21st Annual McGill Biomedical Graduate Conference (AMBGC) | May 11-12, 2021
Experimental Medicine Graduate Students' Society (EMGSS)
Published online: 10 May 2021Finite-element modelling of newborn middle-ear vibrations under quasi-static pressurization
Royan Jafari1, Robert Funnell1,2,31Department of Biological and Biomedical Engineering, McGill University, Montréal, QC, Canada
2Department of Biomedical Engineering, McGill University, Montréal, QC, Canada
3Department of Otolaryngology-Head & Neck Surgery, McGill University, Montréal, QC, Canada
Corresponding Author: Royan Jafari, email royan.jafari@mail.mcgill.ca
Abstract
Unidentified hearing loss in newborns affects speech and language development, academic achievement and social-emotional development. Unfortunately, currently available screening tools for hearing produce large numbers of false positives, largely because of middle-ear conditions. Tympanometry is a promising clinical tool for evaluating the condition of the middle ear by using sounds and quasi-static pressure together as inputs. However, tympanometry is poorly understood in newborns.
To simulate the effects of the large static pressures of tympanometry, a non-linear static model of a 22-day-old newborn ear canal and middle ear was developed for the first time in our lab, without taking into account the dynamic behaviour of the ear. Then a linear version of the 22-day-old model was developed to simulate a broad range of frequencies but without the large static pressures. Most recently, a 1-day-old model was developed. It was a linear model and its responses to static and low-frequency inputs (without inertial and damping effects) were simulated.
Our overall objective is to use mathematical modelling to improve our understanding of how the newborn middle ear responds to different stimuli. The specific objective of this study is finite-element modelling of the 1-day-old newborn middle-ear under high quasi-static pressures and low-amplitude sound pressures, as in tympanometry.
We use FEBio version 2.9 (https://febio.org/), an open-source biomechanical finite-element solver. Locally developed software including Fie, Tr3 and Fad (http://audilab.bme.mcgill.ca/sw/) and open-source software called Gmsh is used for the purpose of 3-D construction and refining of the model. The methods for modelling non-linear viscoelasticity are based on the previous modelling of the gerbil middle ear in our lab.
The static or low-frequency simulation results for the 1-day-old model are obtained in terms of spatial vibration patterns and maximum displacement values. The maximum displacement is located in the anterior region of the eardrum. The model will be verified against our previous models and validated against data obtained from the literature.
This nonlinear viscoelastic mathematical model will help us to gain insight into the response of the middle ear to different loading conditions and allow us to better interpret tympanometry measurements and improve newborn hearing screening.
P2X7 receptor knockout attenuates angiotensin II-induced hypertension, vascular injury and CD8+ T cell infiltration
Brandon Shokoples1, Pierre Paradis1, Ernesto L. Schiffrin1,21Hypertension and Vascular Research Unit, Lady Davis Institute for Medical Research, Montréal, QC, Canada
2Department of Medicine, Sir Mortimer B. Davis-Jewish General Hospital, McGill University, Montréal, QC, Canada
Corresponding Author: Brandon Shokoples, email brandon.shokoples@mail.mcgill.ca
Abstract
Background: Elevated plasma adenosine triphosphate (ATP) levels have been observed in hypertensive patients, which could engage the immune system through activation of the purinergic receptor P2X7 (P2RX7). Pharmaceutical antagonism of P2RX7 or P2rx7 gene knockout reduced blood pressure (BP) in high salt diet- and deoxycorticosterone acetate salt-induced models of hypertension. However, whether P2RX7 activation contributes to angiotensin (Ang) II-induced BP elevation and vascular damage through enhanced immune activation is currently unknown. We hypothesized that deletion of P2rx7 would blunt Ang II-induced BP elevation, vascular injury and infiltration of immune T cells into perivascular adipose tissue (PVAT).
