Friday, October 13th 2017 - McGill University › pharma › files › pharma › ... · Senior Scientist at Princess Margaret Cancer Centre since 1996. Her lab studies tumour microenvironment

23rd ANNUAL PHARMACOLOGY RESEARCH DAY

Friday October 13th 2017

SSMU Ballroom McGill University 3480 rue McTavish Montreal Qc

Department Pharmacology and Therapeutics

Special Thanks To All Of Our Sponsors

Innovative Medicine

GE Healthcare

Pfizer Canada Inc

Faculty of Medicine McGill University

Bruker

Milteny

Promega

Carl Zeiss Canada Ltd

Boehringer Ingelheim

Montreal Biotechnologies Inc

Megs Specialty Gases Inc

Special Mention

Wisent Bioproducts

Laser Quest

N sur MacKay

Table of Contents

PRD Student Committee Welcome1 Keynote Speaker2 Departmental and PRD Prizes 3 PRD Judges 3 Program4-5 Abstracts ndash Oral Presentations 6-13 Poster Session I-Odd Numbers 14-33 Poster Session II-Even Numbers34-53

2017 PHARMACOLOGY RESEARCH DAY COMMITTEE

Dr Lisa Muumlnter Chair of Research Day Dr Bastien Castagner Co-Chair of Research Day

Bobbi Bidochka Tina Tremblay Andrew Bayne Morgan Foret

Chrismita Hegde Anne-Sophie Pepin

Han Yan Noosha Yousefpour

Issan Zhang

A very special Thank You to

Annie Constantin Anna Cuccovia Chantal Grignon Chelsea Dufort

Dear Colleagues

Welcome to the 23rd Annual Pharmacology Research Day

This year we are excited to welcome our keynote speaker Dr Rama Khokha presenting on Mammary stem cells and breast cancer We look forward to learn about Dr Khokhas expertise in stem cell and cancer research She is renowned for her contributions to these fields from development of models to cancer genomics

We are thrilled by the calibre of the student talks and posters that will be showcased throughout the day It is a great opportunity to network and learn about the cutting edge research happening in the Department of Pharmacology and Therapeutics as well as the other departments represented here

We would like to extend special thanks to the event sponsors for their generous contributions and the organizing committee Dr Lisa Muumlnter (Chair of PRD) Dr Bastien Castagner (Co-Chair of PRD) Dr Gerhard Multhaup Tina Tremblay and Bobbi Bidochka for their dedication and hard work put towards the planning and implementation of this fantastic event

Sincerely Students of the Pharmacology Research Day Committee

Left to right Morgan Foret Noosha Yousefpour Issan Zhang Chris Hegde Anne-Sophie Peacutepin Andrew Bayne Missing Aileen Yan

Keynote Speaker

Dr Rama Khokha is a Professor in the Department of Medical Biophysics and the Interim Research Director of Princess Margaret Cancer Centre She received her PhD from University of Western Ontario and her postdoc training at Cancer Research Labs (London Ontario) and at EMBL labs (Germany) She has been a Senior Scientist at Princess Margaret Cancer Centre since 1996 Her lab studies tumour microenvironment and adult stem cell niches with a major interest in breast and bone cancers Her program is also active in developing genetically engineered mouse models for human cancers and novel tools for cancer gene discovery Her work was recognized by the prestigious Robert L Noble Prize from the Canadian Cancer Society Research Institute in 2014

PHARMACOLOGY RESEARCH DAY PRIZES

Melville Prizes This prize established in 1994 is awarded annually to

the Pharmacology students in the MSc and PhD category whose research poster presentation at the

Pharmacology Research Day is judged to be the best A Melville Prize is also awarded to a Pharmacology Postdoctoral Fellow whose research posteroral

presentation is judged to be the best

Best Oral Presentation Prize This prize is awarded annually to a Pharmacology graduate student for the oral presentation that best

exemplifies multidisciplinary approach in Pharmacology

PRD Life Sciences Prize (Anatomy and Cell Biology Biochemistry IPN)

This prize established this year by Pharmacology amp Therapeutics is awarded to a graduate student

presenting the best postersorals at Pharmacology Research Day

Pharmacology Research Day Judges

Dr Mark Aurousseau Dr Bertrand Jean-Dr Oceacuteane Albert Claude Dr Dan Bernard Dr Aimee Katen Dr Derek Bowie Dr Dusica Maysinger

Dr Bastien Castagner Dr Anne McKinney Dr Paul Clarke Dr Alexandre Moquin

Dr Dominic Devost Dr Gerhard Multhaup Dr Sonia Do Carmo Dr Lisa Munter Dr Greg Fitzharris Dr Alfredo Ribeiro-da-

Dr John Gillard Silva Dr Barbara Hales Dr Bernard Robaire Dr Mark Hancock Dr Uri Saragovi Dr Terry Heacutebert Dr Laura Stone

Dr Shireen Hossain Dr Chirine Toufaily Dr Jean-Franccedilois

Trempe

23rd Annual Pharmacology Research Day

Friday October 13th 2017 SSMU Ballroom University Centre 3480 McTavish

Montreal QC H3A 1X9

PROGRAM

08h15 Registration Coffee and Morning Poster Set-up

08h45 Pharmacology Research Day Introduction - Ballroom

- Dr Gerhard Multhaup Professor and Chair

- Dr Lisa Munter PRD Chair

09h00-10h00 ORAL SESSION I

Andrew Bayne MSc Junior lab of Dr Jean-Franccedilois Trempe Dr Patricia Brown Postdoc lab of Dr Derek Bowie Luc Truong MSc Junior lab of Dr Jean-Franccedilois Trempe Chrismita Hegde PhD Junior lab of Dr Bastien Castagner

10h00-11h30 BREAK amp POSTER SESSION 1 Odd Numbered Posters present

11h30-12h30 KEYNOTE ADDRESS Dr Rama Khokha Mammary stem cells and breast cancer Princess Margaret Cancer Centre University of Western Ontario Department of Medical Biophysics University of Toronto

12h30-13h30 LUNCH - Please visit the sponsor booths

13h30-15h00 ORAL SESSION II 13h30 Yining Li PhD Senior lab of Dr Dan

Bernard 13h45 Adamo Mancino MSc Junior-IPN lab of

Dr Derek Bowie 14h00

Jennifer Chen PhD Junior lab of Dr Jason Tanny Dr Filip Liebsch Postdoc lab of Dr Gerhard Multhaup

14h30-16h00 POSTER SESSION 2 Even Numbered Posters present

16h00-19h00 Reception with Departmental Piano Jam Session - bring your instrument Award Ceremony and Closing Remarks

__________________________________________________

Presenter Andrew Bayne 09h00 MSc JrO1

Title Characterizing Human Mitochondrial Processing Peptidase and its Role in Parkinsonrsquos Disease

Authors Bayne Andrew Trempe Jean-Franccedilois

Abstract Mitochondrial processing peptidase (MPP) is a metallopeptidase that cleaves N-terminal targeting signals from the majority of nuclear-encoded mitochondrial proteins Mutations in both MPP and its substrates have been implicated in neurodegenerative diseases including Parkinsons disease (PD) and non-progressive cerebellar ataxia One of these implicated substrates is PINK1 - a kinase whose mutations are known to cause early-onset autosomal PD In healthy mitochondria PINK1 is constitutively imported cleaved first by MPP and then retrotranslocated to the cytosol for proteasomal degradation In this regard MPP acts as a key junction for PINK1 import proteolysis and overall mitochondrial quality control However both the MPP cleavage site on PINK1 and its binding conformation remain unknown To gain insight into the proteolytic mechanisms concerning PINK1 and other disease-implicated substrates we have begun to characterize the human MPP heterodimer We demonstrate that the human MPP dimer can be reconstituted from a co-expression system in E coli We have also developed a mass spectrometry-based method to monitor MPP activity using synthetic N-terminal peptides from a variety of mitochondrial proteins Using this system we have identified the specific MPP cleavage site on PINK1 Research into the mechanism and kinetics of PINK1 processing are currently ongoing in our laboratory

Acknowledgments CIHR GRASP GEPROM

__________________________________________________

Presenter Patricia Brown 09h15PostdocO3

Title Stargazin and Cornichon-3 relieve polyamine block of AMPA receptors by enhancing blocker permeation

Authors Patricia MGE Brown Hugo McGuire Derek Bowie

Abstract Ligand- and voltage-gated ion channels assemble as signaling complexes comprised of both pore-forming and auxiliary subunits In the mammalian brain AMPA-type ionotropic glutamate receptors (AMPARs) co-assemble with several families of auxiliary subunits that regulate channel gating as well as ion channel block and permeation Previous work has shown that auxiliary proteins stargazin (or gamma2) and cornichon-3 (CNIHshy3) attenuate cytoplasmic polyamine channel block of AMPARs though the underlying mechanism has yet to be established Here we show that gamma2 and CNIH-3 relieve channel block by enhancing the rate of blocker permeation Surprisingly the relative permeability of the polyamine spermine (Spm) through the pore of the AMPAR-gamma2 or -CNIH-3 complexes was considerably increased compared to AMPARs expressed alone A modified model of permeant channel block fully accounted for both the voltage- and time-dependent nature of Spm block Estimates of block rate constants revealed that auxiliary subunits do not attenuate block by shifting the location of the block site within the membrane electric field nor do they affect the blockerrsquos ability to reach it Instead gamma2 and CNIH-3 relieve channel block by facilitating the blockerrsquos exit rates from the open channel From a physiological perspective the relief of channel block exerted by gamma2 and CNIH-3 ensures that there is unrestricted signaling by AMPARs at glutamatergic synapses Moreover the pronounced ability of AMPARs to transport polyamines may play an unexpected role in regulating cellular polyamine levels

Acknowledgments CIHR NSERC

__________________________________________________

Presenter Luc Truong 09h30 MSc JrO3

Title Characterizing the importance of S205 autophosphorylation in tcPINK1 activation

Authors Truong L Rasool S Trempe J-F

Abstract Mutations in the kinase PINK1 cause autosomal recessive early-onset Parkinsonrsquos disease (PD) PINK1 selectively accumulates on damaged mitochondria and recruits Parkin by phosphorylating its Ubl domain as well as nearby ubiquitin ultimately leading to targeted mitophagy PINK1rsquos kinase activity therefore acts as a critical signal transducer for mitochondrial damage Similarly to other kinases PINK1 is known to phosphorylate itself (autophosphorylate) namely at residues S402 and S228 Problematically of these two phosphoacceptor sites the role of S228 has not been characterized as emphatically as that of S402 and how its autophosphorylation modulates PINK1rsquos activity was probed in this project We used the Tribolium Castaneum homolog of PINK1 tcPINK1 and the equivalent residue of S228 is S205 We show that the S205A and S205N tcPINK1 mutants have significantly less Ubl phosphorylation activity and fail to phosphorylate ubiquitin in vitro We also use mass spectrometry (MS) to reveal the different autophosphorylation status between these mutants and the WT Indeed although the mutants remain catalytically active they have two fewer phosphate groups than the WT on average Finally MS was again used in a 15N-labelled tcPINK1 experiment to demonstrate that autophosphorylation occurs intermolecularly or in trans Furthermore it was shown that S205 is the only transshyautophosphorylation phosphoacceptor site by trypsin-digest LCshyMSMS Overall this project refines our understanding of kinase activity and by extension mitochondrial quality control by characterizing the importance of S205 autophosphorylation in tcPINK1 The impulse created by these results could reveal more focal points to study in PD and help in the development of pharmacological therapies targeting the PINK1-Parkin pathway

Acknowledgments Andrew Bayne Marta Vranas Nathalie Croteau Nimra Khan

__________________________________________________

Presenter Chrismita Hegde 09h45 PhD JrO4

Title Synthesis of Pathological Calcification Inhibitors

Authors Hegde Chrismita Castagner Bastien Dr

Abstract In chronic kidney disease (CKD) the kidneys are unable to regulate serum mineral concentrations The ensuing rise in serum calcium leads to the progressive calcification of soft tissue(s) and vasculature CKD-dependent vascular calcification is a major factor in the development of cardiovascular disease (CVD) the top cause of death worldwide The calcium in mineralized tissues can bind inositol phosphates a class of phosphorylated poylols Grases et al demonstrated that inositol hexakisphosphate (IP6) inhibits cardiovascular calcification in rats As well preliminary experiments with a PEGylated inositol phosphate (PEG550-IP5) indicated retarded conversion of the serum calcifying agent from the primary to the secondary active form We can surmise that the negative charge density enables these compounds to bind inorganic calcium Here we describe the synthesis of phosphorylated inositols and their analogs which may delay this formation of secondary calcifying agents and have therapeutic potential as calcification inhibitors Eventually we plan to elaborate on the structure-activity relationship of these compounds with respect to their calcium binding ability

Acknowledgments Castagner Lab Zamboni Chemicals CIHR NSERC

__________________________________________________

Presenter Yining Li 13h30 PhD SrO5

Title Enhanced fertility in female mice with a gonadotropeshyspecific deletion of the inhibin co-receptor TGFBR3 (betaglycan)

Authors Li Y Fortin J Boehm U Lin H Bernard D

Abstract Follicle-stimulating hormone (FSH) is an essential regulator of ovarian follicle development and fertility in females FSH synthesis is stimulated by intra-pituitary activins and is suppressed by gonadal inhibins Activins stimulate transcription of the FSHβ subunit gene (Fshb) in gonadotrope cells Inhibins suppress FSH in vitro by acting as competitive activin receptor antagonists particularly in the presence of the co-receptor TGFBR3 (betaglycan) The role of TGFBR3 in inhibin action in vivo has not been determined because Tgfbr3 null mice die during embryonic development We therefore generated mice harboring lsquofloxedrsquo alleles for Tgfbr3 allowing us to ablate the gene specifically in gonadotropes in vivo (hereafter cKO) Pituitaries from cKO mice were insensitive to inhibin A in primary culture Similarly inhibin A suppressed FSH in control but not cKO males in vivo cKO females were supra-fertile ovulating more eggs per cycle and producing approximately two more pups per litter than controls cKO ovaries contained a greater number of large antral follicles and corpora lutea than controls Though these phenotypes were consistent with increased FSH neither synthesis nor secretion of the hormone appeared to differ between control and cKO females or males FSH is a glycoprotein and patterns of glycosylation affect the ligandrsquos actions We are therefore investigating whether cKO mice produce a more active form of FSH than controls We are also examining whether the ovaries of cKO mice are more sensitive to FSH than those of controls Collectively the results indicate that TGFBR3rsquos role in pituitary gonadotropes may be more complex than originally conceived

