant strategy based on vaginal examination and careful imaging review (initial pelvic MRI; CTsim; pre-BT MRI) is devised, acknowledging BT applicator geometry. Post-implant, CTsim (applicator check; IS reconstruction) and MRI (volumetric BT planning) ensue, for same-day BT start. In strategizing procedural parameters likely to achieve optimum target coverage while minding organs at risk, an IC applicator is selected (configuration, size), given the site/extent of tumor residuum, spatial capacity of the vaginal canal, uterine height, and utero-vaginal angle for a given bladder volume. An IS component is inferred via measurements on pre-BT MRI (T2 sagittal, short axis oblique (SA), coronal). After situating the external os, the outer perimeter of the ring/ovoids is estimated, for basic visual mapping of IS radial positions against what are often asymmetric tumors. Along the anticipated axis of insertion, depths are taken for minimum target coverage without transgressing adjacent vital structures. Since 2013, n=45 received pulsed dose rate BT boost with IC/IS technique for cervix cancer. For n=43 single-fraction cases (FIGO stage IB1 – IVA; median HR-CTV volume 33.0 cc [11.1 – 119.0]), ring or ovoid-based, the median number of implanted (depth 2.6 cm [0.8 – 4.7]) and loaded IS catheters were 3 [1 – 6] and 2 [1 – 5], respectively. 18% of non-VPP placed catheters were not loaded. Dose-volume goals were reached: median HR-CTV D90 (EQD2) was 90.7 Gy [76.4 – 96.6], bladder D2cc 75.8 Gy [43.2 – 90.4], rectum D2cc 64.3 Gy [52.3 – 74.2], and sigmoid D2cc 62.2 Gy [46.6 – 73.6]. Challenges remain with a non-VPP approach, which cannot predict for reversibility of rotated/retroflexed uterine lie, risking misjudged IS positions/depths. 1/43 had OAR intrusion (within posterior bladder wall). CONCLUSIONS MR-based IC/IS cervix BT promises better clinical outcomes, but remains resource intensive and still widely prohibitive. The ability to achieve excellent tumor-adapted implant conformation without extra imaging, implant or planning steps can ease access to high quality MR-BT in resource-minded settings.

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73 Lena Bilyk Medicine and Dentistry Oncology Nodal-induced gene reprogramming as a mode of cancer cell plasticity and drug resistance in ovarian cancer cells Lena Bilyk, Laura Lee, Jiahui Lee, Scott Findlay, Martin Coebel, Lynne Postovit Cancer cells can exploit normally dormant embryonic stem cell pathways to promote cancer progression and metastasis. Understanding the impact of these embryonic signals and the mechanisms that reactivate them has significant potential for new cancer therapies. Studying embryonic signaling pathways in aggressive cancer has led to the discovery of the re-expression of the embryonic protein Nodal. Nodal belongs to the TGF-β superfamily of secreted signalling factors, maintains pluripotency and cell plasticity of human embryonic stem cells. We hypothesize that Nodal signaling drives plasticity in OC cells and that disease progression and therapy resistance will be mitigated when Nodal is inhibited. We applied approaches for manipulation of gene expression, series of in vitro assays, clinical data, bioinformatics approach and RNA sequencing to explore the impact of Nodal on cancer cell plasticity, resistance to cytotoxic drugs, biological processes in OC cells and survival of patients with OC. We discovered that Nodal induces transcriptional reprogramming in OC cells via altering immune response, metabolism and drug resistance gene expression. In vitro, we showed that OC cells overexpressing Nodal characterized by increased resistance to cytostatic drugs, cell plasticity, partial EMT transition and tumorigenicity. Analysis of TCGA microarray data, IHC of OC tissue microarrays and clinical data revealed significant association of Nodal expression with the most aggressive type of OC - high grade serous carcinoma and poor overall and progression-free survival of patients. This multi-approach study has discovered a potential biomarker Nodal for OC therapy

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resistance and progression which may hold promise as a therapeutic target to prevent recurrence of the most aggressive type of OC. 74 Mahshad Moshari Science chemistry Novel Colchicine Derivatives Mahshad Moshari, Dr. Jack Tuszynski,Dr. Mariusz Klobukowski Microtubules are one of the major components of the cytoskeleton in eukaryotic cells. These filamentous intracellular structures play a critical role in maintaining cell structure, mitosis and cell division. They are polymers formed from the heterodimers of α and β tubulins. Microtubules are one of the most well-known chemotherapeutic targets in cancer treatment. Depolymerization of microtubules can be triggered by inhibition of β tubulin via binding to a chemotropic agent such as colchicine or taxol. There are some limitations of in vitro approaches for testing chemotherapeutic agents such as complex synthesis methods and compound solubility. Computational methods are reasonable detours to overcome these problems. In this research, a novel library of colchicine derivatives was docked into the βІ tubulin colchicine binding site. We used a combination of different theoretical methods for exploring ligand-tubulin interactions. The ligands structures were first minimized then fully optimized based on the RHF/cc-pVDZ level of theory in GAMESS-US. Since there is no crystal structure for βІ tubulin (TBB5_HUMAN), we obtained its sequence from UniProt (ID: Q13509). We used the tubulin structure 1SA0.pdb as a template to construct the homology model for βІ Tubulin using MOE. We then docked the colchicine library to the protein using Autodock4 program under flexible ligand and rigid receptor conditions. After simulating the ligand-tubulin complexes, for each compound, using molecular dynamics, we calculated the binding free energies of the different ligands to tubulin. In addition, our collaborators provided us with the IC50 values for the compounds from cell-based assays. To have a more realistic correlation coefficient, the

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MLogP of the compounds were calculated using ADMET Predictor 8.0. The linear regression with two independent variables, calculated binding free energies and MLogP, of our dataset showed a good correlation of 0.7 with the experimental IC50 values.

75 Ahmed Elmenoufy Science Chemistry Novel ERCC1-XPF Inhibitors for Resistant Colorectal Cancer Therapy based on Targeting DNA Repair Pathway Ahmed H. Elmenoufy, Francesco Gentile, Feridoun K. Busheri, Olivier M. Soueidan, David Jay, Claudia Weilbeer, Rajam S. Mani, Jack. A. Tuszynski, Michael Weinfeld and Frederick. G. West ERCC1-XPF heterodimer is a 5ʹ-3ʹ structure-specific endonuclease which is essential in multiple DNA repair pathways in mammalian cells. ERCC1-XPF repairs chemically-induced helix-distorting and bulky DNA lesions caused by UV irradiation by removal of cyclobutane pyrimidine dimers (CPDs) result from photoreaction between two adjacent thymine bases. Specific inhibition of ERCC1-XPF has been shown to potentiate radiotherapy cytotoxicity in cancer cells. In this study, the previously described ERCC1-XPF inhibitor F06 was used as a reference compound. Using docking-based virtual screening, we synthesized 7 derivatives of F06 that were identified in silico as being likely to have high binding affinity for the ERCC1- XPF heterodimerization interface. Exploration of the structure-activity relationships (SAR) within the synthesized derivatives yielded compounds 3 and 4 (the structures cannot be disclosed at this time for IP reasons) as potent inhibitors of ERCC1-XPF activity. In vitro ERCC1-XPF endonuclease assay implicated compound 4 as the best ERCC1-XPF inhibitor with an IC50 of 0.3 µM. In addition, Compound 4 has a high binding affinity towards ERCC1-XPF with an experimental Kd of 100 nM determined by monitoring tryptophan fluorescence quenching. Interestingly, compound 4 showed a significant inhibition of the removal of CPDs compared with control cells after UV irradiation of HCT 116 cells. Based on these preliminary results, we hypothesize that compound 4 will be able to sensitize colorectal cancer cells to

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radiotherapy. In further investigations, we are working on the development of new generation compounds based on the structure of compound 4 to improve the inhibitory properties against ERCC1-XPF activity.

76 Logan Fairgrieve-Park Medicine & Dentistry Oncology, Division of Medical Physics Optimization of Magnetic Resonance Spectroscopy Techniques for Quantifying Fat Glycerol Protons Logan Fairgrieve-Park, Clara Fallone, Atiyah Yahya

Magnetic resonance spectroscopy (MRS) is a non-invasive method for fat (lipid) quantification in vivo. In lipid MRS, certain peaks in the spectrum correspond to different protons (1H nuclei) of the molecule. In triglycerides , the lipid glycerol peak (4.3 ppm) can be employed as a reference signal for normalizing the lipid olefinic (5.4 ppm) and diallylic (2.8 ppm) signals allowing for the estimation of relative fat unsaturation, a measure shown to be relevant in cancer. In this study, the response of the glycerol protons were characterized for two MRS in-vivo localization techniques, PRESS (Point RESolved Spectroscopy) and STEAM (Stimulated Echo Acquisition Mode), at 3 T. Spectra were acquired for 9 different oils as a function of PRESS and STEAM echo time (TE), a timing parameter. The scalar coupling interactions of the glycerol protons cause signal modulation with TE. Additionally, water signal in-vivo at overlaps the glycerol signal rendering its quantification challenging. Spectra were acquired over a range of TE values to determine which resulted in minimal scalar coupling evolution loss and water signal interference. The optimized TE values were found to be 180 msec for PRESS and 100 msec for STEAM sequence. Olefinic/glycerol and diallylic/glycerol peak area ratios at the determined TEs were plotted against ratios obtained from higher resolution 700MHz (16.5T) NMR. Linear coefficients of determination were calculated to be 0.79 and 0.89 for PRESS and 0.90 and 0.93 for STEAM, respectively. To verify

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detection of the glycerol signal in-vivo, spectra were acquired from tibial bone marrow of 4 volunteers. It was verified that the water signal had decayed at the optimal TE values.