Methods: Ten to 12-week-old male C57BL/6J male wild-type (WT) and P2rx7–/– mice underwent sham surgery or were infused with Ang II (1000ng/kg/min) for 14 days. BP was determined by telemetry, mesenteric artery function using pressurized myography, aortic stiffening by ultrasound and infiltration of activated immune T cells in PVAT by flow cytometry. IL-1ß secretion from WT and P2rx7–/– bone marrow-derived macrophages (BMDM) and dendritic cells (BMDC) was assessed by ELISA.
Methodology: Lipopolysaccharide plus ATP stimulated IL-1ß release from WT BMDMs and BMDCs, but not from P2rx7–/– cells. Ang II-infusion induced hypertension in WT mice, increased vascular stiffness, induced endothelial dysfunction, and resulted in increased infiltration of activated CD8+ T cells in aortic PVAT (P<0.05). All of the above were reduced or prevented by P2rx7 knockout (P<0.05).
Conclusion: P2rx7 knockout attenuates Ang II-induced hypertension, vascular injury and infiltration of activated CD8+ T cells into aortic PVAT.
The Invisibility of the Asian American Identity in North American Bioethics
Katherine Amber LiCorresponding Author: Katherine Amber Li, email katherine.amber.li@gmail.com
Abstract
This commentary highlights the major barriers behind Asian American invisibility in bioethics, such as the lack of a coherent demographic identity in society, the scarcity of academic literature about their narrative, a perceptual bias against Asians in bioethics which negatively affects Asian American, a noticeable absence from social activism movements and their underrepresentation in clinical trials. It also outlines some troubling moral implications of this invisibility, like perpetuating the model minority myth, ignorance of Asian American cultural values, and the conflation of race and ethnicity in medicine. This leads to a practical knowledge gap for healthcare practitioners about the Asian American demographic and their perspectives in North American bioethics.Ultimately, a new, Asian American bioethics is needed to separate their ethnic and racial differences from their Asian or American predecessors.
Role Of Vγ6+ Gd T Cells in Angiotensin II-Induced Hypertension and Vascular Injury
Ahmad Mahmoud1, Antoine Caillon1, Pierre Paradis1, and Ernesto L. Schiffrin1,21Hypertension and Vascular Research Unit, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis-Jewish General Hospital, McGill University, Montréal, QC, Canada
2Department of Medicine, Sir Mortimer B. Davis-Jewish General Hospital, McGill University, Montréal, QC, Canada
Corresponding Author: Ahmad Mahmoud, email ahmad.mahmoud2@mail.mcgill.ca
Abstract
Background: We previously demonstrated that a small subpopulation of T cells considered "innate-like", expressing the ?d T-cell receptor (TCR) plays a key role in hypertension and vascular injury. ?d T cells can be subdivided according to the TCR variant (V) subtype that is generally specific for a tissue. A subpopulation of lung and skin ?d T cells that are V?6+ and produce interleukin (IL)-17A was shown to respond promptly to pneumococcal infection and skin inflammation. However, ?d T cell V? subtypes involved in hypertension are still unknown. We hypothesized that V?6+ ?d T cells may play a role in angiotensin (Ang) II-induced hypertension.
Methodology: Eleven-to-13-week old C57BL/6J male mice were infused or not with Ang II (490 ng/kg/min, SC) for 14 days and injected IP with control isotype IgG1 or anti-TCR V?6 antibodies. Blood pressure was determined by telemetry, mesenteric artery endothelial function by pressurized myography and T-cell profiling by flow cytometry.
Methodology: The frequency of IL-17 producing effector memory (CCR6+CXCR3–CD69+CD44+) V?6+ ?d T cells was increased in spleen (1.7-fold, P<0.01) and tended to be elevated in MA perivascular adipose tissue in Ang II-infused mice compared to control mice. Anti-TCR V?6 antibody injections in Ang II-infused mice enhanced the early elevation of the systolic and diastolic BP (P<0.05), and reduced the mesenteric artery dilatory response to acetylcholine by 50% compared to the control antibody injections (P<0.05).
Conclusion: V?6+ ?d T cells play a protective role in Ang II-induced hypertension and vascular injury.