Acknowledgments This research was funded by CIHR MOP-133394 to DJB

__________________________________________________

Presenter Adamo Mancino 13h45 MSc Jr IPNO6

Title The Voltage-Gated Sodium Channel Nav15 makes an Unexpected Appearance in the Brain

Authors Mancino Adamo Aurousseau Mark Miraucourt Loiumls Alexander Ryan Yan Edward Bowie Derek

Abstract The voltage-gated sodium channel Nav15 is responsible for the rapid upstroke in the cardiac action potential (AP) Consistent with this mutations in the Nav15 gene have been linked to various forms of cardiomyopathy Despite the emphasis on Nav15 in the heart emerging evidence also suggests a role for Nav15 in neurological disease particularly epilepsy However the detailed location and role of Nav15 in the brain remains unexplored Using RT-PCR we detected Nav15 mRNA from mouse brain extracts as two splice-variants ndash mH1 as found in the adult heart and Nav15e Heterologous expression and whole-cell recordings of Nav15e revealed a ~10 mV depolarizing shift in its activation curve compared to mH1 The stronger depolarizations required to activate Nav15e might make it less likely to contribute directly to the AP Furthermore immunolabeling revealed that Nav15 is expressed in neurons critical to brain function including mitral cells of the olfactory bulb layer 5 pyramidal cells of cortex parvalbumin-positive basket cells of the hippocampus and cerebellar Purkinje neurons Co-labelling experiments also yielded overlapping expression of Nav15 and the auxiliary protein β3 suggesting that they might assemble together as a signalling complex When co-expressed β3 relieved steady-state inactivation and accelerated recovery from inactivation of Nav15 increasing the likelihood that Nav15 can signal at resting membrane potentials Moreover in whole-cell recordings of cerebellar Purkinje cells blockade of Nav15 by ranolazine significantly reduced the firing rate without altering the overall AP shape Taken together this work establishes for the first time the widespread expression pattern of Nav15 in the brain and highlights the non-canonical role that Nav15 plays in fine-tuning neuronal excitability as opposed to direct AP generation

Acknowledgments Many thanks to colleagues who contributed to this vast project namely Mark for his expertise in molecular biology Lois for his knowledge in immunohistochemistry Ryan for his work in acute slices Edward for his help on the recombinant front and Derek for coordinating the project Of course this work could not have been done without our funding sources so thanks to NSERC and CIHR

Presenter Jennifer Chen 14h00 PhD JrO7

Title Mapping Novel Interactions between Rtf1 and Histone Modifying Complexes

Authors Chen Jennifer Mbogning Jean Page Viviane Tanny Jason

Abstract Histone modifications are crucial in regulating many cellular processes including RNA polymerase II gene transcription and DNA repair Therefore a dysregulation of these modifications can lead to cancer pathogenesis promoting interest for research on histone modifications These modifications regulate the balance between active and repressed chromatin states with certain histone modifications predominantly associated with active genes such as histone H2B monoubiquitination (H2Bub1) This modification occurs through conserved E2 and E3 enzymes but the activation of H2Bub1 requires the PAF complex and Rtf1 conserved regulators of transcription elongation whose mechanisms of action are poorly understood PAF complex Rtf1 and H2Bub1 regulate stem cell differentiation organismal development and cancer cell growth making them potentially attractive therapeutic targets Our lab is interested in elucidating the molecular mechanisms linking Rtf1 and the PAF complex in regulating histone modifications and more generally transcription We use a simple eukaryotic model system fission yeast (Schizosaccharoymyces pombe) Preliminary data shows novel interactions in vitro between the +3 domain of Rtf1 and various histone modifying complexes and transcription factors which could explain the roles of Rtf1 and these enzymes in histone modifications Specific subunits of the complexes as well as mutations that perturb these interactions have been identified Surface Plasmon Resonance (SPR) is being used to further confirm these interactions Furthermore assays to monitor Rtf1 function in elongation and termination (RNA polymerase ChIP and qRT-PCR at specific genes) and in promoting H2Bub1 (westerns ChIP) are being used to determine transcriptions defects By understanding the molecular interplay between these different factors we gain further mechanistic insight into how the histone modification and transcriptional machineries are interlinked

Acknowledgments This work was funded by the Canadian Institute of Health Research (CIHR)

Presenter Filip Liebsch 14h15PostdocO8

Title Diagnostic potential of the CSF-Aβ34Aβ42 ratio during pre-symptomatic and prodromal stages of Alzheimerrsquos disease

Authors Liebsch F Kulic L Teunissen C Engelschalt V Hancock M van der Flier W Rosa-Neto P Scheltens P Poirier J Breitner J Hock C Multhaup G Abstract It has been hypothesized that an imbalance between the production and clearance of amyloid-β (Aβ) is a major event to initiate the pathogenesis of Alzheimerrsquos disease (AD) Thus altered Aβ cleavage profiles could provide first signs of biochemical changes during pre-symptomatic stages of AD With a newly generated neo-specific monoclonal antibody we developed a sensitive immunoassay to specifically measure the amyloid cleavage product Aβ34 in cerebrospinal fluid (CSF) We assayed Aβ34 in 192 CSF samples from individuals at different stages of disease expression and assessed its relationship to classical AD biomarkers Individuals with mild cognitive impairment (MCI) who later progressed to AD had elevated Aβ34 when compared with MCI that remained stable or individuals with subjective memory complaints Aβ34 levels correlated positively with other known AD biomarkers in CSF namely total-tau and phosphorylated (P181)-tau in all clinical groups Compared to CSF-Aβ42 alone the ratio of Aβ34Aβ42 increased diagnostic accuracy to distinguish between prodromal AD and stable MCI In cognitively normal individuals at risk for AD (ie from the PREVENT-AD cohort) an increased Aβ34Aβ42 ratio was associated with pre-clinical stages of AD a higher dementia risk score and elevated t-tau and P181-tau Overall this study indicates that combining established biomarkers with C-terminally truncated Aβ cleavage products such as Aβ34 may provide improved accuracy to diagnose preshyclinical and prodromal Alzheimerrsquos disease Acknowledgments Thanks to BrainMcGill the Velux Stiftung and the Canadian Institute of Health Research (MOP-133411) for support awarded to GM CH LK or FL Thanks to the Studienstiftung des Deutschen Volkes and the Groupe de Recherche Axeacute sur la Structure des Proteacuteines (GRASP McGill University) for PhD fellowships awarded to FL GM holds both a Canada Research Chair in Molecular Pharmacology and a Canada Foundation for Innovation (CFI) grant Thanks to Robert Umek and Martin Boissonneault (Meso Scale Discovery) for their help and support with the electrochemiluminescence assay development The McGill SPR-MS Facility thanks the CFI for infrastructure support Data used in the preparation of this article were obtained from the Preshysymptomatic Evaluation of Novel or Experimental Treatments for Alzheimerrsquos Disease (PREVENT-AD) program (httpwwwprevent-alzheimerca) data release 20 (November 30 2015 Update June 07 2016)

POSTER SESSION I 10h00 ndash 11h30

P1 Shafqat Rasool PhD Senior Biochemistry

P3 Erik Larson PhD Senior IPN

P5 Sherilyn Recinto MSc Junior IPN

P7 Ryan Alexander PhD Senior IPN

P9 Ha Eun Kim (Christine) Kim MSc Junior Pharmacology

P11 Xiao Ming (Sindy) Zheng MSc Junior Pharmacology

P13 Fiona Hui MSc Senior Pharmacology

P15 Nicholas Kim MSc Senior Pharmacology

P17 Suleyman Can Akerman PhD Junior Pharmacology

P19 Yubo (Frank) Cao PhD Junior Pharmacology

P21 Han (Aileen) Yan PhD Senior Pharmacology

P23 Sarah MacKinnon PhD Junior Pharmacology

P25 Anne Marie Downey PhD Senior Pharmacology

P27 Sean Jmaeff PhD Senior Pharmacology

P29 Baraa Noueihed PhD Senior Pharmacology

P31 Gauthier Schang PhD Senior Pharmacology

P33 Noosha Yousefpour PhD Senior Pharmacology

P35 Mariana Asslan MSc Junior Pharmacology

P37 Mary Loka MSc Junior IPN

__________________________________________________

Presenter Shafqat Rasool 10h00PhD Sr BiochemP1

Title PINK1 autophosphorylation is required for ubiquitin recognition

Authors Rasool Shafqat Truong Luc Soya Naoto Luckas Gergeley Trempe Jean-Francois

Abstract Mutations in PINK1 cause autosomal recessive Parkinsonrsquos disease (PD) a neurodegenerative movement disorder PINK1 is a kinase that acts as a sensor of mitochondrial damage and initiates Parkin-mediated clearance of the damaged organelle PINK1 phosphorylates Ser65 in both ubiquitin and the ubiquitin-like (Ubl) domain of Parkin which switch on its E3 ligase activity Autophosphorylation of PINK1 is required for Parkin activation but how this modulates the ubiquitin kinase activity is unclear Here we show that autophosphorylation of Tribolium castaneum PINK1 is required for substrate recognition Using enzyme kinetics and NMR spectroscopy we reveal that PINK1 binds the Parkin Ubl with a 10-fold higher affinity than ubiquitin via a conserved interface that is also implicated in RING1 and SH3 binding The interaction requires phosphorylation at Ser205 an invariant PINK1 residue (Ser228 in human) Using mass spectrometry we demonstrate that PINK1 rapidly autophosphorylates in trans at Ser205 Hydrogen-deuterium exchange experiments provide insights into the structure of the PINK1 catalytic domain and reveal how Ser205 phosphorylation affects the dynamics of the canonical C-helix and activation loop in the active site Our findings suggest that multiple PINK1 molecules autophosphorylate first prior to binding and phosphorylating ubiquitin and Parkin

Acknowledgments Nathalie Croteau

__________________________________________________

Presenter Erik Larson 10h00PhD Sr IPNP3

Title Mouse model of Fragile X syndrome has deficient inhibitory GABAergic plasticity

Authors Larson Erik A Accardi Michael V Wang Ying Karimi Benyamin Varaschin Rafael K Siddiqui Tabrez J Bowie Derek

Abstract Fragile X syndrome (FXS) is a neurodevelopmental disorder linked to deficits in several neurotransmitter systems Dysfunction in both glutamatergic and GABAergic signaling are thought to play key roles in the manifestation of the disease yet the exact nature of these defects is still emerging Here we identify a novel plasticity mechanism of inhibitory synapses in cerebellar molecular layer interneurons (MLIs) that is absent from FXS mice We describe a novel activity-dependent plasticity triggered by NMDA receptors (NMDARs) which recruit 3shycontaining GABAA receptors (GABARs) into inhibitory synapses Recruitment of GABARs required an elevation of cytoplasmic reactive oxygen species (ROS) and signaling through a protein kinase CGABARAP-dependent pathway Furthermore we show that excitatory input by NMDARs is diminished in FXS mice and consequently inhibitory synapse strengthening by 3-receptors is functionally lost Since MLI connectivity is important in shaping Purkinje cell activity and consequently cerebellar output our findings identify a disruption in synapse coupling in these neurons which adds to the growing body of evidence connecting cerebellar dysfunction to the pathophysiology of FXS

Acknowledgments This work was supported by Canadian Institutes of Health Research Operating Grants CIHR MOP-342247 to DB and by CIHR MOP-142209 to TJS

__________________________________________________

Presenter Sherilyn Recinto 10h00MSc Jr IPNP5

Title Mechanism of RHBDL4-mediated APP processing linked to Alzheimerrsquos disease

Authors Recinto Sherilyn Paschkowsky Sandra Munter Lisa

Abstract In Alzheimerrsquos disease (AD) excessive amyloid-β (Aβ) deposition is one of the earliest biomarker to appear prior to the emergence of overt clinical symptoms Aβ peptides are generated from amyloidogenic processing of amyloid precursor protein (APP) by β- and -secretases Autosomal dominant inheritance mutations in APP and presenilin genes are the major cause of early-onset AD presumably due to enhanced A production Recently APP was also elucidated to be a substrate of the mammalian rhomboid protease RHBDL4 leading to significantly reduced Aβ levels in cell culture experiments APP metabolites generated from RHBDL4 cleavages differ from those derived from canonical processing suggesting an alternative processing pathway for APP Hence it is important to characterize the functionality of RHBDL4 and its relevance in AD by analyzing cleavage patterns and efficiency of different synthetic APP TMS mutants as well as familial AD (FAD) mutants We hypothesize that RHBDL4 specificity is conferred in the transmembrane sequence (TMS) of its substrate and inheritable APP mutations within or near the TMS are differentially processed by RHBDL4 Using HEK293T cells we show that co-expression of RHBDL4 and synthetic APP mutants resulted in abrogation of RHBDL4shyderived large C-terminal fragments (CTFs) but N-terminal fragments (NTFs) remained intact This suggest a plausible two-tier RHBDL4 substrate recognition mechanism for APP ie RHBDL4 cleaves APP at multiple sites and TMS specificity are only pivotal at certain cleavage sites We also observed enhanced levels of large CTFs in APP FAD mutants upon RHBDL4 cleavage indicating that they may be more stable as compared to wild type Thus familial mutations in APP not only affect amyloidogenic processing but also processing by RHBDL4 In summary our research further investigated the alternative RHBDL4-mediated APP processing which may provide novel insight in the underlying AD pathology

Acknowledgments Munter Lab Dr Lisa Munter Sandra Paschkowsky Felix Oestereich Jing Lias Sasen Efrem Jacqueline Hsiao Funding NSERC RGPIN-2015-04645-15 Scottish Rite Charitable Foundation and Pfizer-FRQS