77 Won Shik (Daniel) Choi Medicine & Dentistry Oncology Overcoming Retinoic Acid Resistance in HER2-enriched Breast Cancer: Role of MYC Won Shik (Daniel) Choi, Rong-Zong Liu and Roseline Godbout Breast cancers are heterogeneous tumours with distinct morphological features and clinical behaviors, making it difficult to predict response to treatment. Breast cancers are divided into four main subtypes based on molecular signatures: luminal A, luminal B, HER2-enriched, and triple-negative. Of all breast cancer subtypes, HER2-enriched cancers have the second poorest prognosis and constitute 15-20% of breast cancers. The development of trastuzumab has greatly improved the clinical outcome for HER2-positive patients; however, intrinsic and acquired resistance to trastuzumab is common. Thus, additional therapeutic interventions are required. Retinoic acid (RA), the active metabolite of vitamin A, is a well-characterized cell differentiation agent. RA has been successfully used in the treatment of acute promyelocytic leukemia. RA signaling is mainly mediated through nuclear transcription factors (RARs and RXRs). Retinoic acid receptor alpha (RARA), a key determinant of RA sensitivity and cell differentiation, is frequently co-amplified with ERBB2, the gene encoding HER2. Surprisingly, HER2-enriched breast cancers are refractory to RA treatment despite having elevated levels of RARa. The mechanism underlying RA resistance in HER2-enriched breast cancer cells remains unclear. The proto-oncogene c-MYC is preferentially amplified and overexpressed in HER2-enriched breast cancers and is associated with a poor prognosis. c-MYC binds RARa, thereby suppressing RA-responsive genes and differentiation. Furthermore, c-MYC upregulates the RA-binding protein gene FABP5. FABP5 facilitates RA cellular trafficking to the PPARb nuclear receptor rather than RARs, thereby promoting PPARb mediated proliferation. We hypothesize that c-MYC attenuates RA action in HER2-

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enriched breast cancer cells by blocking RARa activity and activating the FABP5-PPARb pathway. Our goals are to: i) examine the expression of c-MYC and RA target genes in HER2- enriched breast cancer cells, (ii) determine how c-MYC affects RA-induced cell differentiation and proliferation, and (iii) identify the mechanism underlying c-MYC-mediated inhibition of RA action.

78 Mahsa Alaee Medicine and Dentistry Oncology Plakoglobin Restores Mutant p53 Tumor Suppressor Functions by Sequestering Beta-catenin Mahsa Alaee, Manijeh Pasdar p53 is a tumor suppressor and transcription factor that regulates cell cycle arrest, DNA repair, apoptosis, autophagy and metabolism. p53 is mutated in over 50% of all cancers. Mutant p53 (mp53) proteins not only lose their wild-type functions but they may gain oncogenic functions (GOF). One of the most frequently expressed GOF mp53s is p53-Arg175His (p53- R175H) with well documented roles in tumorigenesis and metastasis. Plakoglobin is a dual cell adhesion and signalling protein and a paralog of beta-catenin. However, beta-catenin has well-known oncogenic function via its role in the Wnt signalling pathway whereas plakoglobin acts as a tumor/metastasis suppressor. We have shown that plakoglobin interacted with both wild type and several mp53s in various carcinoma cell lines of different origins. plakoglobin and p53 interacted with the promoter and regulated the expression of several p53 target genes. Furthermore, plakoglobin interactions with mp53s restored the wild- type tumors suppressor activities in these mutants. GOF mp53s induce the aberrant accumulation and oncogenic activation of beta-catenin and we have shown that one mechanism by which plakoglobin acts as a tumor suppressor is by sequestration of beta-catenin’s oncogenic function. Here, we examined the effects of p53- R175H expression on beta-catenin accumulation and transcriptional activation and assessed how plakoglobin expression modified the beta-catenin-mediated oncogenic function of this

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mutant. Our results showed that p53-R175H expression in plakoglobin null cells increased total and nuclear levels of beta-catenin, its transcriptional activity and the in vitro migration and invasion of the transfectants. Co-expression of plakoglobin in p53-R175H cells promoted beta-catenin’s proteasomal degradation, significantly decreased its total and nuclear levels and its transactivation activity. beta-catenin target genes, c-Myc and S100A4 were upregulated in p53-R175H expressing cells and were significantly downregulated when plakoglobin was co-expressed. The plakoglobin expressing p53-R175H cells also showed significant reductions in their in vitro migration and invasion.

79 Sarah Purcell ALES AFNS Predicting Resting Energy Expenditure in Advanced Colorectal Cancer using Body Composition Sarah A. Purcell, Vickie E. Baracos, Jessica R. Lieffers, Sunita Ghosh, Marina Mourtzakis, Carla M. Prado Introduction: Resting energy expenditure (REE) is variable in cancer and influences energy needs. We assessed the agreement between measured REE (mREE) and predicted REE (pREE) using anthropometric/demographic variables and skeletal muscle and total adipose cross-sectional area (CSA, cm2).

Methods: Patients with advanced colorectal cancer were recruited from the Cross Cancer Institute (Edmonton, Canada). REE was measured by indirect calorimetry. CSAs were quantified via CT images at the 3rd lumbar vertebrae using Slice-O-Matic, V4.3. Multiple linear regression identified the contribution of age, sex, height, weight, skeletal muscle and total adipose tissue CSAs to REE and generated equations using significant (p<0.05) variables. Accuracy of new equations was compared to the Harris-Benedict equation. mREE was compared to predicted using Lin’s concordance and Bland-Altman analyses.

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Results: 24 patients were included (BMI 26.1±5.3kg/m2; age 59±13y); mean mREE was 1502 ± 306 kcal/day. Average pREE from the Harris-Benedict equation was 1542 ± 312 kcal/day; bias = 41 kcal/day; limits of agreement = -212 to 293 kcal/day. Linear regression Model 1 included age, sex, height, and body weight; age (p<0.001), height (p=0.001), and weight (p<0.001) were predictors of REE (p<0.001 R2=0.870, SEE=118kcal/day). Mean pREE from this model was 1502 ± 285 kcal/day; bias = 0 kcal/day; limits of agreement = 216 to 216 kcal/day. In Model 2, skeletal muscle and total adipose tissue CSA were added. Skeletal muscle (p=0.006), age (p<0.001), weight (p=0.009), and height (p=0.014) were predictors of REE (p<0.001, R2=0.914, SEE=99 kcal/day). Mean pREE was 1505 ± 290kcal/day; bias = 3 kcal/day; limits of agreement = -186 to 173kcal/day. Lin’s concordance coefficient was highest for Model 2 equation with a “substantial” strength of agreement (ρc=0.958). Model 1 (ρc=0.9304) and Harris-Benedict (ρc=0.905) equations had "moderate" agreement.

Conclusion: Skeletal muscle from CT images is a better predictor of REE on an individual and group level compared to anthropometric/demographic data.

80 Sarah Purcell ALES AFNS Whole Body Calorimetry Use in Cancer Patients: A Case Study of Agreement between Measured and Predicted Energy Needs Sarah Purcell, Ilana Roitman, Claire F. Trottier, Dr. Carla M. Prado INTRODUCTION Inaccurate estimation of energy needs places cancer patients at risk for weight loss, malnutrition and cachexia. Muscle mass is likely a major driver of changes in energy requirements during cancer, but no study has investigated this. Here, we compared measured resting energy expenditure (REE) and total energy expenditure (TEE) assessed

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using a whole body calorimetry unit (WBCU) to values derived using predictive equations in two individuals with colorectal cancer, one with, one without sarcopenia.

METHODS: Patients were recruited at the Cross Cancer Institute (AB, Canada), as part of an ongoing randomized controlled trial. Body composition was assessed by dual energy X-ray absorptiometry (iDXA). Sarcopenia was defined by previously established cutpoints of low appendicular muscle mass. Patients stayed in the WBCU for 24 hours, where REE and TEE were measured. Four equations were used to predict REE. A stress factor (SF) of 1.3 and a thermic effect of food (TEF) of 1.1 were considered for the estimation of TEE. Energy intake guidelines for individuals with cancer (25-30 kcal/kg body weight/day) were compared with TEE.

RESULTS: Patient 1 had a normal body weight and was sarcopenic; patient 2 had obesity but not sarcopenia. Most equations underestimated REE for both patients. Two commonly used equations would classify patient 2 as hypometabolic. By calculating TEE values using SF and TEF, patient 1 was more likely to have lower than predicted TEE compared to patient 2. Current dietary guidelines underestimated TEE for patient 1 and overestimated for patient 2.

CONLUSION: This case study shows the accuracy of REE and TEE predictive equations is variable for individuals with cancer and affected by muscle mass. The inclusion of body composition into prediction equations may allow for more accurate estimation of energy requirements in clinical settings, benefiting cancer patients at need for individualized nutrition intervention.

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81 Bhumi Bhatt Medicine and Dentistry Laboratory Medicine and Pathology Profiling of small nucleolar RNAs in human skeletal muscle and their potential post- transcriptional regulatory role in cancer cachexia. Bhumi Bhatt1, Ashok Narasimhan1, Oliver Bathe2, Sunita Ghosh1, Vickie E. Baracos1 and Sambasivarao Damaraju1 1University of Alberta and Cross Cancer Institute; 2University of Calgary Background: Small Nucleolar RNAs (snoRNAs) are non-coding RNAs (60-300 nucleotides) involved in functions such as pre-processing and maturation of rRNAs, modifications of snRNAs, tRNAs and alternative splicing. snoRNA dysregulation has been associated with cancer and other disease conditions. snoRNAs also harbor other small RNAs such as miRNAs or piRNAs, both of which regulate gene expression by translational arrest or degradation of mRNA. However, snoRNA profiling and their effects on gene regulation in cancer cachexia have not been explored to-date. Aim: i) to profile snoRNAs in human skeletal muscle biopsies ii) to identify differentially expressed snoRNAs between cachectic and non-cachectic cancer patients iii) to identify snoRNAs harboring piRNAs and to understand their regulatory roles in skeletal muscle. Methods: Study subjects (n=42) were classified as cachectic and non-cachectic based on the international consensus diagnostic framework (high % weight loss and/or sarcopenic status and low BMI). snoRNAs were profiled using the next-generation sequencing platform. Data analysis was performed in Partek Genomics Suite. snoRNAs were annotated using Ensembl database. Raw data was filtered using cut-off of >=5 read counts in 80% samples. The dataset was RPKM normalized and log transformed. Differentially Expressed (DE) snoRNAs were identified with a fold-change >1.2 and p <0.05 (ANOVA). Results: We profiled 426 snoRNAs and 82 of these were retained after filtering. 32 snoRNAs were DE and up-regulated in cachectic cases. ~78% snoRNAs were found in the introns of protein-coding genes. We identified 10 snoRNAs harboring piRNAs; of which two DE piRNAs

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were interrogated for their hypothesized role in down-regulation of mRNAs in muscle. Target mRNAs identified were: ABCC1, GLUL, PUS7L and MTO1 playing a role in Glutamine Biosynthesis. One of the DE piRNA is validated by qRT-PCR. Conclusion: We identified snoRNAs from human skeletal muscle. Representative snoRNAs harboring piRNAs were identified as potential candidates for functional validation studies.