The role of Hippo signaling in stromal-epithelial interactions in acinar-to-ductal metaplasia and pancreatic cancer initiation
Julia Messina-Pacheco1, Alex Gregorieff11Department of Pathology, McGill University, Montréal, QC, Canada
Corresponding Author: Julia Messina-Pacheco, email julia.messina-pacheco@mail.mcgill.ca
Abstract
Background: Pancreatic ductal adenocarcinoma (PDAC) is the 4th leading cause of cancer-related deaths with a 5-year survival rate of approximately 7%. Current models suggest that PDAC may originate from acinar cell trans-differentiation into ductal-like cells. Acinar-to-ductal metaplasia (ADM) is triggered by chronic pancreatitis and/or mutations in K-Ras. The progression to PDAC is associated with a dense fibrotic stroma. The Hippo transcriptional effector YAP1 is a tension-stimulated cancer-associated fibroblast (CAF) activator that promotes ECM stiffening, thereby contributing to pancreatic cancer progression.
Hypothesis: We hypothesize that the Hippo pathway may coordinate fibroinflammatory signals emanating from the stromal compartment during regenerative responses to acinar cell injury in the pancreas and progression towards PDAC.
Methods and Methodology: To unveil the gene expression profile of ADM, we performed a spatial analysis of over 1800 RNA probes in normal, ADM and PDAC areas of human tumour tissue using NanoString’s GeoMx Digital Spatial Profiling (DSP) technology, thus revealing functional enrichment in multiple pathways in ADM, including K-Ras signaling, ECM organization and epithelial-to-mesenchymal transition. Further studies into the role of Yap in ADM will be conducted by immunofluorescence staining with ductal, stromal and fibroblast markers.
To study the role of Hippo signaling in stromal cells, we conditionally deleted Yap/Taz in Collagen1a2-producing cells in a Col1a2Cre-ERT;LSL-Tom murine model of caerulein-induced pancreatitis. Lineage tracing will be completed to verify the contribution of Col1a2-expressing cells to the stromal reaction.
Conclusion: Further insights into the mutational and expression gene profile of ADM and its regulatory network may allow for the discovery of early diagnostic biomarkers and strategies to prevent pancreatic cancer development.
Investigation of the sexual dimorphic tumor suppressor role of DDX3X in melanoma
M. Lingrand1, R. Alkallas1, M. Lajoie1, I. R. Watson1
1Goodman Cancer Research Centre, Department of Biochemistry, McGill University, Montréal, QC, Canada
Corresponding Author: Marine Lingrand, email marine.lingrand@mail.mcgill.ca
Abstract
Patient sex remains a poorly understood prognostic factor in melanoma. At all melanoma stages, men have higher incidence rates and poorer prognosis (Joosse, A. et al., J Clin Oncol 2012 ; Joosse, A. et al., J Clin Oncol 2013). Recently, our group performed a mutational meta-analysis of cutaneous melanoma combining 1,014 exomes from five studies, which identified new significantly mutated genes (Alkallas, Lajoie et al., Nature Cancer, 2020). Interestingly, we found that loss-of-function (LoF) mutations in X-linked DEAD-box RNA helicase, DDX3X, are solely found in male patients. Additionally, we demonstrated that DDX3X can escape from X-inactivation, which would protect females from complete DDX3X loss in the case of a single mutational event. To date, the reported functions of DDX3X include RNA metabolism, regulation of translation and mediators of important cancer signaling pathways. However, the role of DDX3X in melanoma is not entirely clear. We hypothesize that DDX3X is a sexual dimorphic tumor suppressor gene that plays a role in mediating the observed differences in incidence and outcome observed between female and male melanoma patients. To address this, we generated stable CRISPR/Cas9-mediated DDX3X knock-out (KO) in male human melanocyte and melanoma lines. Moreover, we identified DDX3X-null human lines to carryout DDX3X gain of function studies.In these models, we determined DDX3X plays a role in proliferation, migration and invasion in melanoma providing supports for its tumor suppressor role in this malignancy.