__________________________________________________

Presenter Ryan Alexander 10h00PhD Sr IPNP7

Title Long-term modulation of excitability by NMDA receptor signaling in cerebellar stellate cells

Authors Alexander Ryan Mitry John Sareen Vasu Khadra Anmar Bowie Derek

Abstract The action potential (AP) is a fundamental signaling unit used by neurons to communicate within networks The AP is generated through the interplay of several voltage-gated ion channel (VGIC) families including Na+ and K+ channels which determine threshold and frequency of firing Although synaptic activity-driven changes in neurotransmission have been described throughout the brain the long-term influence of synapses on VGIC behaviour is less well characterized We have observed a novel regulation of excitability in cerebellar stellate cells that involves NMDA receptor-mediated modulation of both Na+ and K+ channel activity Local application of NMDA induced a persistent increase in spontaneous action current frequency during on-cell electrophysiological recordings In whole-cell current clamp recordings stellate cells exhibited a time-dependent increase in evoked AP frequency and hyperpolarization of spike threshold both of which were eliminated by pharmacological block of CaMKII To better understand the precise contribution of each VGIC family a neuronal firing model was constructed and compared to experimental data This revealed that the hyperpolarizing shift in stellate cell AP threshold can be primarily explained by changes in Na+ channel gating while modulation of A-type K+ current and delayed rectifier K+ channels will affect spike frequency Our work shows a novel modulation of Na+ channels by excitatory synapses and provides insight into the role of NMDA receptor-dependent signaling in regulating inhibitory neuronal circuits of the cerebellum

Acknowledgments NSERC CIHR Savoy Foundation McGill Faculty of Medicine

__________________________________________________

Presenter Ha Eun Kim 10h00MSc JrP9

Title Hypothyroidism induces relaxin-3 mRNA expression in pituitaries of mice

Authors Kim H Turgeon MO and Bernard DJ

Abstract Thyroid hormones (THs) play fundamental roles in development and adulthood Deficiency in TH production (hypothyroidism) is common affecting approximately 2 of the Canadian population Most often the disorder derives from problems at the level of the thyroid gland (primary hypothyroidism) More rarely the problem lies within the brain andor pituitary gland (central hypothyroidism) The brain produces a tripeptide thyrotropin-releasing hormone (TRH) which stimulates secretion of thyroid-stimulating hormone (TSH) TSH in turn stimulates TH production by the thyroid THs then feed back to the brain and pituitary to suppress TRH and TSH Mutations in the immunoglobulin superfamily member 1 (IGSF1) gene are the most common cause of congenital central hypothyroidism Mice with loss of function mutations in the Igsf1 gene exhibit down-regulation of the TRH receptor (Trhr1) in the pituitary and are therefore less sensitive to TRH However the cellular function of the IGSF1 protein remains elusive To address this gap in knowledge we compared patterns of gene expression in pituitaries of wild-type and Igsf1 knockout mice using RNA-sequencing Animals of both groups were rendered profoundly hypothyroid with a diet low in iodine and supplemented with propylthiouracil (LoIPTU) prior to the analysis Gene expression was highly similar between the two genotypes However we noted that relaxin 3 (Rln3) mRNA levels were significantly lower in knockout relative to wild-type animals As Rln3 expression in pituitary had never before been reported we hypothesized that enhanced TRH signaling in hypothyroid mice may drive expression of the gene Indeed the LoI-PTU diet induced 25-100 fold increases in pituitary Rln3 mRNA levels We are now generating loss of function mutations in the Trhr1 and Rln3 genes using CRISPR-Cas9 to determine whether 1) TRH regulates Rln3 expression and 2) Rln3 regulates TRH or TSH secretion through a novel feedback loop

Acknowledgments Funding Sources Supported by CIHR MOPshy133557

__________________________________________________

Presenter Xiao Ming (Sindy) Zheng 10h00MSc JrP11

Title Gβγ signalling conservation from an evolutionary perspective

Authors Xiao Ming Zheng Shahriar Khan Rory Sleno Phan Trieu Terry Heacutebert

Abstract Heterotrimeric G proteins comprised of α β and γ subunits are responsible for relaying signals from G protein-coupled receptors (GPCRs) to downstream effectors such as enzymes and ion channels In recent years the obligate Gβγ dimers have been found not only on the plasma membrane where the majority of GPCRs are located but also at multiple subcellular sites such as mitochondria Golgi apparatus and the nucleus serving both canonical and novel non-canonical roles Moreover a diverse set of 5 Gβ and 12 Gγ subunits are found in mammals in comparison with invertebrates like Caenorhabditis elegans and lower eukaryotes like Schizosaccharomyces pombe The focus of this project is to understand and characterize the role of Gβ and Gγ subunit diversity from an evolutionary perspective by investigating how invertebrate Gβγ subunits behave in a mammalian cellular environment Tagged constructs were made with the Gβ homologues found in C elegans namely Flag-GPB1 and Flag-GPB2 Analysis of protein expression via western blot showed that both C elegans Gβ homologues were expressed in mammalian cells Previously our lab has shown that Gβ4γ1 served as a modulator of endogenous M3-mAChRs signaling in HEK 293 cells knockdown of Gβ4 resulted in a loss of calcium release upon activation of the receptor Furthermore Gβ1 was found to function as transcriptional regulator acting on the Gβ4 promotor Based on this study in HEK 293 cells we examined whether C elegans Gβ homologues could rescue the loss of signalling under Gβ knockdown conditions We observed that Flag-GPB1 rescued the loss of calcium signalling that resulted from Gβ4 knockdown while Flag-GPB2 had no effect Clearly there are features of Gβγ function that have been altered during evolution

Acknowledgments CIHR

Presenter Fiona Hui 10h00MSc SrP13

Title Heterologous expression of neuropeptide GPCRs NPR-5 from the parasite Brugia malayi

Authors Hui Y Geary T

Abstract Parasitic nematodes cause various debilitating infections in humans creating health and economic burdens in developing countries Current drug treatments have limited efficacy and can cause severe adverse reactions Therefore new potential targets are needed to develop novel anthelmintics The FMRFamide-like peptide (FLP) receptors are GPCRs activated by the FLP family of neuropeptides While FLPs are widely conserved in nematodes and have important roles in feeding reproduction and neuromuscular functions this family is essentially absent from vertebrates making FLP receptors appealing anthelmintic drug targets Existing studies using model organism C elegans have shown that FLP receptor NPR-5 is activated by peptides encoded on the flp-18 precursor gene and that NPR-5 modulates metabolism and feeding behaviour However the relevance of this receptor in a parasitic nematode context has yet to be investigated In this study we are investigating the activity profile of NPR-5 from the parasitic nematode B malayi We hypothesize that the putative B malayi FLP receptor Bm-NPR-5 is activated by FLP-18 peptides and have comparable activity with its orthologue Ce-NPR-5 Bm-NPRshy5 was heterologously expressed using a yeast system GPCR activation induces HIS3 reporter gene expression allowing yeast survival in histidine drop-out medium Yeast proliferation quantified using Alamar Blue reflects the extent of receptor activation This functional assay indicated that while Ce-NPR-5 was activated by the native ligand FLP18-6 Bm-NPR-5 showed no activity Bm-NPR-5 was then tested in Chinese hamster ovary (CHO) cells and showed activation by several FLP-18 derived peptides This supported our hypothesis that Bm-NPR-5 is activated by FLP-18 peptides We plan to further investigate the amino acid sequences required to activate Bm-NPR-5 and to compare the activation profile of Bm-NPR-5 with Ce-NPR-5 assessing NPR-5 as a potential drug target in parasitic nematodes Acknowledgments This research was funded by NSERC and the Canada Research Chair program awarded to Dr Timothy Geary We thank Dr Sven Zels from the Functional Genomics and Proteomics Research Group of KU Leuven for functionally expressing Bm-NPR-5 in CHO cells and testing Bm-NPR-5 against their library of peptides

__________________________________________________

Presenter Nicholas Kim 10h00MSc SrP15

Title Role of heme metabolism in pathological neovascularization implications for retinopathy of prematurity

Authors Kim N Cagnone G Kim J Heckel E Pundir S Gaub P Wunnemann F Szarek W Schipper H Joyal JS

Abstract Aim Retinopathy of prematurity (ROP) is characterized by pathological neovascularization (NV) protruding into the vitreous body threatening retinal detachment Important risk factors of ROP include prematurity and exposure to excess oxygen High oxygen content delays vascular growth and causes vessel loss (vaso-obliteration [VO]) which creates avascular hypoxic retinal pockets that later induce pathological NV Here we are studying whether regulation of heme metabolism may be involved in pathological NV through the action of heme oxygenase-1 (HMOX1) an enzyme induced by hypoxia to degrade heme into iron biliverdin and carbon monoxide With anti-apoptotic anti-inflammatory and anti-oxidant capabilities HMOX1 is notoriously cytoprotective in several disease models but has never been thoroughly investigated in ROP We therefore hypothesize that HMOX1 triggered by hypoxia may contribute to pathological angiogenesis in ROP Methods amp Results Wild-type (WT) C57BL6 pups are exposed to high oxygen concentrations (FiO2 75) from the post-natal day (P) 7 to 12 and subsequently returned to room air until retinal collection oxygen induced retinopathy (OIR) Our pilot data showed increased retinal Hmox1 mRNA expression in OIR from P12 to 19 (n=7 plt001 at P17) coinciding with elevated Hmox1 protein levels measured by Western blot (n=3 plt005) Single-cell RNAseq also identified predominant Hmox1 expression in microglia and Gfap positive Muumleller cells which led us to investigate the impact of Hmox1 in these cell types Finally quantification of retinal areas for VO and NV at P17 allowed us to observe decreased NV by 60 (n=5 Plt001) without significant influence on VO (n=5 not significant) following pharmacological inhibition of Hmox1 Conclusion Our preliminary findings indicate Hmox1 expression in glial cells under hypoxic stress may contribute to pathological NV Glial Hmox1 may therefore have a critical role in the development of diseased vessels

Acknowledgments NSERC VHRN CHU Sainte-Justine McGill University

__________________________________________________

Presenter Suleyman Can Akerman 10h00PhD JrP17

Title Deciphering the Novel Role of Amyloid-β42 in the Nucleus

Authors Akerman S Can Hossain Shireen Multhaup Gerhard

Abstract Alzheimer disease (AD) is a debilitating proteinopathy lacking effective diagnostics prevention and treatment Clinicians must currently rely on the presentation of late-stage events such as cognitive decline whereas the underlying pathophysiological events likely begin decades before the presentation of clinical symptoms The amyloid-β (Aβ) peptides are critical to AD pathogenesis and intraneuronal accumulation of Aβ is seemingly the earliest detectable event (Selkoe and Hardy 2016) There is growing evidence that neurodegeneration may have links to defects in the nucleocytoplasmic transport machinery (Zhang et al 2015) including AD In fact the Ras-related nuclear protein Ran a key molecule that dictates the directionality of this transport was found to be decreased in hippocampal neurons of AD patients (Mastroeni et al 2013) Our laboratory has previously described a novel role for Aβ42 in nuclear signaling and gene regulation (Barucker et al 2015) Aβ42 specifically showed gene regulatory properties in human SH-SY5Y neuroblastoma cells We have investigated whether Aβ was responsible for the reduction of Ran protein levels using the same neuroblastoma cell culture model and have found that Aβ42 specifically caused a reduction in Ran levels Furthermore nuclear envelope irregularities have been observed in AD (Sheffield et al 2006) and protein aggregation diseases such as Huntington disease show disruption of the nuclear envelope (De Grima et al 2017) We have hypothesized that Aβ42 could be causing these observations through genetic regulation of nucleocytoskeleton proteins However our experiments so far revealed that Aβ42 is not causing any alterations of the nuclear pore complex nor the nuclear envelope Hence it is possible that nuclear envelope defects observed in AD may be due to other factors such as tau either in conjunction with Aβ42 or by itself

Acknowledgments I would like to thank Dr Stochaj for her valuable input on this project

__________________________________________________

Presenter Yubo (Frank) Cao 10h00PhD JrP19

Title Studying the Signaling and Trafficking Signatures of Nonsynonymous Mutations of the Human Angiotensin II Type 1 Receptor

Authors Cao Yubo (Frank) Kumar Sahil Namkung Yoon Laporte Stephane

Abstract Hypertension and its associated risks can be attributed to the endogenous hormone angiotensin II (AngII) AngII regulates blood volume and vascular resistance through the angiotensin II type 1 receptor (AT1R) Mutations in the AGTR1 gene that alter the native amino acid sequence are referred to as nonsynonymous mutations (NSMs) While certain NSMs of the AT1R have been reported to reduce AngII binding it is unknown whether mutations can also alter downstream cellular responses Therefore in this study we investigated the effects of three NSMs of the AT1R (A163T T282M and C289W) on signaling and trafficking We hypothesized that the amino acid alterations caused by these NSMs will induce unique receptor conformations that would allow for differential coupling to cognate G proteins (Gαq Gα12 Gαi2 Gαi3) andor β-arrestins To characterize their signaling and trafficking signatures bioluminescence resonance energy transfer (BRET)-based biosensors were used to quantify the receptorsrsquo efficiency for engaging signaling and endocytosis effectors Results show that upon AngII stimulation the A163TshyAT1R displayed similar potency and efficacy in activating G protein-mediated signaling pathways relative to the wild type receptor but increased β-arrestin recruitment to the receptor Conversely the T282M-AT1R and C289W-AT1R had reduced relative activity in all pathways assessed Notably there was a marked biased decrease in Gα12 coupling to the T282M-AT1R In addition although the T282M-AT1R recruited β-arrestin and internalized upon AngII stimulation the receptor trafficked into endosomes without being in complex with β-arrestin Taken together these results suggest that positions A163 and T282 are important for β-arrestin-mediated trafficking of the AT1R while a mutation at the latter causes a bias in G protein-mediated signaling demonstrating that changes in certain residues can not only alter receptor trafficking but also induce a biased signaling profile

__________________________________________________

Presenter Han (Aileen) Yan 10h00PhD SrP21

Title The effects of new generation organophosphate ester flame retardants on endochondral ossification