82 Brittany Umer Medicine and Dentistry Medical Microbiology and Immunology Reprogramming the tumour microenvironment with modified vaccinia viruses Brittany Umer, Ryan Noyce, James Lin, Megan Desaulniers, Mary Hitt, David Evans Oncolytic viruses preferentially replicate in and kill cancer cells. These viruses also promote anti-tumour immunity, classifying them as a form of immunotherapy. We are investigating the oncolytic potential of a genetically modified vaccinia virus (VACV) that selectively targets cancer cells. In breast cancer models VACV delays tumour growth and extends survival, but long-term control is not achieved. We aim to enhance the therapeutic efficacy of VACV for treating breast tumours by increasing the immunogenicity of VACV therapy, leading to improved anti-tumour immune responses. Traditional approaches to enhance the immunogenicity of oncolytic viruses involve encoding immune stimulatory molecules in the virus itself, which recruit and activate immune cells. However, it is important to consider that VACV devotes a large portion of its 200 kb genome to suppressing the immune response of its host. Our approach involves deleting virally encoded immunomodulatory components from the VACV genome. We hypothesize that removal of immunomodulatory genes from VACV will modify the tumour microenvironment and increase anti-tumour immune responses. We anticipate that this response will improve the efficacy of VACV for treating breast cancer. I have produced six mutant VACVs, each with rational gene deletions that encode proteins involved in modulating different immune pathways, such as innate immune

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activation, cytokine signaling, and apoptosis. Here, I show that deletion of virally encoded genes from the VACV genome can reprogram the tumour microenvironment in animal models of breast cancer, resulting in alterations in numbers and activation of CD 4 and CD 8 T cells, myeloid cells, and PD-1 expression. These different mutant VACVs also had statistically significant differences in tumour growth. In the future, I will further characterize the immune response elicited by these different viruses in tumour-bearing mice, with the intention of exploiting these properties to enhance the therapeutic efficacy of VACV therapy.

83 Mohamed Salla Medicine and Dentistry Biochemistry Resveratrol and derivatives: inhibitors of NFkB activity, acute intestinal inflammation and potential anti-tumor drugs Mohamed Salla , Ning Yang , Vrajesh Pandya , Khushwant S. Bhullar Yoke Fuan Wong , Robyn Losch , Joe Ou , Fahad Al dawsari , Carlos Velazquez-Martinez, Audico Thiesen , Jason Dyck , Basil Hubbard , Ing Swie Goping and Shairaz Baksh Background: Resveratrol (3,4,5-Trihydroxy-trans-stilbene) is a naturally occurring polyphenol that exhibits beneficial pleiotropic health effects. It is one of the most promising natural molecules in the prevention and treatment of chronic diseases and autoimmune disorders. One of the limitations of the use of resveratrol is its extensive metabolic processing and the consequent formation of various metabolites as resveratrol glucuronides and resveratrol sulfates. It has been estimated that around 75% of this polyphenol is excreted via feces and urine. To possibly alleviate the extensive metabolic processing, we have added, in collaboration with Dr. Carlos Velazquez-Martinez, segments of acetylsalicylic acid to resveratrol in an attempt to maintain the functional properties of both. Methods: Using an MTT proliferation assay, a dual luciferase gene reporter assay, TMRE analysis and a murine DSS- model of colitis we provide analysis of how resveratrol and its derivatives can inhibit NFκB activation, cytokine production, and growth rate of cancer cells and alleviate intestinal

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inflammation and tumor growth in vivo. Results: We identified resveratrol derivatives C3 and C11 as closely preserving resveratrol bioactivities of growth inhibition and inhibition of NFκB activation. Moreover, C3 shows more promising death inducing and AMPK activating properties, independent of sirtuins. Significance: Maintaining the promising effects of resveratrol in most therapeutic related properties and surpassing it in biological properties such as AMPK activation and cell death induction could make these derivatives more promising therapeutics. We speculate that these derivatives of resveratrol would be more metabolically stable in vivo to result in increased efficacy for treating immune disorders such as IBD and as an anti-cancer agents.

84 Daniel Nisakar Meenakshi Sundaram Faculty of Pharmacy and Pharmaceutical Sciences Faculty of Pharmacy and Pharmaceutical Sciences Reversing Drug Resistance in Chronic Myeloid Leukemia Cells with Integrin-β1 Silencing Daniel Nisakar Meenakshi Sundaram, Cezary Kucharski, Remant Bahadur KC, Hasan Uludag Introduction: Chronic myeloid leukemia (CML) is a clonal myeloproliferative disorder characterized by the fusion gene BCR-ABL, which encodes an oncoprotein containing the activated tyrosine kinase region of ABL leading to uncontrolled proliferation of a sub- population of hematopoietic cells. CML is being successfully treated with tyrosine kinase inhibitors but some patients suffer from drug resistance mediated by attachment of malignant cells to bone marrow via cell surface integrins. In this study, we focused on one such molecular target, namely integrin-beta1, by delivering a dicer substrate siRNA with the aid of lipid-modified low molecular weight polyethyleneimine (PEI) polymer to better understand the role of integrin-beta1 in drug resistant K562 CML cells. Materials and Methods: Integrin-beta1 silencing experiments were carried out through Immunostaining (cell surface levels), qPCR (mRNA levels), fibronectin-binding and colony formation assay to assess clonal proliferation. Unpaired Student’s t-test was used to assess the statistical differences between the group means (p<0.05).

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Results: The in-house designed 1.2 KDa polyethyleneimine polymer, 1.2PEI-tY2.1 exhibited high siRNA uptake [Fig 1]. It effectively decreased the cell surface [Fig 2] as well as mRNA levels of integrin-beta1 [Fig 3] using 60 nM of ITGB1 (integrin-beta1) siRNA. This treatment significantly reduced the binding efficiency towards fibronectin (extracellular matrix protein) for which integrin-beta1 is a primary receptor [Fig 4]. Growth of K562 cells on fibronectin for 2 days induced resistance to BCR-ABL siRNA treatment and a combinational approach with ITGB1 siRNA helped to reduce the proliferation of the cells as studied by colony formation assay [Fig 5]. Discussion and Conclusions: K562 cells develop resistant to BCR-ABL siRNA treatment when cultured on fibronectin, which acts as a model for our drug resistance study and would help us to understand the role of integrins. This acquired resistance can be successfully nullified by ITGB1/BCR-ABL combinational silencing with our polymer system.

85 Jiesi Zhou Medicine and Dentistry Oncology Ribosomal RNA cytosine methyltransferase NSUN5 in translational control in glioblastoma Jiesi Zhou, Krista Vincent, Scott Findlay, Roseline Godbout, Lynne-Marie Postovit, YangXin Fu Glioblastoma is the most common and malignant brain tumor with a median overall survival of 20.5 months. There is an urgent need to develop novel therapeutic strategies. Using a glioblastoma TCGA dataset, we have determined that high NSUN5 mRNA expression is strongly associated with poor survival in glioblastoma patients. NSUN5 is a ribosomal RNA (rRNA) cytosine methyltransferase. In yeast, Rcm1 (the homologue of NSUN5) methylates cytosine 2278 (C2278) of 25S rRNA. Deletion of Rcm1 alters the rRNA structure, ribosomal conformation, and translational fidelity that favors the translation of oxidative stress- responsive mRNAs, leading to increased stress response and lifespan. However, RNA targets of NSUN5 and its role in translational control in mammals are unknown. The objective of this project is to determine whether elevated NSUN5 changes rRNA methylation pattern and

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thereby leads to pro-tumorigenic translational reprogramming and pro-tumorigenic phenotypes in glioblastoma. Because NSUN5 is a targetable enzyme, this study may help identify novel therapeutic targets for glioblastoma.

Thus far, we have determined that NSUN5 is highly expressed in 7 out of 9 glioblastoma cell lines, as well as in two primary patient-derived glioblastoma cell lines. As expected, NSUN5 is localized in the nucleus/nucleolus in glioblastoma cells, which is consistent with its rRNA methyltransferase function. Indeed, bisulfite sequencing confirmed that NSUN5 methylates C3782 of 28S rRNA in glioblastoma cells. Functionally, overexpression of NSUN5 increases, whereas NSUN5 knockout decreases sphere formation in glioblastoma cells. Our results suggest that NSUN5 functions as an rRNA methyltransferase and may regulate cancer stem cell phenotypes in glioblastoma. Our ongoing experiments and future directions will examine the expression of NSUN5 in human glioblastoma tissues and determine whether NSUN5 promotes tumor formation and progression through reprogramming translatome in glioblastoma.