Non-canonical signaling of the meiosis gene HORMAD1 and its role in DNA damage repair in cutaneous squamous cell carcinoma
Jennifer Gantchev1, Amelia Martinez Villarreal1, Brandon Ramchatesingh1, and Ivan V. Litvinov11Research Institute of the McGill University Health Centre, Montréal, QC, Canada
Corresponding Author: Jennifer Gantchev, email jennifer.theoret@mail.mcgill.ca
Abstract
The accumulation of genomic instability through successive DNA strand breaks and mutations is a dynamic mechanism that has been shown to promote carcinogenesis. Recent research has demonstrated that most cancers with high levels of genomic instability also ectopically express meiosis genes, however the role of these genes remain elusive. We focus our work on revealing the expression and function of the meiosis gene, HORMAD1, in cutaneous squamous cell carcinoma (cSCC). In meiosis, HORMAD1specializes in mediating double stand break formation and maintaining genome integrity throughout homologous recombination, an important role in modulating meiosis progression. First, we demonstrated the ectopic expression of HORMAD1 in patient cSCC samples using immunohistochemistry and in patient derived cell lines using western blot analysis. We have revealed that HORMAD1 facilitates a functional level of genomic instability by decreasing the number of double strand breaks, chromatin bridges, and micronuclei, leading to increased cell survival in response to DNA damage elicited by etoposide treatment. In fact, we have gone on to show that HORMAD1 is involved in DNA repair in cSCCs. Most importantly, we have elucidated a pathway by which HORMAD1 is mediated. With the use of shRNA mediated knockdown of the meiosis-specific transcription factor STRA8, we see a subsequent decrease in HORMAD1 protein expression. Additionally, through the use of a MEK inhibitor, we have shown that both STRA8 and HORMAD1 expression is modulated by the MAPK signaling pathway. Our results indicate that STRA8 is a transcription factor that mediates HORMAD1 expression through the non-canonical MAPK pathway to elicit a DNA damage response in cSCC.
Characterization of the CdGAP-ß-PIX interaction in glomerular podocytes
Dina Greenberg1,3, Lamine Aoudjit2, Jun Matsuda2, Nathalie Lamarche-Vane3, Tomoko Takano11Division of Experimental Medicine, McGill University, Montréal, QC, Canada
2Division of Nephrology, McGill University Health Centre, Montréal, QC, Canada
3Department of Anatomy and Cell Biology, McGill University, Montréal, QC, Canada
Corresponding Author: Dina Greenberg, email dina.greenberg@mail.mcgill.ca
Abstract
The glomerulus is the filtration unit of the kidney. When its barrier function (permselectivity) is impaired, leakage of proteins in urine (proteinuria) occurs. This is a hallmark of kidney disease. Visceral glomerular epithelial cells (or “podocytes”) are critical for the permselectivity of the glomerulus. Podocytes have actin-based finger-like projections called foot processes that interdigitate at the glomerular capillary. Dysregulation of the podocyte cytoskeleton can cause foot process effacement or detachment, and consequently, proteinuria.
The Rho family of small GTPases (Rho GTPases) are cytoskeletal regulators. These are in turn regulated by GTPase activating proteins (GAPs, negative regulators) and guanine nucleotide exchange factors (GEFs, positive regulators). Rac1 and Cdc42 are two Rho GTPases; both regulate the actin cytoskeleton and cause renal disease phenotype in vivo when dysregulated. ARHGAP31 (CdGAP) and ARHFEG7 (ß-PIX) act as a GAP and GEF respectively for both Rac1 and Cdc42. Furthermore, recent data by Matsuda et al. has demonstrated the physical interaction of these two proteins in vitro. However, the functional significance of this interaction is unknown.