Authors Yan H Hales BF

Abstract Flame retardants (FR) are added to a plethora of consumer products to slow fire propagation Growing evidence that ldquolegacyrdquo polybrominated diphenyl ether (PBDE) flame retardants are persistent bioaccumulative and toxic has led to a phase out of their usage Organophosphate esters (OPEs) are popular PBDE replacements Triphenyl phosphate (TPhP) and tert-butylphenyl diphenyl phosphate (BPDP) are commonly used OPEs but little is known about their safety We showed previously that the skeleton may be a target of PBDE toxicity In this study we hypothesized that the OPEs are safer alternatives and thus have no effect on bone formation We tested this hypothesis using the 6-day limb bud culture system and a strain of transgenic mice expressing fluorescently tagged collagen markers for the different stages of endochondral ossification COL2A1-eCFP (early chondrogenesis) COL10A1-mCherry (late chondrogenesis) and COL1A1-eYFP (osteogenesis) Limb buds from gestation day 13 embryos were cultured in the presence of vehicle (DMSO) 1μM or 10μM of either FR Limb morphology scoring indicated that exposure to even 1μM of either TPhP or BPDP significantly reduced the extent of limb differentiation compared to controls The fluorescent markers revealed that 10μM of either congener dramatically inhibited the progression of bone formation Furthermore treatment with even 1μM of either OPE significantly reduced the expression of the three master regulators of bone formation Sox9 Runx2 and Sp7 Together these studies provide evidence that OPEs may adversely impact endochondral ossification warranting further study of their potential toxicity

Acknowledgments Supported by funding from CIHR RQR CRRD and McGill University (Alexander McFee Fellowship GEF)

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Presenter Sarah MacKinnon 10h00PhD JrP23

Title Genetic Screen Reveals a Novel Role For Transcription Elongation Factor Spt5 in Heterochromatin Regulation

Authors MacKinnon S Tanny J C

Abstract Histone modifications play a significant role in determining whether genes are actively expressed or silenced One silencing modification methylation of histone 3 on lysine 9 (H3K9me) is associated with repression of underlying genes and is a marker for transcriptionally repressed heterochromatin H3K9me can lead to aberrant gene silencing that has been associated with multiple forms of cancer Small molecule drugs have been developed to prevent the formation of H3K9me in diseases where it is overabundant however their use in humans is likely limited due to their inhibition of physiological H3K9me An understanding of the factors that prevent the formation and propagation of this heterochromatic mark is therefore warranted and could lead to identification of new therapeutic targets A genetic screen completed at the start of this project indicated that the transcription elongation factor Spt5 is required to prevent an accumulation of H3K9me outside of its normal compartment through a process known as heterochromatin spreading While Spt5 plays a described role in promoting transcription elongation and histone modifications associated with gene activation its role in negatively regulating a H3K9me is novel This project aims to (1) determine if Spt5 regulates the extent of H3K9me formation as indicated by the genetic screen and (2) to determine the proteins that partner with Spt5 in this function Chromatin Immunoprecipitation (ChIP) with an anti-H3K9me antibody indicated that heterochromatin spreads outside of its compartment in Spt5 mutant cells We will confirm that the increase in H3K9me leads to inappropriate gene silencing in these mutants Further ChIP experiments will localize candidate proteins involved in H3K9me regulation and will determine if Spt5 functions to recruit these proteins These experiments will dissect the mechanism through which Spt5 prevents the formation of a gene-silencing histone modification

Acknowledgments

__________________________________________________

Presenter Anne Marie Downey 10h00PhD SrP25

Title Zinc Supplementation Reduces Cyclophosphamide Induced Oxidative Stress and DNA Damage In Male Germ Cells

Authors Downey Anne Marie Hales Barbara F Robaire Bernard

Abstract Zinc is required for numerous enzymes and DNA binding proteins involved in oxidative stress defense and DNA damage repair A previous study in our lab suggests that cyclophosphamide (CPA) an alkylating agent and known germ cell toxicant alters zinc homeostasis in germ cells contributing to the damaging effects of the drug We hypothesize that zinc supplementation will protect germ cells from CPA induced oxidative stress and DNA damage Adult male Sprague Dawley rats received one of four treatments 6 daysweek for five weeks saline ZnCl2 (20mgkg) saline only for the first week then with CPA (6mgkg) for 4 weeks or ZnCl2 (20mgkg) only for the first week then with CPA (6 mgkg) for 4 weeks Fluorescent imaging of purified germ cells using CellRox a marker for reactive oxygen species (ROS) revealed that ROS levels were increased in pachytene spermatocytes from animals treated with CPA compared to controls Zinc supplementation in CPA treated animals reduced ROS levels to those of controls A similar trend was observed in round spermatids DNA damage as assessed by the mean intensity of immunofluorescent staining for ɣH2Ax in testis sections was increased after CPA treatment and foci size were more widely distributed compared to controls Zinc supplementation resulted in a trend towards a decrease in ɣH2AX signal when compared to CPA treatment alone and foci size distribution resembled that of controls Although further studies are needed these data suggest a potential role for zinc supplementation in protecting male germ cells against CPA insult

Acknowledgments Supported by CIHR

__________________________________________________

Presenter Sean Jmaeff 10h00PhD SrP27

Title Small molecule novel RET agonistic ligands activate biased neuroprotective signals that unlike GDNF are GFRa1ndash independent and synergize with FGF2

Authors Sean Jmaeff Yulia Sidorova Hayley Lippiatt Pablo F Barcelona Hinyu Nedev Mark A Hancock Mart Saarma H Uri Saragovi

Abstract Glial cell line-Derived Neurotrophic Factor (GDNF) binds to the GFRa1 receptor and the GDNFndashGFRa1 complex binds to and activates the transmembrane RET tyrosine kinase to transduce signals through AktErkSrc pathways To dissect the GDNFndashGFRa1ndashRET signalling complex we developed agents that bind and activate RET directly and independent of GFRa1 expression From a focussed library of aminonaphthalene-sulfonic acids we identified small molecules that are genuine ligands that bind to the RET extracellular domain These ligands activate RET kinase irrespective of GFRa1 co-expression afford biased trophic signals and synergize with basic Fibroblast Growth Factor 2 (FGF2) RET activation is constrained by GFRa1 likely via an allosteric mechanism that can be overcome by increasing RET ligand concentration or by using GFRa1 modulators In a mouse model of Retinitis Pigmentosa monotherapy with a small molecule RET agonist activates survival signals and reduces neuronal death significantly better than GDNF suggesting therapeutic potential

Acknowledgments This work was supported by a CIHR grant to HUS and the Lundbeck Foundation and Sigrid Juseacutelius Foundation to MS We are thankful for reagents provided by Dr Brian Pierchala (pRET antibodies) Dr Eero Castren (MG87Trk cell lines) Dr Carlos Ibanez (MG87RET cell lines) and Dr T Li (RHOP347S transgenic mice) Jenni Montonen assisted with the Ret phosphorylation assays Dr Enrique de la Rosa assisted with retinal explants and TUNEL assays Ms Lucia M Saragovi counted and quantified TUNEL assays The McGill SPR-MS Facility thanks the Canada Foundation for Innovation (CFI grant 228340) for the BIACORE T200 SPR infrastructure

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Presenter Baraa Noueihed 10h00PhD SrP29

Title Mesenchymal stem cells repair retinal vascular damage in Retinopathy of Prematurity mouse model

Authors Noueihed Baraa Rivera Jose Carlos Chemtob Sylvain

Abstract Objective Retinopathy of prematurity is a leading cause of visual impairment in infants Exposure of premature babies to the hyperoxic extrauterine environment leads to vascular decay known as vaso-obliteration (VO) Ischemia subsequently ensues and triggers pathological intravitreal neovascularization (NV) in the immature retina Current treatments only target the aberrant neovessels without repopulating the avascular regions Thus there is a dire need for new therapies that promote healthy vessel growth Mesenchymal stem cells (MSCs) have shown the ability to migrate to the damaged tissue in different animal models and enhance vascularization We therefore investigated whether MSCs can promote vascular repair of ischemic retinas Methods Oxygen-induced retinopathy (OIR) model was used herein Postnatal day 7 (P7) mice were subjected to 75 O2 until P12 to induce VO followed by 5 days of room air leading to NV Compact bone-derived MSCs were isolated from adult mice and cultured either in hypoxia (5 O2) or normoxia (21 O2) Conditioned media (CM) was collected 24hours later and injected intravitreally in P12 OIR retinas to assess vascular repair To determine possible factors involved in MSC-induced revascularization qPCR was performed on P17 OIR retinas In vitro we investigated the effect of MSC-CM on microglial polarization using qPCR and flow cytometry Results Hypoxic MSC-CM significantly decreased both VO and NV areas Levels of IGF-1 and VEGF were significantly high in MSC-injected retinas Moreover mRNA levels of pro-inflammatory cytokines (IL-1β TNFα) dropped whereas levels of anti-inflammatory cytokines (IL-10 IL-4) increased Treatment of pro-inflammatory M1 microglia with MSC-CM decreased the gene expression of IL-1β TNFα and iNOS (M1 marker) Conclusion In this study we demonstrated that MSCs promote healthy vessel growth in OIR retinas via a paracrine fashion by regulating expression of angiogenic factors and modulating inflammation

Acknowledgments FRSQ Reseau Vision

Presenter Gauthier Schang 10h00PhD SrP31

Title Loss of GATA2 impairs FSH production in male mice in vivo

Authors Schang Gauthier Brucircleacute Emilie Boehm Ulrich Bernard Daniel J

Abstract Mammalian reproduction is dependent on follicle-stimulating hormone (FSH) and luteinizing hormone (LH) secreted by pituitary gonadotrope cells The dimeric hormones share a common α-subunit (CGA) linked to hormone-specific β-subunits (LHβ and FSHβ) Relative to LHβ the mechanisms regulating FSHβ (Fshb) synthesis are poorly resolved Activins are potent and selective regulators of Fshb transcription and act at least in part via the transcription factors FOXL2 SMAD3 or SMAD4 However all three of these proteins are co-expressed in other tissues that do not produce FSH indicating that they alone are insufficient to explain mechanisms of Fshb expression Here we will test the hypothesis that GATA2 plays a fundamental role in FSH synthesis by gonadotrope cells This hypothesis derives from at least three observations First the mature gonadotrope-like cell line LβT2 expresses both Gata2 and Fshb while the immature cell line αT3-1 expresses neither Second Gata2 is the only member of the GATA factor family expressed in gonadotropes Third conditional deletion of Gata2 in mouse pituitaries decreases serum FSH levels though the gene was deleted in multiple pituitary lineages precluding an assessment of GATA2rsquos specific role in gonadotropes As such we crossed floxed Gata2 animals with GRIC mice which only express the Cre recombinase in gonadotropes Resulting knockout animals (cKO) were compared to littermate controls cKO females exhibited normal timing of vaginal opening (ie puberty) estrous cyclicity and fertility cKO males exhibited ~50 lower serum FSH levels which were paralleled by reduced pituitary Fshb and Cga mRNA expression levels as well as by a decrease in testicular weight Seminal vesicle weight was unchanged as was pituitary Lhb expression These data suggest that Gata2 expression in gonadotropes is dispensable for female fertility but essential for quantitatively normal FSH production in males Acknowledgments The authors would like to acknowledge undergraduate students Nittha Lalang and Jonas Lehnert for their ontribution as well as Ying Wang and Xiang Zhou for their technical assistance This work was funded by CIHR operating grant MOP-133394 and fellowship 152308 FRQS fellowship 31338 and the Dr Samuel Solomon Fellowship in Endocrinology

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Presenter Noosha Yousefpour 10h00PhD SrP33

Title Role of microglia in structural plasticity of touch circuitry in neuropathic pain

Authors Yousefpour Noosha Aparicio Maria Ribeiro-da-Silva Alfredo

Abstract Injury or disease affecting the somatosensory nervous systems often results in the development of chronic neuropathic pain This can cause long-lasting pain and sensory abnormalities such as pain in response to innocuous mechanical stimulation (mechanical allodynia) One of the proposed mechanisms underlying mechanical allodynia is reduced dorsal horn inhibitory control of touch input transmitted by myelinated low-threshold mechanoreceptors (A-LTMRs) In this study we performed a detailed anatomical study to assess inhibitory synaptic changes with respect to A-LTMRs and dorsal horn neurons in a model of neuropathic pain We hypothesize that mechanical allodynia is maintained by spinal cord disinhibition mediated by selective loss of synapses between A-LTMRs and GABAergic inhibitory neurons and this selective loss is mediated by the activity of microglia and components of the complement cascade Using light and electron microscopy we assessed changes in inhibitory appositions on dorsal horn neurons and synapses on A-LTMR central terminals in a rat model of neuropathic pain We showed that following nerve injury there was a reduction in the number A-LTMR central terminals and a loss of inhibitory synapses in those that remained A-LTMR terminals showed signs of degeneration and glia profiles were observed to partially or completely surround these degenerating terminals Moreover complement factor C1q was upregulated in microglia and co-localized with inhibitory synapses in the ipsilateral side of the dorsal horn of neuropathic animals Chronic inhibition of microglia using minocycline reduced mechanical allodynia and prevented the loss of inhibitory appositions on dorsal horn neurons This study provides evidence that microglia contribute to mechanical allodynia by removing dorsal horn inhibitory synapses through a complement dependent pathway in a model of neuropathic pain

__________________________________________________

Presenter Mariana Asslan 10h00MSc Jr35

Title Promising combinatorial approach to treat CLL by combining TPL2 and BCL2 inhibitors