86 Nicole Del Alba Medicine & Dentistry Oncology RNF166 as a Novel Regulator of Homologous Recombination Nicole A. Del Alba, Jean-Yves Masson, & Ismail H. Ismail Repair of DNA damage is essential for cells to maintain the integrity of their genome. DNA double-stranded breaks (DSBs) are repaired by two major pathways: non-homologous end joining (NHEJ), and homologous recombination (HR). HR is active in the S and G2 phases of the cell cycle, and has a low error rate as it utilizes the available sister chromatid as a repair template for the DNA break site. Due to the low error rate, HR is an ideal DNA repair pathway for cells to prevent the accumulation of gene mutations that can lead to genomic instability over time. Recently, RNF138 -- an E3 ubiquitin ligase, was discovered in a short interfering

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RNA screen for E3 ubiquitin ligases involved in HR. Furthermore, RNF138 has been characterized to recruit to DSBs and promote cells into performing HR over NHEJ. We have found that a poorly characterized paralogue of RNF138, RNF166, also recruits to DSBs. Our research seeks to determine the potential role of RNF166 in DSB repair, and whether its function is redundant to that of RNF138 with respect to the DNA damage response. Micro- irradiation of U2OS cells transiently transfected with GFP-, or mcherry-tagged versions of RNF166 showed that the protein recruits to DSBs in similar fashion to RNF138. Furthermore, similar experiments with RNF166 truncation mutants showed that the Zinc Finger Domains of the protein mediate its recruitment to DNA damage sites. An electrophoretic mobility shift assay with radiolabelled oligonucleotides proved that RNF166 binds DNA, preferentially single-stranded DNA over double-stranded. Finally, an in vivo reporter assay was completed with the use of flow cytometry to show that RNF166 depletion causes a reduction in HR function. As a result, this research has shown that RNF166 is recruited to the sites of DNA damage, has the ability to bind DNA, and is a novel regulator of homologous recombination.

87 Zelei Yang Medicine & Dentistry Biochemistry Role of cytomegalovirus infection on breast tumor growth in mice Zelei Yang, Xiaoyun Tang, Matthew G.K. Benesch, Martina Mackova, Denise G. Hemmings and David N. Brindley Cytomegalovirus (CMV) infects 40-70% of women, but has been reported in >90% of breast cancer patients. Breast tumors cause inflammation, which stimulates autotaxin secretion from surrounding adipose tissue. Autotaxin produces lysophosphatidate, which causes further inflammation in a feed forward cycle. CMV infection also produces inflammation. Our preliminary results from treating mouse breast adipose tissue with mouse CMV (mCMV) in culture showed an increased gene expression of interleukin-6, cyclooxygenase-2, and

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autotaxin. We hypothesized that CMV infection enhances the inflammatory cycle in the breast, which promotes tumor progression.

Syngeneic mouse breast cancer models were established by injecting E0771 or 4T1 breast cancer cells into the mammary fat pad of C57Bl/6J or Balb/c mice, respectively. A transgenic MMTV-PyVT mouse model with spontaneous tumor growth was also used. Mice were treated with mCMV or medium where virus was removed at 10 weeks prior to tumor inoculation or spontaneous tumor development to establish a latent infection. Tumor growth was monitored throughout. Tumor mass, vascularity and metastasis were determined at the endpoint. Tumors, tumor-associated and contralateral adipose tissues were analyzed for inflammatory mediators. mCMV-infected C57Bl/6J mice showed tumor suppression initially, but then grow rapidly like the uninfected controls. However, the onset of this growth was unpredictable and can delay up to an additional 20 days. Autotaxin expression increased in tumor-associated adipose tissues from mCMV-infected mice, suggesting enhanced inflammation. mCMV-infected Balb/c mice showed a 3-fold increase in the number of nodules on lung, indicating greater metastasis. In MMTV-PyVT model, mCMV-infected mice showed a trend of increased total tumor burden at the endpoint and the tumors showed significantly greater vasculature and multiple tumor lobes. Mechanisms for these observations are being investigated through analysis of inflammatory mediators and immune cell subtypes.

Conclusion: mCMV infection plays a role in breast tumor progression and metastasis by modulating the host inflammatory responses.

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88 Jack Wang Medicine & Dentistry Experimental Oncology Role of DEAD Box 1 Protein in Splicing and Intron Retention Yixiong (Jack) Wang, Lei Li and Roseline Godbout DEAD box proteins are RNA helicases that are involved in all aspects of RNA metabolism. DEAD Box 1 (DDX1) is a RNA unwinding protein that was originally cloned from a subtracted retinoblastoma cDNA library. DDX1 has been reported to function as a regulatory component of the microprocessor complex. We hypothesize that DDX1 functions as an enhancer for the microprocessor complex to compete with the spliceosome complex, thereby altering the splicing of a subset of pre-mRNAs. Upon transfection of HeLa cells with siRNAs targeting either DDX1 or DROSHA (a key component of the microprocessor complex), we observed a reduction in APOBEC3H variant 4 and an increase in APOBEC3H variant 1 compared to cells transfected with scrambled siRNA. These data suggest that DDX1 may play a role in APOBEC3H alternative splicing by enhancing the activity of the microprocessor complex. Interestingly, we also have evidence showing DDX1 binds and potentially exports intron-containing APOBEC3H transcripts to the cytoplasm. Several groups have reported that intron retention is increased in cancer compared to normal tissue. We are in the process of carrying out RNA sequencing on cytoplasmic and nuclear RNAs purified from control and DDX1 knockdown cells, with the goal of shedding light on DDX1’s role in cancer-related intron retention and splicing. We are also investigating the role of DDX1 in the transport of intron-containing RNAs.

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89 Lekhini Bhatt Science Microbiology Role of ID2 in Regulating KLF4 by the Ubiquitin Proteasome Pathway Lekhini Bhatt, Michelle Wu, Armin Gamper Most cancer-associated deaths are not due to the primary tumour, but because of metastases. Breast cancers originate from epithelial cells and these must undergo a dedifferentiation process to be able to spread. The inhibitor of differentiation (ID2) and Krüppel-like factor 4 (KLF4) are both proteins involved in transcriptional regulation and have been found to be crucial in the induction of a more naïve cell state (and maybe “cancer stem cells”). The VBC ubiquitin ligase complex, containing the Von Hippel-Lindau protein (pVHL), cullin-2 and the elongins B and C, regulates KLF4 turnover. ID2 was recently found to interact with elongin C and pVHL and to inhibit the degradation of HIF1α by the VBC complex. We investigated the relationship between ID2 and KLF4, particularly whether ID2 stabilizes KLF4, a protein with a short half-life in the cell. We hypothesize that ID2 inhibits pVHL-mediated KLF4 degradation by the ubiquitin proteasome pathway. We overexpressed ID2 in MCF7, a breast cancer cell line, and used western blotting to test the effect on KLF4 levels. Overexpression of ID2 led to strong increases in KLF4 levels. qPCR of KLF4 RNA levels was used to confirm that ID2 affected KLF4 levels post-transcriptionally. We next will test mutants of ID2 that lack the basic helix-loop-helix domain (and thus transcriptional activity) or the domain found to interact with pVHL. To investigate whether increased KLF4 levels are due to ID2 interaction with pVHL, we will also knock down pVHL in MCF7. Because increased levels of KLF4 due to ID2 overexpression may increase cell metastasis in breast cancer, we will perform scratch assays with MCF7 cells overexpressing ID2. As high KLF4 or ID2 levels were found to predict poor clinical outcome in breast cancer, insights into their interplay in breast cancer can lead to improved clinical biomarkers.

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90 Allison McNamara Medicine and Dentistry Oncology Role of phospholipid targeting antibodies in anti-tumor mediated immunity in colorectal cancer Allison McNamara, Desmond Pink, John Lewis and Kristi Baker Targeted therapy is difficult for colorectal cancer (CRC) due to lack of conserved tumor antigens (TA). Immunotherapies for CRC directed at TA that can overcome intestinal tolerance need to be developed to improve survival of CRC patients. Exosomes are extracellular vesicles secreted by CRC that carry TA and display unique surface phospholipids (PPL). These PPL can be bound by immunoglobulin G (IgG) to generate immune complexes (IC). We hypothesize that engagement of the Fc region of the IC with Ig-binding receptors on dendritic cells (DC) can prime CD8+ T-cell anti-tumor immunity strong enough to overcome mucosal immunosuppression. DC were stimulated with exosomes from MC38 murine CRC cells complexed or not with anti-PPL IgG. Greater expression of inflammatory genes COX-2 and IL-12 were observed in DC stimulated with IC than exosomes alone. We co-cultured CD8+ T cells with DC exposed to IC or exosomes alone in the presence or absence of IgG receptor- blocking antibodies. IC led to greater release of IFN-λ than exosomes alone and was abrogated when Fc receptors were blocked. When mice bearing an MC38 tumor transfected with OVA were injected with IC and given anti-OVA OT-I T cells, splenic DC demonstrated greater expression of the activation marker CD86 when immunized with IC than exosomes alone. No difference in IFN-λ expression by T cells was detected. This suggests that exosome IC could stimulate an anti-tumor immune response. Further investigation of anti-PPL will allow us to understand whether this strategy could help increase anti-tumor immunity and survival in CRC patients.

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91 Nicholas Ruel Medicine & Dentistry Pharmacology Role of the nucleobase transporter SLC43A3 in cellular accumulation and cytotoxicity of 6- mercaptopurine Nicholas Ruel, Khanh Hoa Nguyen, Xenia Cravetchi, Gonzalo Vilas, James Hammond Background: 6-Mercaptopurine (6-MP) is a nucleobase analog used in the maintenance treatment of acute lymphoblastic leukemia (ALL). SLC43A3 is a recently identified nucleobase transporter involved in the cellular uptake of purine nucleobases. We hypothesize that variability in 6-MP uptake by leukemic cells via SLC43A3 leads to changes in 6-MP cytotoxicity and may contribute to inter-individual variability in 6-MP therapeutic efficacy.

Methods: HEK293 cells, which have low endogenous SLC43A3 expression and activity, were stably transfected with myc-tagged SLC43A3. 6-MP cytotoxicity was assessed over a range of concentrations using the MTT assay. SLC43A3 function was assessed via the rate of uptake of [3H]adenine and [14C]6-MP. This was correlated with cell expression of SLC43A3, as well as other potential transporters involved in 6-MP uptake/efflux, as defined by RT-PCR.