Here, we examine the nature of this interaction by visualizing the colocalization of CdGAP and ß-PIX. We show that the spatial intracellular relationship of these proteins is altered upon EGF stimulation. Lastly, we find that overexpression of CdGAP alone, ß-PIX alone, or CdGAP and ß-PIX together affects cell migration dynamics. Together, these data suggest that, despite its GAP activity, CdGAP may be required for the normal function of ß-PIX in the regulation of the downstream Rho GTPase targets Rac1 and Cdc42.
Characterizing The Interplay Between Angiogenic And Immunoactive Factors Of Hepatocellular Carcinoma
Audrey Kapelanski-Lamoureux1, Flemming Kondrop1, Stephanie Petrillo2, Thomas Mayer2, Anthoula Lazaris2, Peter Metrakos2,31Anatomy & Cell Biology, McGill University, Montréal, QC, Canada
2Research Institute of the McGill University Health Centre, Montréal, QC, Canada
3Department of Surgery, McGill University, Montréal, QC, Canada
Corresponding Author: Audrey Kapelanski-Lamoureux, email audrey.kapelanski-lamoureux@mail.mcgill.ca
Abstract
Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death globally. It is considered a multistep process evolving from underlying liver cirrhosis most commonly due to hepatitis (B or C) virus infection or non-alcoholic steatohepatitis (NASH). Patients typically present at an advanced stage and less than 50% reach the maximum 1-year survival rate, when given as first-line treatment, Sorafenib. This highlights the need for novel therapeutic targets as well as predictive biomarkers to increase overall survival for patients with HCC. In this era of personalized therapy, immunotherapy is revolutionizing treatment of a variety of cancers, including HCC. Given the important role of angiogenesis in HCC from its early stage and its rich immune composition, anti-angiogenic and immune checkpoint inhibitors (ICI), are two therapeutic approaches when combined have shown efficacy with response rate never observed before in patients with HCC. While combination of agents inhibiting angiogenesis and ICI have entered clinical trials for HCC, the interplay between angiogenic factors and immunity in the context of this approach remains poorly understood. Here we focus on understanding the interplay between the vascular state of the tumor and the immune response in HCC. As a first step, we focus on characterizing the immune and vasculature landscape with a primary focus on the correlation to histopathological features and HCC subtypes. This project presents preliminary evidence for the interaction of vascular factors with immune cells, thus providing insight into the biological rationale of why 30% of patients responded to combined angiogenic and immunotherapy treatment.
eIF4E phosphorylation drives the production and spatial organisation of collagen type I in the mammary gland
Samuel E J Preston1,2, Christophe Gonçalves2, Vincent Richard3, René Zahedi3,4, Christoph Borchers3, Wilson Miller1,2,4, Sonia del Rincón1,2,41Division of Experimental Medicine, McGill University, Montréal, QC, Canada.
2Department of Oncology, Segal Cancer Centre, Lady Davis Institute, Jewish General Hospital, McGill University, Montréal, QC, Canada.
3Segal Cancer Proteomics Centre, Lady Davis Institute, Jewish General Hospital, Montréal, QC, Canada.
4McGill Centre for Translational Research in Cancer (MCTRC), McGill University, Montréal, QC, Canada.
Corresponding Author: Samuel Preston, email samuel.preston@mail.mcgill.ca
Abstract
Background: The ECM is a highly dynamic component of the tumor microenvironment (TME) that adapts to reinforce neoplastic progression. The MNK1/2-eIF4E axis exemplifies how translation initiation can act abnormally to promote cancer. eIF4E phosphorylation, uniquely by MNK1/2, induces the translation of a subset of mRNA involved in invasion and metastasis, some of which are known matrisomal proteins. To date, no one has reported on how translational control impacts ECM homeostasis.
Methods: We aim to define ECM signatures that are regulated by the MNK1/2-eIF4E axis. Primary ECM samples were isolated from the mammary glands (MGs) of wild-type and phospho-eIF4E-deficient female mice and subjected to proteomic analysis. In tandem, ECM preparations were used to treat breast cancer cells and primary MG fibroblasts in vitro for immunoblot, qPCR, transwell migration-invasion and functional assays. Tail vein injection and orthotopic mammary fat pad injection models were also used, as well as IHC staining on patient-derived samples.