Authors Asslan Mariana Rousseau Simon

Abstract Therapeutic strategies for treating CLL have significantly changed from using alkylating agents to novel therapeutics BCL2 inhibitors BH3 mimetics represent an attractive treatment for lymphoid tumors as BCL2 proteins are a major promoter of cell survival that are mostly overexpressed in CLL Moreover TPL2 inhibitors have been considered an appealing avenue to reduce the proliferation of malignant cells via blockade of MAPKERK pathway a pathway involves a series of protein kinase cascades playing a critical role in regulating cell proliferation The aim of this study is to investigate the effectiveness of dual BCL2 and TPL2 inhibition on CLL cells As we hypothesize that the association of BH3 mimetics such (ABT199 and ABT263) and the TPL2 inhibitors (C1 and C34) could be a more efficacious clinical strategy than single target therapy MTT (cell viability) assay was performed on OCI-ly2 cell line MTT data from this study showed that targeting both pathways simultaneously with ABT263 and C1 or C34 has synergistically enhanced tumour apoptosis in vitro with no toxic effects on normal cells This study provides insight into innovative strategy and suggests that combination between BCL2 and TPL2 inhibitors acts synergistically so it might help to improve the treatment of CLL in the future

Acknowledgments Funding agency Canadian Institute of Health Research Rousseau Lab Dr Simon Rousseau

__________________________________________________

Presenter Mary Loka 10h00MSc Jr IPNP37

Title The Efficacy of Oral Versus injectable Administration of Analgesics

Authors Mary Loka Chulmin Cho Vassilia Michailidis Matthew Danesh Loren Martin

Abstract In several fields of research animals often undergo surgeries as part of manipulations to answer certain scientific questions As such research institutions have established committees that set mandates regarding the administration of analgesia to laboratory rodents undergoing invasive procedures These are typically administered via injection which can induce stress and alter an animals behaviour Administering medication in an animalrsquos water may be more effective at alleviating post-surgical pain since it is a less invasive and remains readily accessible as opposed to receiving medication all at once through injection For this experiment we compare the efficacy of Carprofen Meloxicam Buprenorphine and saline given both orally and through injection for alleviating craniotomy pain in mice We test these using minimally invasive measures of well being including the Mouse Grimace Scale to look at spontaneous pain Results have shown that control animals expressed the most pain behavior on surgery day as compared to animals receiving analgesics though injections seem more effective in mitigating the pain response than oral administration We also show that buprenorphine is the most effective for treating pain in both males and females though the pain trajectories for other drugs differed between the sexes

Acknowledgments This project was supported by NSERC and by the University of Toronto Mississauga Undergraduate Research Grant

POSTER SESSION II 14h30 ndash 16h00

P2 Emilie Brule MSc Junior Anatomy and Cell Biology

P4 Valerie Bourassa PhD Junior IPN

P6 Samantha Locke PhD Senior IPN

P8 Celia Bouazza MSc Junior Pharmacology

P10 Iulia Pirvulescu MSc Junior Pharmacology

P12 Xinwen Zhu MSc Junior Pharmacology

P14 Jeff Ji MSc Senior Pharmacology

P16 David McCusty MSc Senior Pharmacology

P18 Kyla Bourque PhD Junior Pharmacology

P20 Colin Cheng PhD Junior Pharmacology

P22 Jenna Giubilaro PhD Junior Pharmacology

P24 Anne-Sophie Peacutepin PhD Junior Pharmacology

P26 Morgan Foret PhD Senior Pharmacology

P28 Jace Jones-Tabah PhD Senior Pharmacology

P30 Sandra Paschkowsky PhD Senior Pharmacology

P32 Heather Fice PhD Junior Pharmacology

P34 Issan Zhang PhD Senior Pharmacology

P36 Helene Hall Postdoctoral Fellow Pharmacology

P38 Daniel Sapozhnikov PhD Junior Pharmacology

__________________________________________________

Presenter Emilie Brucircleacute 14h30MSc Jr Anat amp CellP2

Title Does IGSF1 play a role in thyroid hormone transport

Authors Brucircleacute Emilie Silander Tanya Wang Ying Bernard Daniel J

Abstract The hypothalamic-pituitary-thyroid axis regulates growth development and metabolism In this axis hypothalamic thyrotropin-releasing hormone (TRH) stimulates the secretion of pituitary thyroid-stimulating hormone (TSH) TSH stimulates thyroid hormone (TH) production by the thyroid gland THs composed of the prohormone T4 and its bioactive metabolite T3 negatively regulate expression of the TSH subunit and TRH receptor (Trhr1) genes in pituitary thyrotrope cells We recently reported that mutations in the X-linked immunoglobulin superfamily member 1 (IGSF1) gene cause a novel form of congenital central hypothyroidism In two mouse models of IGSF1-deficiency we observed reduced pituitary Trhr1 resulting in impaired TRH stimulation of TSH secretion Reduced Trhr1 may derive from enhanced sensitivity to TH negative feedback in Igsf1 knockout mice TH action is regulated by hormone transport metabolism andor receptor activation A potential role for IGSF1 in TH transport was suggested by our observation that IGSF1 physically interacts with a TH transporter monocarboxylate transporter 8 (MCT8) However MCT8-mediated transport of THs was unaffected by co-expression of IGSF1 in heterologous cells Moreover MCT8 expression is low in murine pituitary We therefore hypothesized that IGSF1 functionally interacts with and inhibits the activity of a MCT8-related TH transporter In the absence of IGSF1 this transporter can more readily transport THs into cells leading to greater suppression of Trhr1 However in vivo wild-type and Igsf1-knockout mice exhibit similar responses to exogenous T3 challenging this idea Since TH transporters can have different affinities for T3 or T4 we will test the sensitivity of Igsf1-knockout mice to exogenous T4 In conclusion although IGSF1 can physically interact with MCT8 this interaction is not functional in vitro Moreover our current in vivo data suggest that mice do not transport T3 differently in the absence of IGSF1

Acknowledgments Funding Source Supported by CRRD (EB) NSERC (EB) ENDO (EB) CIHR (DJB)

__________________________________________________

Presenter Valerie Bourassa 14h30PhD Jr IPNP4

Title Studies of Mechanisms of Osteoarthritis using Intra-Articular Injection of MIA in the Rat Ankle Joint

Authors Bourassa Valeacuterie Deamond Haley Ribeiro-da-Silva Alfredo

Abstract Osteoarthritis (OA) is a complex disease of the whole joint and affects over 3 million Canadian adults Typically OA patients have mechanical hypersensitivity and cold allodynia most often in the knee ankle hip or shoulder joints To this day there is no satisfactory method of relieving OA pain One of the most used animal models of OA in the rodent is the intra-articular injection in the knee joint of the chondrocyte glycolytic inhibitor Mono-Iodoacetate (MIA) This project characterizes the behavioral and histopathological changes of cartilage necrosis and degeneration that occur in a new model of MIA-induced OA in the rat ankle joint which we believe will more closely replicate the clinical symptoms of the human disease We hypothesize that the changes in OA are comparable to those previously described by our group for inflammatory arthritis including the occurrence of anomalous sensory and sympathetic fibre sprouting in joints and adjacent skin although appearing at later time points A dose of 24 mg of MIA in 40 microl of saline administered via intra-articular injection in the right ankle joint induced significant mechanical hypersensitivity and cold allodynia at 5 weeks post injection This is important as cold allodynia had not been previously reported in the rat OA knee joint Histological studies of decalcified joints showed significant cartilage loss correlating with the behavior pattern and replicating known pathologic features of the human disease X-ray microtomography scans of the ankle joint performed at 5 and 10 weeks confirmed that bone fragmentation and remodeling in our MIA model are similar to those in human OA Furthermore preliminary results show an increase innervation of sympathetic fibers of the subchondral bone of the talus at 10 weeks as detected by DAB immunohistochemistry against VMAT2 confirming our prediction that anomalous sprouting correlates with pain states in the disease and is likely to contribute to pain in OA

Acknowledgments We would like to thank the funding agencies Dr Ribeiro-da-Silvas CIHR operating grant and the Louise and Alan Edwards Foundationrsquos Edwards PhD Studentship

__________________________________________________

Presenter Samantha Locke 14h30PhD Jr IPNP6

Title Forced walking results in pain-related changes in a rat model of inflammatory arthritis

Authors Locke Samantha YOUSEFPOUR Noosha MANNARINO Matthew RIBEIRO-DA-SILVA Alfredo

Abstract Pain upon movement is a main complaint of arthritis patients and in inflammatory arthritis inflammation and pain do not always correlate suggesting that a mechanism other than overt peripheral inflammation is driving pain There is an increasing recognition of a neuropathic component of arthritis pain and there is a link between arthritis patients that have an identified neuropathic component to their pain and greater movement-related issues During chronic pain such as in arthritis pain does not only arise in the periphery there is also a contribution of central sensitization In models of arthritis changes in neuronal activation in the spinal dorsal horn related to nociception have been shown but have not yet been investigated in freely moving animals Following intra-articular CFA injection to the ankle joint weight bearing assessment was performed before and after forced walking on a treadmill Sham and CFA-treated animals were divided into forced walking or non-walking groups At 4 weeks post-CFA 2 hours after forced walking animals were sacrificed and processed for immunohistochemistry Spinal neuronal activation was assessed using anti-fos antibodies a marker of neuronal activation and inhibitory interneurons detected with anti-Pax2 antibodies Weight bearing showed a greater distribution of weight to the uninjured paw following walking as compared to non-walking animals In the dorsal horn fos expression was maximal in walked CFA-treated rats compared to non-walked CFA-treated animals with the least fos expression in sham animals The number of inhibitory interneurons positive for fos immunoreactivity was decreased in CFA-treated animals that underwent walking compared to sham animals that walked These data suggest that forced walking induces changes in the spinal cord and weight bearing reflective of a pain state and that neuropathic-changes may underlie pain in relation to movement in this model

Acknowledgments Research funded by CIHR grant MOP-136903 and Samantha Locke was a recipient of a studentship from the Louise and Alan Edwards Foundation

__________________________________________________

Presenter Celia Bouazza 14h30MSc JrP8

Title The effects of Gβγ signaling on global RNA synthesis in cardiac fibroblasts

Authors Bouazza Ceacutelia Khan Shahriar Martin Ryan Heacutebert Terry

Abstract Heterotrimeric G proteins act as intermediates in G protein-coupled receptor signaling relaying extracellular signals received by cell surface receptors to downstream signaling pathways Our lab has shown that Gβγ subunits act as direct regulators of multiple signaling pathways and can localize to many intracellular organelles including the nucleus In the nucleus we have uncovered a novel interaction of Gβγ subunits with Rpb1 the largest subunit of RNA polymerase II (RNAPII) but the function of this interaction remains incompletely characterized Cardiac hypertrophy and fibrosis major features of heart diseases occurs when the cardiomyocytes and fibroblasts are continuously exposed to external stimuli Our first goal was to undertake a qPCR array-based analysis of fibrotic genes in rat neonatal cardiac fibroblasts in response to angiotensin II (Ang II) We next examined the impact of knocking down Gβ1- and Gβ2shycontaining Gβγ dimers Upon RNAi knockdown of Gβ12 we observed increases in basal levels of many genes upregulated in cardiac fibrosis From there we sought to understand the impact of knocking down these dimers on global gene expression measured using incorporation of ethynyl uridine RNAi knockdown of Gβ1 and Gβ2 individually or together revealed their potential involvement in global RNA transcription upon stimulation of the AT1R Knockdown of Gβ12 resulted in increased basal levels of global RNA transcription and inhibited the Ang II-induced increase seen under control conditions Taken together our data suggest a novel role for Gβ12γ as critical modulators of global transcription in primary cardiac fibroblasts This further supports the effects on gene expression modulated by Gβγ association with RNAPII which may serve a role in cardiac pathologies Identifying which pairs of Gβγ dimers are relevant in our model and how different combinations may impact transcription could potentially determine novel targets in treating heart disease

Acknowledgments This work was supported by grants from The Heart and Stroke Foundation

__________________________________________________

Presenter Iulia Pirvulescu 14h30MSc JrP10

Title Mechanisms of action of the truncated TrkCT1 neurotrophin receptor

Authors Pirvulescu Iulia Brahimi Fouad Pidgeon Reilly Saragovi Uri

Abstract Neurotrophins play a key role in the growth and function of neurons They activate Trk and p75 receptors where Trk are considered to be neuroprotective and p75 activation leads to proshyapoptotic signaling a process initiated by p75 cleavage The neurotrophin NT-3 binds to TrkC TrkC-full length (TrkC-FL) has tyrosine kinase activity that promotes trophic signaling TrkC is alternatively-spliced into truncated receptor TrkCT1 which lacks the tyrosine kinase domain The up-regulation of TrkCT1 has been noted in neurodegenerative disorders Work in our laboratory showed that TrkCT1 leads to production of the neurotoxic cytokine TNF-α I aim to uncover ways in which the ldquoneuroprotectiverdquo TrkC-FL and the ldquoneurotoxicrdquo TrkCT1 cooperate with and cross-regulate with p75 co-receptors I used cells expressing p75 TrkC-FL or TrkCT1 alone as well as p75 co-expressed with each TrkC isoform Co-IP showed interaction between each isoform and p75 The cells were treated with combinations of a p75 antagonist the neurotrophin NT-3 and pro-neurotrophin proNGF the latter of which only has affinity for p75 Apoptotic signaling in response to the ligands was assessed via MTT cell viability assay and Annexin-PI staining p75 activation and proteolytic cleavage were studied through biochemical analysis We found that p75 cleavage occurs in ligand-dependent and independent ways Ligand-independent cleavage of p75 is significantly increased in cells co-expressing TrkCT1 whereas it is decreased in the presence of TrkC-FL Unlike TrkCT1 ligand-dependent activation of the tyrosine kinase-containing TrkC-FL enhances cleavage of p75 We show that the isoforms of TrkC functionally interact with p75 differently leading to increased paracrine neurotoxic signals Hence it is of interest to further our study on the specific biology of the truncated isoform of TrkC We believe that continuing this study may uncover implications relevant in neurodegenerative diseases