Results: SLC43A3 mediated the uptake of both [14C]6-MP and [3H]adenine with Km values of 139µM and 36µM, respectively. SLC43A3-HEK293 cells showed greater sensitivity to the cytotoxic effect of 6-MP (IC50 = 700nM) than untransfected HEK293 cells (IC50 = 7µM). In addition, expression of SLC43A3 in a range of leukemic cell lines correlated positively with [14C]6-MP accumulation into the cells (r2= 0.82, n=7).

Conclusion: SLC43A3 mediates the cellular uptake of 6-MP. Changes in SLC43A3 expression influences 6-MP rate of uptake and the sensitivity of cells to the cytotoxic effects of 6-MP. These data support the hypothesis that differences in expression of SLC43A3 in ALL patients may contribute to the variability observed in the therapeutic efficacy of 6-MP treatment.

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92 Jenilee Woodfield Medicine and Dentistry Oncology Small molecule inhibition of hypoxia inducible factor 1 alpha in triple negative breast cancer. Jenilee D. Way, Cody Bergman, Jennifer Defour, Scott Findlay, Ingrit Hamann, Lynne Postovit, and Frank Wuest Abstract: Hypoxia inducible factor 1 alpha is an over-expressed transcription factor in triple negative breast cancers. HIF-1a gets activated under hypoxic conditions and binds to its partner of HIF-1a via the PAS-B region of the HIF-1b protein. By developing small molecules that are able to target HIF-1a in the PAS-B region specifically, we can create first ever imaging agents of HIF-1a as well as possible therapeutics for the treatment of TNBCs. Herein, we design and synthesize a small library of compounds that bind to and target HIF-1a, including in vitro analysis in transfected cell lines. A compound library was synthesized from lead structures from literature1-3 yielding 13 structures with synthesis yields from 10-60%. These compounds were then tested using microscale thermophoresis with solubilized HIF-1a in order to determine their binding affinity. Binding affinity to HIF-1a was shown to be from 20- 1000 nM for many derivatives, yielding a few potentially promising candidate structures for radiolabelling. Lead structures where then radiolabelled and tested in vitro in transfected cell lines for cell uptake. Several radiolabelled HIF-1a binding compounds have been synthesized and their use has been proven in vitro, in future these will be used for imaging and treatment of TNBC in animal imaging models. Acknowledgements: The authors gratefully acknowledge the Dianne and Irving Kipnes Foundation, the National Science and Engineering Research Council of Canada (NSERC), Alberta Innovates Health Solutions (AIHS) and Alberta Cancer Foundation (ACF) for supporting this work. Reference: 1. Okamoto K, et al. (2012) Int. J. Mol. Med., 4, 541-549. 2. Nagao S, et al. (2014) Bioorg. Med. Chem., 22, 5513-5529. 3. Miranda E, et al. (2013) JACS, 135, 10418-10425.

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93 Joanne Smith Medicine & Dentistry Oncology Targeting DNA damage response in multiple myeloma Joanne Smith, Dr. Chris Venner and Dr. Ismail Ismail Multiple myeloma (MM) is an incurable cancer of antibody-producing plasma cells. The outcome for MM patients has improved substantially over the past 20 years due to several therapeutic advances. Unfortunately, even with these exciting new options available for the treatment of MM, approximately 20% of newly diagnosed MM patients are inherently resistant to proteasome inhibitors (bortezomib). Moreover, most will develop resistance over time. Therefore, discovering therapeutic agents that target specific pathways aberrantly expressed in drug sensitive and drug resistant cells will be necessary to complement existing therapeutic strategies toward a cure for this disease. Here, we show that MM cells showed signs of ongoing DNA damage and replicative stress, and activate the DNA damage response without overt apoptosis. We also found that targeting the ATM- and RAD3-related (ATR) kinase using a small molecule inhibitor selectively kills myeloma cells. The goal of this study is to develop an entirely novel therapeutic strategy for the treatment of this disorder involving agents that harness DNA replication stress and interrupt an important survival signaling pathway. If successful, these studies could lead to the development of a more effective therapy for patients with refractory multiple myeloma and potentially other blood cancers.

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94 Mahsa Mohseni Medicine and Dentistry Medicine Targeting STAT5 in Acute leukemic cells with siRNA Mahsa Mohseni, Cezary Kucharski, Remant Bahadur KC, Hasan Uludag, Joseph Brandwein* Introduction: Development of novel acute leukemia therapy is urgently needed due to poor prognosis and high relapse rates of current therapies1, 2. Transcription factors including Signal Transducer and Activator of Transcription (STAT)- protein family members are key molecular targets for acute leukemia, since they can activate expression of oncogenes leading to aberrant proliferation of cancer cells1. In hematological malignancies, downregulation of STAT5 can decrease proliferation of leukemia cells1, 2. Small interfering RNA (siRNA) mediated silencing of these targets has become a promising alternative due to its specificity and high degree of safety 3. In this study, we evaluated therapeutic role of STAT5 inhibition in acute leukemic cell lines by polymeric siRNA delivery systems.

Methods: Acute myeloid MOLM13 and Acute lymphocytic NALM-6 leukemia cells were used. Lipid- modified low molecular weight polyethyleneimine (PEI) polymers were used as siRNA carriers. Cell proliferation was assessed by MTT and DNA assays, Cellular uptake by Flow Cytometry and STAT5 knockdown at mRNA level by RT-qPCR.

Results: Specific lipid substituted 2 and 1.2 kDa PEI (2PEI and 1.2PEI) displayed excellent complexation properties with siRNAs to form nanoparticles and gave high siRNA uptake in both cells with negligible toxicity. There was a good correlation in uptake between the two cell types. Cell growth was reduced (90%) by STAT5 siRNA delivery in MOLM13 cells using 1.2PEI-lipid polymer, however, STAT5 downregulation was not enough to cause cell death in NALM6

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cells. Though some polymers showed higher uptake, STAT5 gene expression was strongly downregulated (60-70%) with leading polymers and silencing effect was higher on day 6.

Conclusions: We demonstrated effective delivery of STAT5 siRNA by polymeric nanoparticles into leukemia cells, accompanied by marked inhibition of STAT5 gene. Further experiments will be directed at evaluating STAT5 protein silencing by siRNA therapy and exploring the effect of STAT5 downregulation on leukemic patient samples.

95 Sadra Aghazadeh Science Psychology The association of FABP7 protein expression with progression and differentiation of the malignant brain tumour glioblastoma Sadra Aghazadeh, Epsita Shome, Ryan Fung, Roseline Godbout, David D. Eisenstat Introduction: Glioblastoma (GBM) is the most common malignant brain tumour and is among the most aggressive forms of cancer with less than 10% survival at 5 years. To date, only one prognostic biomarker has been identified for GBM patients, MGMT promoter methylation. This project builds on prior work from our lab studying cell death genes and patient outcomes in GBM, as well as Dr. Godbout’s lab, which focused on a protein called brain fatty acid binding protein (BFABP). My focus is to determine whether BFABP, also known as FABP7, which is expressed in radial glia, influences the progression of pediatric and adult GBM tumor cells as determined by proliferation and migration. We expect to gain insight into FABP7 as a potential prognostic biomarker for GBM.

Methods: Immunohistochemistry was used to study FABP7 expression on a cohort of 81 formalin-fixed

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paraffin-embedded (FFPE) GBM patient samples treated with surgery and chemoradiation with adjuvant cis-retinoic acid and both concurrent and adjuvant temozolomide. Expression patterns were scored and correlation with patient outcomes determined. Immunostained patient samples will also be used to compare the expression of FABP7 with GFAP and developmental markers, such as DLX2, PAX6 and PROX1. Lastly, siRNA or shRNA transfection methods will be used to assess how knockdown of FABP7 affects GBM cell lines that express high endogenous levels of FABP7 in vitro (U251, U373). Flow cytometry will be used to assess cell cycle markers, whereas cell death and invasion assays (ex. Boyden chamber) will be used to evaluate the effects on tumor cell migration in vitro.

Results: Out of 81 tissue samples, 54 showed diffuse protein expression and 27 showed focal. Six of all the samples showed high levels of staining, while 62 and 13 samples showed medium and low levels of staining, respectively. Analysis of this data is currently underway.

96 Jasdeep Mann Medicine & Dentistry Biochemistry The BCL2 family member BAD regulates breast cancer cell growth Jasdeep Mann, Tim Buckland, Rachel Montpetit, Dr. John Mackey and Dr. Ing Swie Goping We have shown that the Bcl-2 family protein, BAD is required for taxane-induced apoptosis of breast cancer cells. We further tested BAD protein levels in tumor samples of primary breast cancer patients treated with taxane chemotherapy, and identified that BAD expression is a prognostic indicator for disease-free and overall survival. We are now assessing whether BAD protein levels can specifically predict patient outcome to taxane versus non-taxane treatment, with the goal of generating a tool that will guide decisions for taxane therapy. To understand the biology of BAD in breast cancer, we investigated the molecular signaling

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pathways that were modulated by BAD. We identified that BAD regulated opposing cellular programs of apoptosis and cell growth. Specifically, phosphorylation of Ser118 stimulated cell growth, while loss of phosphorylation stimulated cell death. Hence, differing signals converge on BAD to ultimately determine cell fate, implicating that BAD functions as a novel molecular switch to regulate breast cell homeostasis. Our work suggests that BAD integrates survival and apoptotic signals to maintain homeostasis in the mammary gland, and that this pathway is deregulated in breast cancer. We propose that discrimination of these roles will elucidate a new pathway for breast cell growth and refine potential application of BAD as a diagnostic breast cancer biomarker.