Methodology: Matrisomal profiling highlights several proteins that are differentially expressed between conditions. This includes collagen-I which is downregulated in phospho-eIF4E-deficient MG ECM. Phospho-eIF4E-deficient MG fibroblasts produce less collagen-I and produce a less organized 3D matrix as compared to wild-type. Breast cancer cells treated with phospho-eIF4E-deficient MG ECM have suppressed invasive capacity in vitro and in vivo, when compared to wild-type. Breast cancer patients also show a positive correlation between phospho-eIF4E and collagen-I.
Conclusion: We have shown that ECM from a phospho-eIF4E deficient MG fosters a less invasive TME, through its modulation of collagen I production and topology.
Change in Skeletal Health Determinants and Outcomes Among Canadians: Comparison of the Canadian Multicenter Osteoporosis Study and the Canadian Longitudinal Study on Aging cohorts
Hassanabadi N1, Berger C2, Papaioannou A3, Cheung AM4, Rahme E1,2, Leslie WD5, Goltzman D1,2, Morin SN1,21Department of Medicine, McGill University, Montréal, QC, Canada
2Research Institute of the McGill University Health Centre, Montréal, QC, Canada
3Department of Medicine, McMaster University, Hamilton, ON, Canada
4Department of Medicine, University of Toronto, Toronto, ON, Canada
5Department of Medicine, University of Manitoba, Winnipeg, MB, Canada
Corresponding Author: Nazila Hassanabadi, email nazila.hassanabadi@mail.mcgill.ca
Abstract
Background: It is unknown whether bone mineral density (BMD), fracture rates and the osteoporosis caregap have changed over time in Canada.
Methodology: We compared baseline data of participants 50-85 years from Canadian Multicenter Osteoporosis Study (CaMos, N=6,479; 1995-1997) and Canadian Longitudinal Study on Aging (CLSA, N=17,662; 2012-2015) and use of anti-osteoporosis therapy in those at high risk for fractures over a 20-year period. We created sex-stratified linear and logistic regression models to determine differences between cohorts in femoral neck BMD (FN-BMD) (g/cm2; 95%CI) and prevalent MOF, adjusting for age, body mass index (BMI) and other important covariates.
Methodology: Participants from CaMos were older and had lower BMI than CLSA. Adjusted linear regression models revealed higher FN-BMD in CLSA (women; 0.014; 0.010-0.018, men; 0.006; 0.000-0.011), compared to CaMos. Adjusted Odds Ratios (aOR; 95%CI) for prevalent MOF were significantly lower in CLSA than CaMos (women; 0.50; 0.43-0.59, men; 0.43; 0.33-0.55). Results were similar when restricting the analyses to participants of White ancestry. The prevalence of women at high risk for fractures who received anti-osteoporosis treatment was lower in CaMos women (supplements 32.0%, bisphosphonates 5.8%) than in CLSA (supplements 47.8%, bisphosphonates 16.4%). Anti-osteoporosis treatment was low in men in both cohorts and did not change over time.
Conclusion: Although we documented a higher BMD and a lower fracture prevalence in the CLSA cohort compared to CaMos; over a 20-year period the osteoporosis caregap remained high in Canadians at high risk for fractures, particularly in men.