Acknowledgments This work is funded by CIHR

__________________________________________________

Presenter Xinwen Zhu 14h30MSc JrP12

Title P-TEFb dependent pathways in cancer

Authors Zhu X Alattar R Tanny JC Heacutebert TE

Abstract Regulators of transcriptional control are crucially involved in the progression of cancer For instance mixed lineage leukemia (MLL) has been shown to be caused by fusion proteins which induce abnormal transcriptional elongation Following transcription initiation RNA polymerase II (RNAPII) can be maintained in a promoter proximally paused state by two associated protein complexes DRB sensitivity inducing factor (DSIF) and negative elongation factor (NELF) Entrance of RNAPII into productive elongation requires the activity of positive transcription elongation factor b (P-TEFb) a cyclin-dependent kinase which phosphorylates the Spt5 subunit of DSIF and a component of NELF as well as RNAPII itself To determine the functional significance of P-TEFb dependent pathways and their relevance in various cancers we are examining the effects of various small molecule inhibitors of P-TEFb pathway elements on the proliferation and viability of established cancer cell lines Preliminary results suggest that cells which carry MLL fusion proteins show increased sensitivity to such inhibitors We are also investigating the relationship between histone 2B monoubiquitination and Spt5 phosphorylation To this end we are using CRISPRCas9 to knock phosphorylation-deficient Spt5 mutants in to cancer cells Elucidation of the mechanisms underlying P-TEFb dependent cancer progression will lead to the identification of potential therapeutic targets

Acknowledgments We would like to thank Julien Leconte for his assistance and guidance with the flow cytometry This work is supported by funding from the Canadian Institutes of Health Research the Heart and Stroke Foundation of Canada and the Faculty of Medicine of McGill University

__________________________________________________

Presenter Jeff Ji 14h30MSc SrP14

Title Impact of gold nanostructures on neuroglia and synaptic remodeling

Authors Ji J Moquin A Chang PK Winnik FM McKinney RA Maysinger D

Abstract Gold nanoparticles have attracted considerable interest as delivery vehicles and imaging agents due to their surface tunability and optical properties As therapeutic agents gold nanoparticles are used for the photothermal ablation of gliomas and as contrast agents in the diagnosis of neurodegenerative diseases Although the effect of gold nanoparticles on peripheral tissue and immortalized cell lines are well studied how gold nanoparticles affect the central nervous system is largely undocumented We investigate the effect of gold nanoparticle morphologies (clusters spheres rods and flowers) and coating (PEG GSH CTAB) on synaptic structures in the hippocampus an area of brain involved in learning and memory By employing organotypic hippocampal cultures generated from transgenic mice expressing membrane-tagged green fluorescent protein in a subset of excitatory neurons we studied the impact of gold nanoparticle shapes and on dendritic spine density and morphology Furthermore we investigated the uptake and toxicity of gold nanoparticles in enriched primary astrocyte microglia and neuronal cultures We show that spine populations are unaffected by medium sized (10-100 nm) gold nanoparticles coated with PEG or GSH despite extensive intracellular uptake but are adversely affected by gold nanoparticles coated with a synthetic precursor CTAB Interestingly small-sized (lt2 nm) gold nanoclusters significantly lowered total spine density with the mature mushroom spines being particularly susceptible In summary dendritic spines are sensitive to both size and surface coating of gold nanoparticles Future research will focus on exploring the intracellular effects of gold nanoclusters which led to changes in spine populations and its effect on different neural cell types

Acknowledgments We gratefully acknowledge CIHR and NSERC for funding this study We also thank Francois Charon for his technical assistance in preparing the organotypic cultures

__________________________________________________

Presenter David McCusty 14h30MSc SrP16

Title Stimulus-Responsive Imaging of Mineralized Tissue

Authors McCusty David LeBlanc Emanuelle Kurdieh Reem Castagner Bastien

Abstract Fluorescent probes are widely to investigate biological processes A major limitation in their use however is the delivery of the probe to the location of interest and maintaining the probe in that location long enough to be able to detect a useful fluorescence signal To mitigate this issue a fluorescent probe conjugated to the bone-targeting drug alendronate via a maleimide linker has been synthesized and successfully purified which facilitates probe binding to the inorganic hydroxyapatite mineral in bone tissue By using a Foumlrster resonance energy transfer (FRET) internal peptide the probe is also made stimulus responsive Upon enzymatic cleavage of the peptide by the protease Cathepsin K a quencher is separated from a fluorescent dye resulting in an increase of fluorescence which is measured in a fluorescence plate reader or imaged using fluorescent microscopy Cathepsin K is an important protease as it is the major protease expressed by osteoclasts the only cell capable of resorbing bone tissue and it is largely responsible for the break down of the organic collagen tissue in bone This probe is designed to not only indicate the presence of cathepsin K but it is also capable of detecting its activity important for proteases like Cathepsin K with both active and inactive forms Furthermore the probe is also a proof-of principle of a bone-targeting probe that can be applied to other proteases by changing the amino acid sequence of the peptide different proteases could be investigated using a similar design While the alendronate-conjugated peptide has been successfully synthesized and purified in vitro biological assays have proven challenging and remain in progress

Acknowledgments Dr Mark Hancock Dr Svetlana Komarova Zamboni Chemical Solutions NSERC

Presenter Kyla Bourque 14h30PhD JrP18

Title G proteins and mimetic nanobodies as allosteric modulators of the β2AR

Authors Bourque Kyla Peacutetrin Darlaine Sleno Rory Devost Dominic Heacutebert Terry

Abstract As highly dynamic proteins G protein-coupled receptors (GPCRs) attain a plethora of different conformational states with certain conformations linked to specific signal transduction pathways The connection between receptor conformation and activity is not completely understood but can be studied using bioluminescence resonance energy transfer (BRET) We used the β2-adrenergic receptor as a model system and introduced the FlAsH binding tetracysteine tag along different points on the intracellular surface of the β2AR with a C-terminally fused Renilla luciferase as pairs for BRET to monitor receptor conformation in real-time Our panel of eight β2AR FlAsH BRET-based biosensors were expressed in a parental HEK 293 cell line and in a ∆Gαs CRISPRCas9 KO cell line In the parental line the response to isoproterenol was transient with respect to conformational changes in ICL2 and ICL3 while more robust responses were captured in two of the three C-tail positions However in the ∆Gαs line all three ICL3 biosensor positions showed a sustained increase in BRET upon isoproterenol stimulation These results suggest that the G protein Gαs may act as an allosteric modulator of β2AR conformation and function given that two distinct conformational signatures were observed in its presence and absence Additionally we are also screening various nanobodies (Nb60 71 74 80 84) in order to better understand their roles as G protein mimics and stabilizers of specific receptor conformations Their expression in parallel with the conformation-sensitive biosensors have shown that in response to isoproterenol Nb60 primes the allosteric stabilization of a distinct receptor state whereas the allosteric effect of the other four nanobodies were not as robust As such exploring their effect using the other conformational perspectives might guide us to understand how nanobodies modulate receptor conformation and function Acknowledgments This work was supported by the Canadian Institutes for Health Research We also thank Jan Steyaert (VIB Belgium) for the generous gift of the different nanobodies and Asuka Inoue for the ∆Gαs cell line

__________________________________________________

Presenter Colin Cheng 14h30PhD JrP20

Title IL-1 Modulators From Pre-term Labour to Retinopathy of Prematurity

Authors Cheng Colin Rivera Carlos Hou Xin Geran Azade Nadeau-Valleacutee Mathieu Quiniou Christiane Lubell William Chemtob Sylvain

Abstract Preterm birth (before 37 weeks gestation) affects up to 18 of all pregnancies and over 40 of cases are attributed to inflammation It is associated with many sequelae including retinopathy of prematurity (ROP) a major cause of blindness in infants Interleukin-1 (IL-1) is an inflammatory cytokine that contributes to both premature labour in pregnant women as well as the retinopathy that subsequently develops in the neonate Current therapies to prevent or treat premature labour and ROP including competitive antagonists of IL-1 are only modestly effective and are associated with adverse effects To this end our lab has developed rytvela a heptapeptide and an allosteric modulator of the IL-1 receptor rytvela has been previously shown to prevent preterm labour and confer protection against retinal vaso-obliteration a hallmark sign of retinopathy To better characterise the structural-activity relationships of rytvela we developed a library of derivatives with modifications in chirality at threonine-3 and valine-4 and a lactam ring cyclisation of threonine-3 We then tested these derivatives in our established murine models of preterm labour and oxygen-induced retinopathy While the efficacy of our derivatives varied our best compound AGU25 is at least as effective as rytvela in delaying preterm labour and preventing retinal vaso-obliteration We also observed that efficacious compounds possessed R-chirality in the αshycarbons of valine-4 and the lactam ring These results offer early insights into structure-activity relationships of rytvela and will complement future structural and signalling transduction studies that are required prior to submitting our derivatives for clinical testing

Acknowledgments I would like to express my thanks to my supervisor Dr Sylvain Chemtob and my co-supervisor Dr Alfredo Ribeiro-da-Silva as well as my thesis committee members Dr Barbara Hales Dr Jean-Francois Trempe and Dr Sylvie Girard I would also like to thank the current and recent members of the Chemtob Lab for their support Alexandra Beaudry-Richard Ankush Madaan Amarilys Boudreault Baraa Noueihed Estefania Marin Sierra(Ellen) Tianwei Zhou Franccedilois Duhamel Houda Tahiri Isabelle Lahaie (Joanna) Xiaojuan Yang Mohammad Ali Mohammad Nezhardy Prabhas Chaudhari Rabah Dabouz Samy Omri Tang Zhu and Vikrant Bhosle

__________________________________________________

Presenter Jenna Giubilaro 14h30PhD JrP22

Title Characterizing a Novel Inhibitor of Receptor Trafficking and Intracellular Signaling

Authors Giubilaro J Namkung Y Laporte SA

Abstract G protein-coupled receptors (GPCRs) are the largest family of cell surface receptors and the target of 40 of therapeutic drugs Upon agonist stimulation GPCRs can signal via G-protein coupling or via a β-arrestin-dependent mechanism β-arrestins are adaptor proteins that bind to activated receptors to desensitize G protein signaling promote receptor internalization and activate signal transduction cascades such as MAPK and PI3K that can be independent of G protein coupling However there are currently no well-characterized tool compounds to assess the mechanisms by which β-arrestins mediate receptor internalization and intracellular signaling Therefore we used trafficking Bioluminescence Resonance Energy Transfer (BRET)shybased biosensors to run a high-throughput phenotypic screen and selected trafficking molecule 21 (Traf 21) for its inhibitory effects on angiotensin II type 1 receptor (AT1R) internalization in HEK293 cells Using BRET biosensors we show that Traf 21 does not affect βshyarrestin recruitment to receptors nor the formation of the βshyarrestin2AP-2 complex in clathrin-coated pits By western blot we show that Traf 21 blocks receptor-mediated RasERK and PI3KAkt pathways We use various BRET biosensors to look upstream of the MAPK pathway and show that AT1R can still activate Gq protein PKC and PLC in the presence of Traf 21 By western blot we show that MEK1-mediated phosphorylation of ERK is not affected Using various BRET biosensors we show that Traf 21 inhibits the activation of small G proteins Ras and Arf6 but not Rho and Rac consistent with their roles in cellular signaling and membrane trafficking respectively

Investigating how Traf 21 pharmacologically modulates responses downstream of GPCRs may lead to a better understanding of how receptor internalization and intracellular signaling are mediated and may address their physiological role in pathologies such as cardiovascular disease and cancer

Acknowledgments This work was supported by the CIHR and the RQRM

__________________________________________________

Presenter Anne-Sophie Peacutepin 14h30PhD JrP24

Title Using a transgenic mouse model combined with an environmental challenge (high-fat diet) to delineate molecular mechanisms implicated in epigenetic inheritance

Authors Peacutepin A-S Lafleur C Tanwar DK Kennedy K Ribeiro T Sloboda DM Kimmins S

Abstract Within the last four decades an obesity epidemic has spread worldwide It is well established that genetics and lifestyle play an important role in the development of obesity Nonetheless epidemiological studies and diet-induced obesity rodent models have shown that overweight fathers are more likely to have overweight children even if the child consumes a healthy diet The underlying molecular mechanisms are still poorly defined We hypothesized that histone methylation in sperm is one such mechanism implicated in the epigenetic inheritance of obesity and associated metabolic disorders To test this hypothesis we used our existing transgenic mouse model with an altered sperm epigenome and demonstrated transgenerational inheritance of abnormal development and reduced survivability (Siklenka et al 2015) We aimed to assess whether there are cumulative deleterious effects on histone methylation in sperm resulting from an environmental challenge (high-fat diet) that would lead to enhanced phenotypes in transgenic offspring Transgenic non-transgenic littermates and C57BL6NCrl control males (n=10 per group) were fed either a low- or high-fat diet (10 or 60 kcal fat respectively) for 12 weeks starting at weaning Males were then bred to chow-fed C57BL6NCrl females All offspring were fed a chow diet Fathers and offspring metabolic function was assessed As expected high-fat fed males became obese with glucose intolerance reduced insulin sensitivity and elevated fasting blood glucose Sex- and genotype-specific metabolic disturbances in offspring associated with paternal diet were observed ChIP-sequencing targeting H3K4me3 in sperm revealed changes associated with diet These findings implicate histone methylation in sperm in environment-induced heritable phenotypes

Acknowledgments CIHR CRRD RQR

Presenter Morgan Foret 14h30PhD SrP26

Title Understanding Oxidative Stress in Early Alzheimers Disease

Authors Foret M Do Carmo S Greene L Lincoln R Zhang W Cosa G Cuello AC

Abstract The earliest stages of Alzheimers disease (AD) occur decades before clinical symptoms are present and remain elusive with no cures nor effective preventative strategies for AD currently available To unravel these early molecular events driving AD the Cuello lab generated the McGill-R-Thy1-APP transgenic (Tg) rat model which exhibits a progressive AD-like amyloid pathology In this model we have observed an accumulation of intraneuronal amyloid beta (iAβ) during the early silent stages of the disease well before extracellular Aβ plaque deposition and cognitive deficits This project aims to shed light on the earliest preshysymptomatic pathology that drives amyloid-associated molecular events occurring in AD More specifically we aim to investigate how iAβ drives production of reactive oxygen species (ROS) Importantly ROS and subsequent oxidative stress has been increasingly associated with neurodegenerative diseases and has shown to be increased in AD patients albeit at later stages of the pathology The role of ROS during early disease progression in AD has yet to be understood We hypothesize that the early iAβ accumulation triggers a pathological cascade leading to increased ROS production and subsequent mitochondrial dysfunction which then culminates in the well known AD pathology The main objectives of this project are to determine when where and how this cascade occurs In collaboration with Dr Cosa in the McGill Chemistry department we are using novel fluorescent probes to detect ROS production in primary neuronal cultures and ex vivo hippocampal slices of Tg and wild type (Wt) rats Preliminary results presented here demonstrate live cell imaging in primary neurons using total internal reflection fluorescence microscopy and characterization of these cultures using immunocytochemistry Future aims of the project include applying these probes to ex vivo slices at various time points in the AD-like amyloid pathology of the Tg rat Acknowledgments This work was supported by the Canadian Institute of Health Research Acknowledgement to Dr Bowie and his lab for their assistance and guidance in producing acute hippocampal slices as well as those at the Advanced BioImaging Facility (ABIF) including