97 Xiaoyun Tang Medicine & Dentistry Biochemistry The effect of lipid phosphate phosphatase 2 (LPP2) on breast cancer cell proliferation Xiaoyun Tang, Basil P. Hubbard, David N. Brindley Background: Lipid phosphate phosphatases (LPPs) consist of three isoforms: LPP1, LPP2 and LPP3, which dephosphorylate a wide variety of bioactive lipid phosphates and pyrophosphates. One of the major bioactive lipids dephosphorylated by LPPs is lysophosphatidate (LPA). LPA is produced by autotaxin (ATX) and signals through six G protein-coupled receptors (GPCRs). In cancers, LPA signaling is hijacked by cancer cells to promote tumor development and resistance to chemotherapy and radiotherapy. Overexpression of LPP1 or LPP3in breast cancer cells decreased tumor growth and metastasis in mouse models which partially correlates to the increased degradation of extracellular LPA by LPPs. In addition, LPPs showed some non-enzymatic effects which are not clearly understood so far. For example, LPP1 and LPP2 decreased ERK phosphorylation mediated by thrombin receptor activation in HEK293 cells. LPP1 inhibited protease-activated receptors (PARS) stimulated calcium transit in MDA-MB-231 cells. It is quite possible that these non- redundant functions of LPPs play important roles in cancer cell signaling transduction and

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tumor development. Significantly, the effect of LPP2 on cell growth is different from LPP1 and LPP3 which is probably caused by the non-enzymatic function of LPP2. Fibroblasts overexpressing LPP2 showed a premature entry into S-phase of cell cycle. LPP2 knock-down delayed the entry of cells into S-phase and inhibited cell growth. Genome screen data has identified PLPP2/LPP2 expression is elevated in different carcinomas and sarcomas and this contributes to the transformed phenotypes.

Hypothesis: Therefore, we hypothesized that LPP2 has distinct effects on breast cancer development.

Methods: mRNA expression of LPP1-3 in normal human breast tissues and human breast tumors were determined by quantitative PCR. LPP2 knockout MDA-MB-231 and MCF 7 cell lines were generated by CRISPR. Cell growth was compared between wild type and LPP2 knockout cells. Cell cycle was analyzed by flow cytometry. Levels of cell cycle proteins were determined by western blot.

Results: LPP1 mRNA level was ~12 times lower in human breast tumor tissue relative to normal human breast tissues (p<0.01). LPP3 mRNA level showed a trend of decrease relative to normal breast tissues, but was not statistically significant. LPP2 mRNA was ~8 times higher in breast tumors than normal breast tissue (p<0.01). LPP2 knockout MDA-MB-231 cells showed a ~40% decrease in cell growth relative to wild type cells. In cell cycle, LPP2 knockout caused a ~33% increase of cells in G0/1phase and a ~39% decrease of cells in S phase. LPP2 knockout caused decreased protein levels of phospho-Rb, cyclin A2 and cyclin B1in MDA-MB- 231 cells. Similarly, LPP2 knockout MCF 7 cells showed a ~60% decrease in cell growth in 3D culture relative to wild type cells.

Conclusions: Human breast tumors express higher level of LPP2 mRNA relative to normal breast tissue. LPP2 is involved in cell proliferation of MDA-MB-231 and MCF 7 cells, which appears to be a potential target for breast cancer treatment.

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98 Zackariah Breckenridge Medicine Oncology The Evolving Impact on Survival Outcomes in Multiple Myeloma with Greater Access to Novel Treatment Regimens: Experience in a Real-World Cohort Zackariah Breckenridge, BSc, Victor Jimenez-Zepeda, MD, Nizar J. Bahlis, MD, Paola Neri, MD, Irwindeep Sandhu, MD, FRCPC, Michael P. Chu, MD, Ismail Ismail, Jason Tay, MD, FRCPC, Peter Duggan, MD, FRCPC, Andrew Belch, Joanne D. Hewitt, NP, PhD, Linda M. Pilarski, PhD, Tatiana Nikitina and Christopher P. Venner Introduction: Novel agents (NA) are now established standards of care in the treatment of multiple myeloma (MM) in all lines of therapy. The use of proteosome inhibitors (PIs) and immunomodulatory drugs (IMiDs) has led to marked improvements in survival outcomes. Furthermore, these agents have evolved as the standard backbones for newer regimens incorporating novel approaches like monoclonal antibody-based (MAb) therapies and other targeted anti-plasma cell agents. Data is evolving with respect to patient outcome after exposure to standard PI and IMiD-based regimens in the real-world setting. We previously presented our provincial experience examining outcomes after subsequent lines of therapy in a setting where PI and IMiD-based approaches are routine. Herein, we update and expand the original series with a further 2 years of follow-up and with the cohort having greater access to newer more targeted agents. Patients and methods: This series includes patients seen through the provincial myeloma program in Alberta, Canada. Only patients who initiated first line NA-based therapy between 2005-2015 were included to allow at least 2 years of follow-up. We examined double exposed (DE) patients, classified as those who had been treated with, but were not necessarily refractory to, both an IMiD and PI. We also examined patients who were double exposed by the end of their second line of treatment (DE2) as well as those who were double refractory (DR). Overall survival (OS) was measured from the time each line of therapy was started until death or last follow-up. Progression free survival (PFS) was from the time of

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therapy initiation to relapse, death or last follow-up. The category of MAb-based approaches was specifically examined given the unique mechanism of action of these agents. This category includes patients treated with daratumumab, elotuzumab, durvalumab and nivolumab-based combinations. Results: Outcomes were examined after the 1st, 2nd, 3rd and 4th line therapy. Six hundred and nineteen patients have received upfront therapy of which, 311 and 308 were ASCT eligible and ineligible. The median follow-up for the whole cohort is 41 months. There was a statistically significant difference in favour of ASCT eligibility for frontline therapy with a median OS of 103 months vs 51 months (P < 0.001)) and a median PFS of 42 months vs 19 months (p < 0.001). Three hundred fifty five patients had relapsed. Survival outcomes between patients eligible and ineligible for frontline transplant were similar and thus were analyzed as a single cohort. Outcomes are presented in Table 1. From 2nd, 3rd and 4th line therapy the median OS were 36 months, 20 months, and 12 months and the median PFS were 12 months, 8 months and 6 months respectively. In the DE, DE2 and DR cohorts the median OS were 34 months, 36 months and 17 months and the median PFS were 11 months, 11 months and 4 months respectively. Forty-two patients received MAb based therapy for relapsed disease. Thirty-five patients received daratumumab, 4 received elotuzumab, 2 durvalumab, and 1 nivolumab. Forty-eight percent of patients in this cohort achieved a ≥PR (29% ≥VGPR). In this cohort the median OS and PFS from the time of first MAb exposure were 13 months and 3 months respectively. Focusing on the most widely used MAb daratumumab, 14% achieved ≥PR (9% ≥VGPR). In this cohort the median OS and PFS from the time of daratumumab exposure were 12 months and 4 months respectively. Of note, 71% of patients received their MAb in 4th line or later making this a heavily pretreated population. Summary: In this updated series we continue to see remarkable survival outcomes for both transplant eligible and ineligible patients. In the former group, the median OS is approaching 10 years. In addition, we have seen gains in OS after each line of therapy likely owing to increased therapeutic options available for relapsed patients. This is notable given that patients are becoming double exposed to both traditional NA classes much earlier in their disease course. However, once double refractory the expected median PFS is only 4 months,

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again emphasizing that new approaches will be required to improve outcomes in this challenging population. The data presented here reveals a glimpse into the impact of MAb in a real-world setting; an important class of drugs owing to their entirely different mechanism of action. This data will help further benchmark expected outcomes as the therapeutic landscape in MM evolves.

99 Courtney Mowat Medicine and Dentistry Experimental Oncology The influence of genetic instability on the mucosal immune system in colorectal cancer Courtney Mowat, Jianyin Liu, Sharmin Sumi, Kristi Baker The prevalence of unhealthy lifestyles is increasing the incidence of colorectal cancer (CRC) associated deaths and evidence indicates that the microbiome plays an important role in this. The tolerogenic mucosal immune system maintains homeostasis with the gut microbiome, but tolerance can ultimately hinder tumour detection. Interestingly, 15% of CRCs are microsatellite instable (MSI) and have a favourable prognosis compared to other CRCs. This is partially due to MSI CRC having defective DNA mismatch repair (dMMR), mainly in MLH1, resulting in a hypermutable phenotype and therefore an accumulation of aberrant proteins that are recognized as tumour associated antigens (TAAs) and trigger an anti-tumour immune response. Based on preliminary data this cannot be the whole story because tumours with mutations in POLE do not have high infiltrating activated cytotoxic T cells like MLH1 deficient tumours despite also being hypermutable. We are using CRISPR to disrupt different DNA repair pathways in murine intestinal epithelial cells (IECs) to investigate whether MSI CRCs have microsatellite mutations in pattern recognition receptors (PRR) that regulate the tumour's interaction with the microbiome. We expect this will induce immunological changes that enhance tumour detection and may explain MSI CRCs improved prognosis. We are stimulating PRR on mutated IECs with bacterial antigens and whole bacteria to evaluate if mutations in mismatch repair or other DNA repair pathways alter the anti-tumor immune

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response triggered by the microbiome. If we can resolve the immune consequences of dMMR on CRC, then perhaps we can enhance the susceptibility of all CRC to available immunotherapies.