Emergence of B1 integrin-deficient mammary tumors from dormancy involves both epithelial cell intrinsic and extrinsic mechanisms
Yu Gu1,2, Tung Bui1,2, Frederic Ancot1, Virginie Sanguin-Gendreau1, Dongmei Zuo1, and William J. Muller1,2,31Rosalind and Morris Goodman Cancer Research Centre, McGill University, Montréal, QC, Canada
2Department of Biochemistry, McGill University, Montréal, QC, Canada
3Faculty of Medicine, McGill University, Montréal, QC, Canada
Corresponding Author: Yu Gu, email yu.gu3@mail.mcgill.ca
Abstract
The molecular and cellular mechanisms behind mammary tumour dormancy are unclear and how these processes are dynamically orchestrated to allow for tumor resurgence remains to be elucidated. In concordance with our previous studies, we report that mammary epithelial specific disruption of ß1 integrin in a murine model of Luminal B human breast cancer drastically impairs tumour growth. This phenotype is accompanied with proliferation block, apoptosis induction and cellular senescence, altogether resulting in tumour mass dormancy. Molecular analyses from the sequencing of ß1 integrin-deficient dormant lesions show activation of p53 and Retinoblastoma (Rb) tumour suppressors, and tumours that eventually escape dormancy possess mutations in these pathways analogous to those in human disease. We further demonstrate that mammary epithelial deletion of p53 alone and in ß1 integrin-deficient mice fully rescues dormant tumour phenotype, yet deletion of Rb only displays limited effect on tumour dormancy. Hence, we show that ß1 integrin modulates key tumour suppressor pathways that exert differential effects on tumorigenesis and cancer dormancy. In addition to cell autonomous networks, single cell RNA-sequencing of resurgent ß1 integrin-deficient dormant tumours unveils extensive stromal activation, involving recruitment and activation of cancer-associated fibroblasts and increased tissue fibrosis. We identify a potential druggable target, Osteopontin, as a cytokine mediator of this paracrine signaling that permit dormancy exit. Collectively, these results reveal both novel cell intrinsic and extrinsic factors regulated by ß1 integrin that govern mammary tumour dormancy which henceforth should be considered in future therapeutics to avert cancer recurrence caused by tumour dormancy.
Mechanisms of early life RSV infection-mediated enhancement of Type 2 allergic lung inflammation
Lydia Labrie1,2, Haya Aldossary1,2, Vanessa Moarbes1,3, Veronique Gaudreault1, Jichuan Shan1, Brian Ward1,2,3, Elizabeth Fixman1,31Research Institute of McGill University Health Centre (RI-MUHC), Montréal, QC, Canada
2Department of Microbiology and Immunology, McGill University, Montréal, QC, Canada
3Department of Experimental Medicine, McGill University, Montréal, QC, Canada
Corresponding Author: Lydia Labrie, email lydia.labrie@mail.mcgill.ca
Abstract
Respiratory syncytial virus (RSV) infection leads to millions of hospitalizations and many thousands of deaths per year. Early childhood RSV infection is also associated with development of wheezing and asthma. Currently, neither antivirals nor vaccines are available for RSV; thus, novel preventative and therapeutic strategies are urgently needed. A potential preventative strategy is a cell-penetrating peptide we developed called STAT6-IP, which inhibits Type 2 inflammation in allergy and RSV infection models. To understand the potential impact of STAT6-IP in the induction of asthma associated with respiratory virus infection, we have developed a clinically relevant murine model to examine the impact of early life RSV infection on house dust mite- (HDM-) induced Type 2 inflammatory responses in the lung. My data demonstrate that prior exposure to RSV markedly enhanced HDM-induced lung eosinophil (Eos) numbers and activation (by flow cytometry) as well as production of Type 2 cytokines by ex vivo-stimulated lung draining mediastinal lymph node (MLN) cultures (by ELISA). RSV infection enhanced responses in mice exposed acutely to HDM to induce Th2 adaptive immunity. The degree of Eos activation was more prominent in females. Delivery of STAT6-IP selectively at the time of RSV infection reduced both lung and MLN inflammatory responses induced by HDM. These data suggest that STAT6-IP has the potential to provide protection against the development of allergic airways disease associated with early life RSV infection. Further investigation is required to elucidate the mechanisms of action of STAT6-IP as well as the mechanisms underlying sex differences in this model.