__________________________________________________

Presenter Jace Jones-Tabah 14h30PhD SrP28 Title Fiber-optic recording of FRET biosensors for detecting GPCR signalling in vivo

Authors Jones-Tabah Jace Benaliouad Faiza Clarke Paul Heacutebert Terry

Abstract G protein-coupled receptors (GPCRs) mediate neuronal responses to neurotransmitters and neuromodulators and are major drug targets in neuropsychiatric disease Individual GPCRs signal via multiple downstream effectors only some of which may mediate therapeutic effects in vivo Furthermore the specific complement of signalling cascades engaged by a given GPCR is determined by several factors including the particular ligand as well as the cellular and tissue context Linking specific intracellular signalling events in defined cell populations to biologic effects in the whole animal would advance our understanding of GPCR function in disease states and facilitate the development of novel functionally selective ligands (ie those that only modulate a subset of pathways downstream of a given GPCR) We have developed a method for imaging Foumlrster resonance energy transfer (FRET)-based biosensors that report signalling downstream of GPCRs in real time in live animals We combine fiber-photometry-based fluorescent recording with genetically-encoded FRET biosensors that report GPCR-mediated second messenger production (cAMP Ca2+) and protein kinase activity (PKA ERK12) with high spatial and temporal resolution Biosensors are expressed in wild-type animals using viral vectors and cell-type selective expression is achieved using specific promoters Flexible fiber-optic patch cords allow imaging to be performed in freely moving animals allowing the simultaneous measurement of behavioral and signalling responses to pharmacological manipulation

Acknowledgments This work was supported by funding from NSERC and CIHR

__________________________________________________

Presenter Sandra Paschkowsky 14h30PhD SrP30

Title RHBDL4-mediated APP processing - a clue into APP physiology or AD pathology

Authors Paschkowsky Sandra Michalski Bernadeta Fahnestock Margaret Yang Jingyun Bennett David A Munter Lisa-Marie Abstract Introduction Although known for over a century and despite extensive research Alzheimer disease (AD) is to date arguably one of the most debilitating neurodegenerative disease with no disease modifying treatment available Accumulation of toxic amyloid-beta (Abeta) peptides is one of the hallmarks of AD pathology thus lowering Abeta levels is widely considered a therapeutic strategy Abeta is generated through sequential cleavages of the amyloid precursor protein (APP) by beta- and gamma-secretase the so called amyloidogenic processing pathway Recently we discovered that RHBDL4 a mammalian rhomboid intramembrane protease residing in the endoplasmic reticulum cleaves APP multiple times in its ectodomain This novel processing pathway circumvents amyoidogenic processing and in vitro leads to a reduction of Abeta peptide levels (Paschkowsky et al JBC 2016) We therefore aimed to study the impact of RHBDL4-mediated APP processing in AD by further performing more detailed biochemical analysis of novel APP-fragments and biochemical analysis of human brain tissue Results In cell culture RHBDL4 activity decreased levels of classical Abeta peptides but may generate longer Abeta-like species instead that become secreted These Abeta-like peptides seem to be generated independently of beta- and gammashysecretase activity potentially hinting towards a membrane cleavage of RHBDL4 Analysis of RHBDL4 expression in human brain tissue revealed that RHBDL4 mRNA expression associates significantly with AD (n=450) On protein level we found a tendency of increased RHBDL4 in AD versus age-matched controls (n=10) Conclusion We propose that the upregulation of RHBDL4 in AD may be a compensatory response to the stressors occurring in AD RHBDL4 may serve as a switch to shift APP processing away from amyloidogenic processing towards a pathway secreting longer Abeta-like species

Acknowledgments This work was supported by CFI-LOF NSERC Pfizer-FRQS Scottish Rite foundation and the Alzheimer Society of Canada

__________________________________________________

Presenter Heather Fice 14h30PhD JrP32

Title Developing a Germ Cell Fast Halo Assay

Authors Fice H Robaire B

Abstract Various DNA damage tests have been developed in recent years but most are laborious expensive and require specific expertise The Fast Halo Assay (FHA) was designed to fill the niche for a less expensive quick method to examine the extent of DNA damage within a cell This test has been used with somatic but not germ cells Therefore we chose to adapt the FHA for use in germ cell DNA integrity tests Here the FHA has been modified and optimized to be compatible with germ cells including pachytene spermatocytes and round spermatids Two cell lines HepG2 (from liver) and C18-4 (spermatogonia) were used for initial optimization The deviations from the original FHA protocol include the slide type used the low melting point agarose conditions the cell concentration used an added final fixing step with 70 ethanol and a 48-hour period for the cells to settle before imaging A major development of the assay was to image the cells using the Operetta High Content Screening microscope This allows for more halos to be imaged and a greater representation of the extent of DNA damage for a given sample Columbus analysis software has been used to create an analysis script through which the halos are detected and the nuclear diffusion factor is measured This is the first instance of the halo assay being used for detection of DNA damage in cells going through different phases of spermatogenesis This method also allows for DNA dispersion and the binding of specific antibodies or probes to regions of interest within the chromatin such as the telomeres

Acknowledgments Supported by a grant from CIHR

__________________________________________________

Presenter Issan Zhang 14h30PhD SrP34

Title SAHAquines regulate metalloproteinases in human glioblastoma

Authors Zhang I Moquin A Beus M Zorc B Maysinger D

Abstract Background Glioblastoma multiforme is the most common and deadly type of brain tumors Its metastatic potential relies on high levels of matrix metalloproteinases (MMPs) that degrade the extracellular matrix and basement membranes to promote cancer invasion Although the role of MMPs in cancer is well established little is known about the effect of chemotherapeutic drugs on their expression and activity SAHAquines novel structural analogs of SAHA and primaquine are lipophilic weak acids found in lysosomes Lysosomal exocytosis plays a role in cancer drug resistance Data from our pilot studies suggest that one of the SAHAquines reduces the viability of A1235 glioblastoma but its effect on lysosomes and MMPs has not been explored Hypothesis SAHAquines inhibit glioblastoma migration by reducing MMP expression and activity To test this hypothesis we prepared and characterized a novel fluorescence resonance energy transfer (FRET)-based nanosensor and tested it in parallel with commonly used gelatin zymography in human glioblastoma cells Results We show that SAHAquines reduce human glioblastoma cell viability in the low micromolar range which is significantly more effective than temozolomide the current first-line chemotherapy for glioblastoma Life-time measurements and FRET analysis revealed high MMP-9 activity in glioblastoma Gelatin zymography data did not show significant reduction of MMP-2 and MMP-9 expression in response to SAHAquines Conclusion SAHAquines are potent anti-cancer agents that significantly decrease human glioblastoma viability Mechanisms of SAHAquine action eg MMP regulation lysosomal relocation and enzymatic activity are being explored

Acknowledgments The authors wish to thank NSERC CIHR and the JP Collip Fellowship (IZ) for their support

Presenter Helene Hall 14h30PostdocP36

Title Overexpression of human tau in the locus coeruleus of rats a tool to study the early Alzheimerrsquos Disease pathology

Authors Hall Heacutelegravene Breuillaud Lionel Flores Aguilar Lisi Cuello AClaudio Abstract Aims The locus coeruleus (LC) is located in the brainstem and is the main source of noradrenaline (NA) innervating the neocortex and hippocampus In Alzheimerrsquos Disease (AD) abnormal intraneuronal accumulation of tau protein starts in the LC decades before onset of clinical symptoms In addition NA neurons in the LC degenerate early in the course of AD leading to reduced cortical NA levels Interestingly NA plays an important role in the modulation of neuroinflammation a process that appears early on in AD and is disease-aggravating Taken together this would suggest that the LC is a key player in the early AD pathogenesis In order to further elucidate the consequences of an early LC demise on the evolution of the AD pathology we generated rats overexpressing human tau in the LC using somatic transgenesis Methods Recombinant adeno-associated viral vectors encoding for either human wild-type tau P301S mutant tau or GFP were stereotaxically injected in the LC of 3 month-old rats 12 months post injection rats were perfused and their brains collected for analyses Results Histological analyses revealed stable protein expression in the LC persisting 12 months after injection Transgene was expressed throughout the LC where NA neurons were selectively transduced Widespread GFP immunoreactivity was seen in fiber terminals of target regions including cerebral cortex and hippocampus ascending from LC neurons through the dorsal noradrenergic bundle Tau hyperphosphorylation and conformational changes as seen in human AD and tauopathies were detected in the LC Ongoing analyses of inflammatory markers will determine whether tau pathology induced in the LC can trigger a neuroinflammatory process in the naiumlve brain Conclusions This model constitutes a valuable tool to study the consequences of tau overexpression in the LC Applied to an AD-like model of amyloid pathology it should shed light on the impact of an early LC dysfunction on AD pathology

Acknowledgments The authors wish to acknowledge the following agencies for financial support CIHR FRQS the Richard and Edith Strauss Canada Foundation FQRNT and CONACyT

__________________________________________________

Presenter Daniel Sapozhnikov 14hPhD JrP38

Title Developing CRISPRCas9-based tools for the targeted manipulation of DNA methylation

Authors Sapozhnikov Daniel M amp Szyf Moshe

Abstract It has been known for decades that within promoters cytosine methylation is associated with transcriptional repression DNA methylation is a crucial epigenetic mark and deviations from the tightly-regulated developmental patterns have been reported in nearly every disease state across every tissue Despite a large body of DNA methylation research it has yet to be shown that methylation of a promoter is necessary or sufficient for silencing of the downstream gene Though there is a myriad of correlational evidence that demonstrates the inverse relationship between promoter methylation and gene expression the issue of whether DNA methylation or silencing occurs first continues to be a controversial question in the field aggravated by multiple studies suggesting that silencing can in some cases precede DNA methylation Often used to demonstrate causality genetic and pharmacological hypo- or hyper-methylating agents cause genome-wide changes in methylation confounding conclusions concerning individual promoters by countless concurrent genome-wide changes Therefore we sought to develop a tool that could cause demethylation of targeted CpGs without modulating any additional confounding transcriptional pathways We show that the nuclease-deficient Cas9 (dCas9) can be used to prevent methylation of specific CpGs in vitro We demonstrated that specific CpGs are important for the inhibition of transcription in a luciferase-based promoter-reporter assay Furthermore we determined the genetic distance with which dCas9 prevents methylation to guide future experimental design We then applied the same approach in mammalian cells to cause specific demethylation of targeted CpGs We show that we are able to completely demetylate specific CpGs We hope to use these tools to provide the first demonstration that DNA methylation of specific CpGs is necessary for endogenous downstream gene silencing

Acknowledgments We would like to thank Mr Julien LeConte for performing flow cytometry services Genome Quebec and McGill University for performing Sanger Sequencing services and undergraduate students Tatsuya Corlett and Claire Lin for general experiment assistance

Table of Contents

PRD Student Committee Welcome

Keynote Speaker

Departmental and PRD Prizes

PRD Judges

Program

Abstracts ndash Oral Presentations

Poster Session I-Odd Numbers

Poster Session II-Even Numbers

Keynote Speaker

Dr Rama Khokha is a Professor in the Department of Medical Biophysics and the Interim Research Director of Princess Margaret Cancer Centre She received her PhD from University of Western Ontario and her postdoc training at Cancer Research Labs

Melville Prizes

Best Oral Presentation Prize

PRD Life Sciences Prize

(Anatomy and Cell Biology Biochemistry IPN)

Abstract Mitochondrial processing peptidase (MPP) is a metallopeptidase that cleaves N-terminal targeting signals from the majority of nuclear-encoded mitochondrial proteins Mutations in both MPP and its substrates have been implicated in neurodege

__________________________________________________

Abstract Ligand- and voltage-gated ion channels assemble as signaling complexes comprised of both pore-forming and auxiliary subunits In the mammalian brain AMPA-type ionotropic glutamate receptors (AMPARs) co-assemble with several families of aux

__________________________________________________

Abstract Mutations in the kinase PINK1 cause autosomal recessive early-onset Parkinsonrsquos disease (PD) PINK1 selectively accumulates on damaged mitochondria and recruits Parkin by phosphorylating its Ubl domain as well as nearby ubiquitin ultimately

__________________________________________________

Abstract In chronic kidney disease (CKD) the kidneys are unable to regulate serum mineral concentrations The ensuing rise in serum calcium leads to the progressive calcification of soft tissue(s) and vasculature CKD-dependent vascular calcificatio

The calcium in mineralized tissues can bind inositol phosphates a class of phosphorylated poylols Grases et al demonstrated that inositol hexakisphosphate (IP6) inhibits cardiovascular calcification in rats As well preliminary experiments with a

__________________________________________________

Abstract Follicle-stimulating hormone (FSH) is an essential regulator of ovarian follicle development and fertility in females FSH synthesis is stimulated by intra-pituitary activins and is suppressed by gonadal inhibins Activins stimulate transcri

__________________________________________________

Abstract The voltage-gated sodium channel Nav15 is responsible for the rapid upstroke in the cardiac action potential (AP) Consistent with this mutations in the Nav15 gene have been linked to various forms of cardiomyopathy Despite the emphasis

__________________________________________________

Abstract Histone modifications are crucial in regulating many cellular processes including RNA polymerase II gene transcription and DNA repair Therefore a dysregulation of these modifications can lead to cancer pathogenesis promoting interest for