100 Min Hsuan Wu Medicine and Dentistry Oncology The regulation of transcription factor Sox2 via genotoxic stress and hypoxia. Min Hsuan Wu, Amirali Bukhari, Lekhini Bhatt, Kareena Nanda, Armin Gamper Within a tumour, only a subset of cells, often called tumour initiating cells or Cancer Stem Cells (CSCs), are able to form neoplasms following failed eradication or localization to distant organs. CSCs are characterized by high cellular plasticity, which is a trait of normal tissue stem cells and induced-pluripotent stem cells. Yamanaka and colleagues identified four transcription factors that are crucial for dedifferentiation and the establishment of cellular plasticity. One of these factors, Sex-determining region Y box protein 2 (Sox2), is the focus of my research. Upregulation of Sox2 has been observed to promote tumour progression in certain tissues and its activity has been associated with CSC maintenance. Recent studies suggest that some DNA damaging therapies may trigger dedifferentiation of cancer cells to a stem cell-like state and possibly an increase in CSCs. In addition, tumours often have regions of hypoxia due to disorganized angiogenesis. Hypoxic conditions promote both stemness and resistance to radiotherapy. Interestingly, hypoxia was reported to upregulate Sox2. As both DNA damage and hypoxia have been reported to regulate cancer cell plasticity, we hypothesize that DNA damage signalling can regulate Sox2 levels and activity and that hypoxia can modulate this regulation. The resulting alteration in Sox2 levels have important consequences for the establishment (or maintenance) of cells with high plasticity, resulting in adverse outcome for genotoxic therapies. My preliminary data show depletion of Sox2 protein following genotoxic stress and induction of its activity with IR and hypoxia. We intend to study the specific upstream signalling pathways regulating Sox2

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protein levels following genotoxic stress and hypoxia as well as the control mechanisms of their transcriptional activity. I thus hope to not only gain insight into mechanisms underlying Sox2 regulation, but also establish whether DNA damage-induced changes in Sox2 activity affect cancer cell plasticity.

101 Powel Crosley Medicine & Dentistry Oncology TRAIL-expressing oncolytic vaccinia virus in combination therapy against cancer Powel Crosley; Kate Agopsowicz; Kyle Potts; Brittany Umer; Marjut Pihlajoki; Markku Heikinheimo; Anniina Farkkila; David Evans; Mary Hitt PAC1 is a small-molecule drug shown to sequester inhibitory zinc ions from Caspase-3 (CASP3) allowing CASP3 to auto-mature and execute apoptosis. PAC1 has demonstrated safety and efficacy in vivo and is currently in Phase I trials for advanced malignancies. TRAIL is a pro-apoptosis ligand that can bind membrane-bound death receptors. This binding triggers the extrinsic apoptotic pathway resulting in activation of CASP3 to execute its proteolytic role in programmed cell death. TRAIL has been found to induce apoptosis selectively in cancer cells and has been well tolerated in human patients. Our preliminary in vitro results indicate that combining low-dose TRAIL with PAC1 in treatment of granulosa cell tumour (GCT) cells (both model cell-line and patient-derived cells) induces more loss of viability after 48 hours than either PAC1 or TRAIL alone. Interestingly, adding TRAIL 24 hours after PAC1 treatment activates similar levels of caspase activity and cytotoxicity within a few hours. Our group has also developed and evaluated novel oncolytic vaccinia virus (VACV) mutants that are selective for replication in many types of cancer, resulting in cancer cell lysis. Some of the VACV mutants have been assessed in vitro in the GCT model cell line KGN and results suggest that VACV enhances toxicity of PAC1/TRAIL combined treatment. Our current project is investigating generation of a novel VACV mutant to include expression of

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secreted soluble TRAIL for use in vitro and in vivo against GCT that has been pre-treated with PAC1.

102 Marawan Ahmed Pharmacy Pharmaceutical Sciences The Too Many Faces Of PD-L1: A Comprehensive Conformational Analysis Study Marawan Ahmed, and Khaled Barakat In the current study, we focused on the immune-checkpoints PD-1 pathway and in particular on the ligand, PD-L1 [1]. We studied the conformational dynamics of PD-L1 through principal component analysis (PCA) of existing crystal structures combined with classical and accelerated molecular dynamics simulations. We identified the maximum structural displacements that take place in all PD-L1 crystal structures and in the MD trajectories. We found that these displacements are attributed to specific flexible regions in the protein. We also investigated the conformational preference for small molecule binding and highlighted a Methionine residue at the binding site, which plays a key role in drug binding [2]. The binding mechanism of PD-L1 to other binding partners is also discussed in details from a computational perspective. We hope that the data presented here supports the ongoing efforts to discover effective therapies targeting the PD-1 immune-checkpoint pathway. References

1-Chen, Y.; Liu, P.; Gao, F.; Cheng, H.; Qi, J.; Gao, G. F., A dimeric structure of PD-L1: functional units or evolutionary relics? Protein Cell 2010, 1, 153-60. 2-Zak, K. M.; Grudnik, P.; Guzik, K.; Zieba, B. J.; Musielak, B.; Domling, A.; Dubin, G.; Holak, T. A., Structural basis for small molecule targeting of the programmed death ligand 1 (PD-L1). Oncotarget 2016, 7, 30323-35.

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103 Sindhuja Pattabhi Raman Medicine and Dentistry Experimental Oncology Theranostic evaluations of bioreductively-activated Tirapazamine (TPZ) prodrugs for the management of hypoxic solid tumors Raman SP, Elsaidi H, Postovit LM, Weinfeld M, Kumar P Solid tumours have large areas with low levels of oxygen (known as hypoxic regions), which are associated with poor prognosis and treatment response. Tirapazamine (TPZ), a hypoxia targeting anticancer drug, started as a promising candidate to deal with this issue. However, it was withdrawn from the clinic due to severe neurotoxic side effects and impaired target delivery. Hypoxic cells overexpress GLUT transporters - a key feature during hypoxic tumour progression. My project aims at conjugating TPZ with glucose to exploit the upregulated GLUTs for its delivery, to facilitate both imaging and therapeutic management of hypoxic tumors. Glucose-conjugated TPZ (G-TPZ) will be selectively recruited to these receptors, facilitating its uptake in poorly oxygenated cells only. This targeted approach will minimize the toxicity to the oxygenated normal cells. The molecule also has the features to bestow both diagnosis and in situ radiotherapy, which adds further value to TPZ. Our hypothesis will be validated using selected head and neck and glioblastoma cancer cell lines, followed by its evaluation in pre-clinical animal models. I have already confirmed the overexpression of GLUT-1 in oxygen-depleted cells, which forms the basis for future studies with G-TPZ. Effects of TPZ therapy will be evaluated using key molecular markers of hypoxia such as the hypoxia selective transcription factor HIF1-alpha. The cytotoxicity of the conjugate will be determined by clonogenic and live/dead assays to evaluate its chemo/radiotherapeutic potential. Together this approach aims to overcome the limitations of TPZ therapy in the context of cancer treatment.

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104 Faisal Rashed Medicine and Dentistry Oncology Therapeutic Targeting of Tumour Hypoxia Using 2-Nitroimidazole Imaging Agents Faisal B. Rashed; Dawn Macdonald; Alexandru C. Stoica; Carolynne Ricardo; Ramanaguru S. Piragasam; Richard P. Fahlman; Lynne-Marie Postovit; Piyush Kumar; Michael Weinfeld Abnormal vasculature in solid tumours often results in poor oxygenation; a phenomenon that is known as tumour hypoxia. Hypoxic cells are resistant to chemo- and radiotherapy, and thus may contribute to treatment failure and relapse. Targeting hypoxia, therefore, is vital for therapeutic management of solid tumours. 2-Nitroimidazole compounds (2-NI) undergo hypoxia selective entrapment in oxygen deficient cells through bioreduction and subsequent formation of protein-drug adducts. As a result, radiolabeled 2-NIs are widely used as radiodiagnostics to monitor tumor hypoxia. Clinically established examples include our compounds of interest; 123I-iodoazomycin arabinoside (123I-IAZA) and 18F- fluoroazomycin arabinoside (18F-FAZA). This study explores the unique premise of using these 2-NI hypoxia radiotracers in a therapeutic setting. In our experiments, both IAZA and FAZA act as hypoxic cytotoxins, and can further sensitize oxygen deprived cells to ionizing radiation. We hypothesized that hypoxic sensitization by 2-NI compounds is achieved partly through interaction with critical cellular proteins. To identify the potential interaction partners of our drugs, we have adopted a click chemistry based approach using a modified analogue of our parent compounds. The hypoxia selectivity of this method has been confirmed in cell and xenograft mouse models. A biotin labelled alkyne allowed us to pull down the drug-protein adducts, which were then analyzed through mass spectrometry. Our pilot study identified heat shock protein 90 (Hsp90) as a potential candidate. Hsp90 is required for stabilization of the hypoxia inducible factor 1 alpha subunit (HIF-1α), which provides growth selective advantages to hypoxic cells. Our preliminary data suggests that 2-NI compounds can decrease HIF-1α protein levels in irradiated hypoxic cells. The mechanism of this regulation is currently being investigated in details.

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105 Nasim Ghasemi Pharmacy and Pharmaceutical sciences Pharmacy Towards Development of Biodegradable Chemically Crosslinked Hydrogels for Stimulus Responsive Drug Delivery Ghasemi N, Vakili MR, Lavasanifar A Purpose: The long-term objective of this study is to develop biodegradable stimuli-responsive gels with optimal properties for depot and smart drug delivery. In this study, we developed the method of polymerization for synthesis of triblock copolymers based on poly (ethylene glycol) (PEG) and functionalized poly(ε-caprolactone) (PCL). We then evaluated the effect of dihydroxy PEG 200 Da as cross linker on the gelation of triblock copolymers. Methods: Triblock copolymers of poly (α-carboxyl-co-benzyl carboxylate-ε-caprolactone)-b-PEG-b-poly (α-carboxyl-co-benzyl carboxylate-ε- caprolactone) (PCBCL-PEG-PCBCL) were prepared through ring opening polymerization of α-benzyl-ε-caprolactone (BCL) and PEG (1450 kDa) through bulk polymerization method with and without PEG 200 Da as cross linker; this step followed by hydrogenation of block copolymer with palladium on activated charcoal under hydrogen gas stream. Prepared block copolymers were characterized to find out molecular weight, polydispersity index (PDI), and the cross linking degree by gel permeation chromatography (GPC).1H NMR was also used to characterize the structure and molecular weight of the polymer. The sol-gel transition of polymer solutions in water was measured using inverse flow method, differential scanning calorimetry (DSC) and rheometric analysis. Results: GPC results proved that PEG 200 Da acted as cross linker during the ring opening polymerization and polymers prepared by pure monomer and PEG 200 Da as cross linker yielded PCBCL-PEG-PCBCL with sol-gel transition temperatures around 35°C at concentration 10-15%. Conclusion: The present study illustrates successful synthesis of triblock copolymers based on PEG and functionalized PCL with sol-gel transition close to physiological body temperature. The cross linker improved the mechanical property of the gelation state of the gel. It also decreased gel-forming concentration of the triblock copolymer.