Abstract It has been hypothesized that an imbalance between the production and clearance of amyloid-β (Aβ) is a major event to initiate the pathogenesis of Alzheimerrsquos disease (AD) Thus altered Aβ cleavage profiles could provide first signs of bioc

Overall this study indicates that combining established biomarkers with C-terminally truncated Aβ cleavage products such as Aβ34 may provide improved accuracy to diagnose pre-clinical and prodromal Alzheimerrsquos disease

Abstract Mutations in PINK1 cause autosomal recessive Parkinsonrsquos disease (PD) a neurodegenerative movement disorder PINK1 is a kinase that acts as a sensor of mitochondrial damage and initiates Parkin-mediated clearance of the damaged organelle P

__________________________________________________

Abstract Fragile X syndrome (FXS) is a neurodevelopmental disorder linked to deficits in several neurotransmitter systems Dysfunction in both glutamatergic and GABAergic signaling are thought to play key roles in the manifestation of the disease ye

__________________________________________________

Abstract In Alzheimerrsquos disease (AD) excessive amyloid-β (Aβ) deposition is one of the earliest biomarker to appear prior to the emergence of overt clinical symptoms Aβ peptides are generated from amyloidogenic processing of amyloid precursor prote

__________________________________________________

Abstract The action potential (AP) is a fundamental signaling unit used by neurons to communicate within networks The AP is generated through the interplay of several voltage-gated ion channel (VGIC) families including Na+ and K+ channels which de

__________________________________________________

Abstract Thyroid hormones (THs) play fundamental roles in development and adulthood Deficiency in TH production (hypothyroidism) is common affecting approximately 2 of the Canadian population Most often the disorder derives from problems at the

__________________________________________________

Abstract Heterotrimeric G proteins comprised of α β and γ subunits are responsible for relaying signals from G protein-coupled receptors (GPCRs) to downstream effectors such as enzymes and ion channels In recent years the obligate Gβγ dimers ha

__________________________________________________

Abstract Parasitic nematodes cause various debilitating infections in humans creating health and economic burdens in developing countries Current drug treatments have limited efficacy and can cause severe adverse reactions Therefore new potential

Abstract Aim Retinopathy of prematurity (ROP) is characterized by pathological neovascularization (NV) protruding into the vitreous body threatening retinal detachment Important risk factors of ROP include prematurity and exposure to excess oxyge

Methods amp Results Wild-type (WT) C57BL6 pups are exposed to high oxygen concentrations (FiO2 75) from the post-natal day (P) 7 to 12 and subsequently returned to room air until retinal collection oxygen induced retinopathy (OIR) Our pilot data s

Conclusion Our preliminary findings indicate Hmox1 expression in glial cells under hypoxic stress may contribute to pathological NV Glial Hmox1 may therefore have a critical role in the development of diseased vessels

__________________________________________________

Abstract Alzheimer disease (AD) is a debilitating proteinopathy lacking effective diagnostics prevention and treatment Clinicians must currently rely on the presentation of late-stage events such as cognitive decline whereas the underlying pathop

__________________________________________________

Abstract Hypertension and its associated risks can be attributed to the endogenous hormone angiotensin II (AngII) AngII regulates blood volume and vascular resistance through the angiotensin II type 1 receptor (AT1R) Mutations in the AGTR1 gene tha

__________________________________________________

Abstract Flame retardants (FR) are added to a plethora of consumer products to slow fire propagation Growing evidence that ldquolegacyrdquo polybrominated diphenyl ether (PBDE) flame retardants are persistent bioaccumulative and toxic has led to a phase o

__________________________________________________

Abstract Histone modifications play a significant role in determining whether genes are actively expressed or silenced One silencing modification methylation of histone 3 on lysine 9 (H3K9me) is associated with repression of underlying genes and i

A genetic screen completed at the start of this project indicated that the transcription elongation factor Spt5 is required to prevent an accumulation of H3K9me outside of its normal compartment through a process known as heterochromatin spreading Wh

__________________________________________________

Abstract Zinc is required for numerous enzymes and DNA binding proteins involved in oxidative stress defense and DNA damage repair A previous study in our lab suggests that cyclophosphamide (CPA) an alkylating agent and known germ cell toxicant al

__________________________________________________

Abstract Glial cell line-Derived Neurotrophic Factor (GDNF) binds to the GFRa1 receptor and the GDNFndashGFRa1 complex binds to and activates the transmembrane RET tyrosine kinase to transduce signals through AktErkSrc pathways To dissect the GDNFndashG

__________________________________________________

Abstract Objective Retinopathy of prematurity is a leading cause of visual impairment in infants Exposure of premature babies to the hyperoxic extrauterine environment leads to vascular decay known as vaso-obliteration (VO) Ischemia subsequently e

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Abstract Mammalian reproduction is dependent on follicle-stimulating hormone (FSH) and luteinizing hormone (LH) secreted by pituitary gonadotrope cells The dimeric hormones share a common α-subunit (CGA) linked to hormone-specific β-subunits (LHβ an

Abstract Injury or disease affecting the somatosensory nervous systems often results in the development of chronic neuropathic pain This can cause long-lasting pain and sensory abnormalities such as pain in response to innocuous mechanical stimulati

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Abstract Therapeutic strategies for treating CLL have significantly changed from using alkylating agents to novel therapeutics BCL2 inhibitors BH3 mimetics represent an attractive treatment for lymphoid tumors as BCL2 proteins are a major promote

The aim of this study is to investigate the effectiveness of dual BCL2 and TPL2 inhibition on CLL cells As we hypothesize that the association of BH3 mimetics such (ABT199 and ABT263) and the TPL2 inhibitors (C1 and C34) could be a more efficacious

MTT (cell viability) assay was performed on OCI-ly2 cell line MTT data from this study showed that targeting both pathways simultaneously with ABT263 and C1 or C34 has synergistically enhanced tumour apoptosis in vitro with no toxic effects on norma

This study provides insight into innovative strategy and suggests that combination between BCL2 and TPL2 inhibitors acts synergistically so it might help to improve the treatment of CLL in the future

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Abstract In several fields of research animals often undergo surgeries as part of manipulations to answer certain scientific questions As such research institutions have established committees that set mandates regarding the administration of anal

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Abstract The hypothalamic-pituitary-thyroid axis regulates growth development and metabolism In this axis hypothalamic thyrotropin-releasing hormone (TRH) stimulates the secretion of pituitary thyroid-stimulating hormone (TSH) TSH stimulates thy

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Abstract Osteoarthritis (OA) is a complex disease of the whole joint and affects over 3 million Canadian adults Typically OA patients have mechanical hypersensitivity and cold allodynia most often in the knee ankle hip or shoulder joints To thi

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Abstract Pain upon movement is a main complaint of arthritis patients and in inflammatory arthritis inflammation and pain do not always correlate suggesting that a mechanism other than overt peripheral inflammation is driving pain There is an incr

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Abstract Heterotrimeric G proteins act as intermediates in G protein-coupled receptor signaling relaying extracellular signals received by cell surface receptors to downstream signaling pathways Our lab has shown that Gβγ subunits act as direct reg

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Abstract Neurotrophins play a key role in the growth and function of neurons They activate Trk and p75 receptors where Trk are considered to be neuroprotective and p75 activation leads to pro-apoptotic signaling a process initiated by p75 cleavage

I used cells expressing p75 TrkC-FL or TrkCT1 alone as well as p75 co-expressed with each TrkC isoform Co-IP showed interaction between each isoform and p75 The cells were treated with combinations of a p75 antagonist the neurotrophin NT-3 and

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Abstract Regulators of transcriptional control are crucially involved in the progression of cancer For instance mixed lineage leukemia (MLL) has been shown to be caused by fusion proteins which induce abnormal transcriptional elongation Following

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Abstract Gold nanoparticles have attracted considerable interest as delivery vehicles and imaging agents due to their surface tunability and optical properties As therapeutic agents gold nanoparticles are used for the photothermal ablation of gliom

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Abstract Fluorescent probes are widely to investigate biological processes A major limitation in their use however is the delivery of the probe to the location of interest and maintaining the probe in that location long enough to be able to detect

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Abstract As highly dynamic proteins G protein-coupled receptors (GPCRs) attain a plethora of different conformational states with certain conformations linked to specific signal transduction pathways The connection between receptor conformation an

Abstract Preterm birth (before 37 weeks gestation) affects up to 18 of all pregnancies and over 40 of cases are attributed to inflammation It is associated with many sequelae including retinopathy of prematurity (ROP) a major cause of blindnes

To this end our lab has developed rytvela a heptapeptide and an allosteric modulator of the IL-1 receptor rytvela has been previously shown to prevent preterm labour and confer protection against retinal vaso-obliteration a hallmark sign of r

These results offer early insights into structure-activity relationships of rytvela and will complement future structural and signalling transduction studies that are required prior to submitting our derivatives for clinical testing

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Abstract G protein-coupled receptors (GPCRs) are the largest family of cell surface receptors and the target of 40 of therapeutic drugs Upon agonist stimulation GPCRs can signal via G-protein coupling or via a β-arrestin-dependent mechanism β-arr

Using BRET biosensors we show that Traf 21 does not affect β-arrestin recruitment to receptors nor the formation of the β-arrestin2AP-2 complex in clathrin-coated pits By western blot we show that Traf 21 blocks receptor-mediated RasERK and PI3K

Investigating how Traf 21 pharmacologically modulates responses downstream of GPCRs may lead to a better understanding of how receptor internalization and intracellular signaling are mediated and may address their physiological role in pathologies su

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Abstract Within the last four decades an obesity epidemic has spread worldwide It is well established that genetics and lifestyle play an important role in the development of obesity Nonetheless epidemiological studies and diet-induced obesity ro

We hypothesized that histone methylation in sperm is one such mechanism implicated in the epigenetic inheritance of obesity and associated metabolic disorders To test this hypothesis we used our existing transgenic mouse model with an altered sperm

Transgenic non-transgenic littermates and C57BL6NCrl control males (n=10 per group) were fed either a low- or high-fat diet (10 or 60 kcal fat respectively) for 12 weeks starting at weaning Males were then bred to chow-fed C57BL6NCrl females Al

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Abstract The earliest stages of Alzheimers disease (AD) occur decades before clinical symptoms are present and remain elusive with no cures nor effective preventative strategies for AD currently available To unravel these early molecular events d

We hypothesize that the early iAβ accumulation triggers a pathological cascade leading to increased ROS production and subsequent mitochondrial dysfunction which then culminates in the well known AD pathology The main objectives of this project are t

Abstract G protein-coupled receptors (GPCRs) mediate neuronal responses to neurotransmitters and neuromodulators and are major drug targets in neuropsychiatric disease Individual GPCRs signal via multiple downstream effectors only some of which ma

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Abstract Introduction Although known for over a century and despite extensive research Alzheimer disease (AD) is to date arguably one of the most debilitating neurodegenerative disease with no disease modifying treatment available Accumulation of t

Results In cell culture RHBDL4 activity decreased levels of classical Abeta peptides but may generate longer Abeta-like species instead that become secreted These Abeta-like peptides seem to be generated independently of beta- and gamma-secretase

Conclusion We propose that the upregulation of RHBDL4 in AD may be a compensatory response to the stressors occurring in AD RHBDL4 may serve as a switch to shift APP processing away from amyloidogenic processing towards a pathway secreting longer Ab

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Abstract Various DNA damage tests have been developed in recent years but most are laborious expensive and require specific expertise The Fast Halo Assay (FHA) was designed to fill the niche for a less expensive quick method to examine the exte

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Abstract Background Glioblastoma multiforme is the most common and deadly type of brain tumors Its metastatic potential relies on high levels of matrix metalloproteinases (MMPs) that degrade the extracellular matrix and basement membranes to promot

Hypothesis SAHAquines inhibit glioblastoma migration by reducing MMP expression and activity

To test this hypothesis we prepared and characterized a novel fluorescence resonance energy transfer (FRET)-based nanosensor and tested it in parallel with commonly used gelatin zymography in human glioblastoma cells

Results We show that SAHAquines reduce human glioblastoma cell viability in the low micromolar range which is significantly more effective than temozolomide the current first-line chemotherapy for glioblastoma Life-time measurements and FRET analy

Conclusion SAHAquines are potent anti-cancer agents that significantly decrease human glioblastoma viability Mechanisms of SAHAquine action eg MMP regulation lysosomal relocation and enzymatic activity are being explored

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Abstract Aims The locus coeruleus (LC) is located in the brainstem and is the main source of noradrenaline (NA) innervating the neocortex and hippocampus In Alzheimerrsquos Disease (AD) abnormal intraneuronal accumulation of tau protein starts in the

Methods Recombinant adeno-associated viral vectors encoding for either human wild-type tau P301S mutant tau or GFP were stereotaxically injected in the LC of 3 month-old rats 12 months post injection rats were perfused and their brains collected

Results Histological analyses revealed stable protein expression in the LC persisting 12 months after injection Transgene was expressed throughout the LC where NA neurons were selectively transduced Widespread GFP immunoreactivity was seen in fib

Conclusions This model constitutes a valuable tool to study the consequences of tau overexpression in the LC Applied to an AD-like model of amyloid pathology it should shed light on the impact of an early LC dysfunction on AD pathology

Abstract It has been known for decades that within promoters cytosine methylation is associated with transcriptional repression DNA methylation is a crucial epigenetic mark and deviations from the tightly-regulated developmental patterns have been

We show that the nuclease-deficient Cas9 (dCas9) can be used to prevent methylation of specific CpGs in vitro We demonstrated that specific CpGs are important for the inhibition of transcription in a luciferase-based promoter-reporter assay Furtherm

We hope to use these tools to provide the first demonstration that DNA methylation of specific CpGs is necessary for endogenous downstream gene silencing

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Special Thanks To All Of Our Sponsors

Innovative Medicine

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Faculty of Medicine McGill University

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Megs Specialty Gases Inc

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Wisent Bioproducts

Laser Quest

N sur MacKay

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Friday, October 13th 2017 - McGill University › pharma › files › pharma › ... · Senior Scientist at Princess Margaret Cancer Centre since 1996. Her lab studies tumour microenvironment