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106 JoAnn Thai Faculty of Medicine & Dentistry Oncology, Surgery The Role of Hypoxia Induced Transcription Factors in Renal Cell Carcinoma Angiogenesis JoAnn Thai, Hua Chen, Abul Azad, Ron Moore Clear cell renal cell carcinoma (ccRCC) is often characterized by a mutation in the von Hippel- Lindau (VHL) gene, which is associated with the activation of hypoxia-induced transcription factors (HIFs) such as HIF2α. These transcription factors activate pro-tumorigenic target genes that eventually lead to angiogenesis and survival pathways for the cell. Angiogenesis is the process by which blood vessels emerge from pre-existing blood vessels in normal physiological conditions. In phases of rapid tumour growth, cancer cells release growth factors to encourage endothelial cells to grow and differentiate to aid in the growth of the tumour, thus leading to blood vessel formation. Cells use the process of autophagy to degrade proteins and damaged organelles through lysosomal degradation. Autophagy has been identified to be one of the mechanisms through which HIF2α is degraded.

In this study, we investigated the role of HIF2α in ccRCC angiogenesis and the behaviour of these transcription factors within the cell. We used an angiogenesis fibrin bead assay to observe that growth factors released by cancer cells promote angiogenesis in human umbilical vein endothelial (HUVEC) cells. Using western blots and cell cytotoxicity assays in 786-O ccRCC cells both with and without a VHL mutation, we compared the presence of HIF2α in ccRCC cells. We found that 786-O cells treated with autophagy inhibitor bafilomycin A1 had increased levels of HIF2α and survived better compared to parental 786-O re- expressed with VHL. These results indicate that HIF proteins like HIF2α are a possible therapeutic target for kidney cancer. Although the specific mechanism of action remains to be established, these HIFs offer a possible route to treat unwanted angiogenesis within a tumour.

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107 Aishwarya Iyer Medicine & Dentistry Medicine TRANSLATING GENOMIC INTRATUMOR HETEROGENEITY INTO PROGNOSTIC BIOMARKERS IN T-CELL LYMPHOMA Aishwarya Iyer, Weiwei Wong, Dr. Gane Wong, Dr. Robert Gniadecki, Mycosis fungoides (MF) is a common extranodal T-cell lymphoma primarily arising in the skin. In early disease stages, MF presents as skin patches and plaques that in some cases may progress to tumor and disperse to lymph nodes and other internal organs. Early diagnosis is difficult as the histology overlaps with features of inflammatory skin diseases. Even when the diagnosis is established there are no prognostic markers that predict whether the disease will be aggressive or indolent. Lastly, there are no curative treatments and MF will invariably relapse even after aggressive chemotherapy. The disease is a diagnostic, prognostic and therapeutic challenge. The main objective of this study is to address the question of tumor heterogeneity in MF. To date, MF is considered to be monoclonal, derived from a transformed, mature memory T- cell. However, clinical observations and preliminary data suggest that MF comprises multiple subclones, which may be of importance for understanding of disease evolution and resistance to therapy. We plan to address this objective using Whole Exome Sequencing (WES) of MF tissue prepared by laser microdissection (LMD). Following are our study objectives: Description of mutational landscape of early disease (plaques) and late stages (tumors). The emphasis of this objective is to find genetic differences between early and late stage disease and to identify key signaling pathways likely to be affected by these mutations. Deciphering the genetic evolution of MF. Patients with MF usually develop multiple lesions and it is not clear whether advanced lesions (tumors) develop by evolution from plaques or rather emerge from lymphoma precursor cells. We will compare mutational landscapes between plaques and a tumors

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sampled from the same patient to describe the developmental tree of the tumor. Achieving these objectives will assist in the precision diagnosis and prognostication of the disease.

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MEETING PARTICIPANTS LIST

Acevedo-Morantes, Claudia Y Danielson, Brita Aceytuno, Danny De Zanche, Nicola Aghazadeh, Sadra Del Alba , Nicole Agopsowicz, Kate DIAZ DUSSAN, DIANA Ahmadi, Fatemeh Dietz, Bryson Ahmed, Marawan Dinakaran, Deepak Ajidagba, Morenike Dobberthien, Brennen Alaee, Mahsa Dutta, Indrani Alharbi, Abdulsalam Dutta, Pranamika Ali, Mohammad Elfiky, Abdo Ali, Noureen Elian, Fahed Ali, Declan Elmenoufy, Ahmed Andersson, Jan Fairgrieve-Park, Logan Anoveros, Ana Fallone, Clara Ansari, Aysha Fallone, B. Gino Arbabimoghadam, Sahar Ferguson, Meghan Augustine, Aruna Field, Catherine Azad, Abul Kalam Findlay, Scott Babu, Dinesh Fitieh, Amira Baker, Kristi Fliegel, Larry Baksh, Shairaz Fu, YangXin Barakat, Khaled Gamper, Armn Barta, Radim Ganesan, Aravindhan Basonbul, Asmaa Gentile, Francesco Bell, Clayton Ghasemi, Nasim Bhatt, Bhumi Ghila, Andrei Bhatt, Lekhini Githaka, John Bhavsar, Amit Glover, Mark Bhullar, Amritpal Godbout, Roseline Bilyk, Lena Goping, Ing Swie Brander , Kristina Graham, Kathryn Breckenridge, Zackariah Grose, Laura Brindley, David Gupta, Nidhi Brodeur-Robb, Kathy Hamedi, Bahareh Bukhari, Amirali Hammond, James Campbell, Shelagh Han, Gawon Chan, Gordon HAO, YUBIN Chen, Huachen Hejazi, Maryam chen, hua Hemmings, Denise Choi, Won Shik (Daniel) Hewitt, Joanne Chu, Michael Hitt, Mary Chwyl, Brian Huang, Fleur Coatham, Mackenzie ILARRAZA, RAMSES Constantinescu, Gabriela Islam, Md Touhidul (Apu) Cristi Munoz, Francisca Ismail, Ismail Crosley, Powel Ismail, Alaa

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Iyer, Aishwarya Mriouah, JIhane Jain, Saket Mula-Hussain, Layth Jans, Hans-S. Murray, David Jay, David Murray, Allan Jenish, David Nami, Babak Julien, Olivier Narine, Kelly Kalantar, Al Nevin, Mikaela Kalyaanamoorthy, Subha Newell, Marnie Kang, Dong-Woo Nguyen, Hoa KC, Remant Noyce, Ryan Kedarisetti, Pradyumna Olayinka, Lily Kim, Hyeong Jin Omar, Sara Kin, Tatsuya Ostergaard, Hanne Kirkham, Amy Pacheco-Pereira, Camila Kramer, David Paiva, Igor Krys, Daniel Palmaria, Cynthia Kumar, Piyush Pandya, Vrajesh Kumaran, Mahalakshmi Panigrahi, Rashmi Lavasanifar, Afsaneh Pasdar, Manijeh Lee, Laura Pattabhi Raman, Sindhuja Lehner, Richard Peters, Anthea Lewis, Cody Pink, Desmond Lewis, John Postovit, Lynne LI, LEI Potts, Kyle Li, Christie Prado, Carla Li, Xiuju Purcell, Sarah Litchfield, Marcus Purchase, Aaron liu, jiahui Rajput, Sunil Liu, Jianyin Ramamonjisoa, Nirilanto Liu, Rong-Zong Rashed, Faisal Liu, Michelle Refaei, Mohammad Locke, Andrew Robson, Paula MacGillivray, Melinda Ruel, Nicholas Maki, Rachel Sadat, Sams Mann, Jasdeep Salla, Mohamed McDonald, Rory Seelam, Sudhakara Reddy McNamara, Allison Shen, Michael Meenakshi Sundaram, Daniel Nisakar Siegers, Gabrielle Meng, Guanmin Simmen, Thomas Menghisteab, Isaac Singhal, Sandeep Menon, Geetha Smith , Joanne Mercier, Rebecca Souza, Nilian Mohseni, Mahsa Srayko, Martin moradipoor, sara Storozynsky, Quinn Moshari, Mahshad Swan, Amanda Mowat, Courtney taheri rouhi, seyedeh zeinab

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Tandio, David Tang, Xiaoyun Taylor, Michael Thai, JoAnn Thapa, Bindu Tuszynski, Jack Ubeda, Anyeld Uludag, Hasan Umer, Brittany Underhill, Alan Valencia, Juliana VanderSluis, Laura Vega, Hector Vena, Jennifer Vo, Kevin Walter , Mike Wang, Jack Wee, Ping Weilbeer, Claudia Weinfeld, Michael Wiebe, Ericka Willms, Reegan Woodfield, Jenilee Wu, Min Hsuan Xiao, Jingjie Xu, Xia Yahya, Atiyah Yang, Zelei Yasmin, Lubna Yip, Wan Kong Yuen, Richard zare, Alaa Zemp, Roger zhang, guihua Zhou, Jiesi Ziegler, Kirby

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ORGANIZING COMMITTEE

Aruna Augustine

Gareth Armanious

Dr. Shairaz Baksh

Dr. Mary Hitt

ACKNOWLEDGEMENTS

We would like to thank our sponsors for contributing to this event, the Office of the Dean, Faculty of Medicine & Dentistry, for their support, our abstract selection committees, poster judges, and all of the CRINA trainee volunteers, particularly Sara Omar and Saket Jain, involved in making CRINA Research Day 2017 such a success.

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