MedSafer to Support Deprescribing for Residents of Long-Term Care: A Mixed-Methods Study
Giulia-Anna Perri MD1*, Émilie Bortolussi-Courval CPN2*, Christopher D. Brinton BSc1, Anna Berall RN1, Anna Theresa Santiago MPH MSc1, Mareiz Morcos RPh, PharmD, PMP4, Todd C. Lee MD MPH2,3, Emily G. McDonald MD MSc2,3*Equal contributions for the purposes of authorship
1Baycrest, Toronto, Ontario, Canada
2Division of Experimental Medicine, McGill University, Montréal, QC, Canada
3Clinical Practice Assessment Unit, McGill University Health Centre, Montréal, QC, Canada
4Clinical Pharmacist, Edmonton, Alberta, Canada
Corresponding Author: Émilie Bortolussi-Courval, email emilie.bortolussi-courval@mail.mcgill.ca
Abstract
Objectives: This study evaluated the feasibility and applicability of physicians using an electronic deprescribing tool facilitating deprescribing in a long-term care home (LTCH) during the quarterly medication review (QMR).
Design: Semi-structured interviews obtained physicians’ feedback on the usefulness and applicability of MedSafer in the LTCH setting. Retrospective chart reviews collected medication changes during the MedSafer QMR and during the previous standard QMR for comparison. A mixed-method approach evaluated the pilot implementation of MedSafer during the QMR for two units in an academic LTCH in Ontario, Canada.
Measures: Semi-structured interviews obtained physicians’ feedback on MedSafer-LTCH in the LTCH setting. Resident data collected included demographics, comorbidities, medications, and recent lab values. Residents' prescriptions were collected to compare changes made at the MedSafer-LTCH QMR for each pilot unit to the standard QMR.
Results: Physicians found the software helpful and of potential use in LTCHs and provided suggestions for improving MedSafer’s use in LTCHs. The standard QMR deprescribed an average of 0.5 (SD = 0.9) medications per resident versus the MedSafer-LTCH QMR average of 1.1 (SD = 1.3, p < 0.05 for comparison). Standard QMR and MedSafer-LTCH QMRdeprescribing rates differ: MedSafer-LTCH deprescribed 0.5 (SD = 1.1) more medications per resident than the standard QMR (p = 0.01) with a moderate effect size (0.5).
Conclusions and Implications: MedSafer, a scalable deprescribing intervention, has potential to augment medication management in LTCHs. Software improvements suggestions are currently being studied. Further research should investigate its impact on polypharmacy and reduction of adverse drug events in older adults.
Investigating the Receptor Mediated Effects of Select Cannabinoids on the Innate Inflammatory Response
Matthew Preteroti1,3, Dr. Carolyn Baglole1,2,3,41Research Institute of the McGill University Health Centre, Montréal, QC, Canada
2Department of Medicine, McGill University, Montréal, QC, Canada
3Department of Pathology, McGill University, Montréal, QC, Canada
4Department of Pharmacology and Therapeutics, McGill University, Montréal, QC, Canada
Corresponding Author: Matthew Preteroti, email matthew.preteroti@mail.mcgill.ca
Abstract
Derived from the plant Cannabis Sativa, exist in excess of 120 compounds called cannabinoids. Cannabinoids represent a class of terpenophenolic compounds which interact with the endogenous cannabinoid system to exert an assortment of physiological effects including those that modulate psychoactivity in addition to immune reactivity. ?9-tetrahydrocannabinol (?9-THC) and cannabidiol (CBD) represent two of the most abundant and extensively studied cannabinoids found in Cannabis sativa. The endogenous cannabinoid system consists of two primary receptors; the cannabinoid receptor 1 (CB1) and the cannabinoid receptor 2 (CB2). CB1 is predominantly expressed within nervous tissues and is known to mediate the psychoactive effects of ?9-THC. CB2 conversely, is a highly inducible receptor shown to be expressed on immune cell surfaces with potential roles in inflammatory and oxidative stress processes. Here we will be investigating the receptor-mediated properties of cannabinoids; ?9-tetrahydrocannabinol and cannabidiol on the inflammatory response of alveolar macrophages. Initially, through the use of qPCR and Multiplex Assay, we found that both ?9-THC and CBD had no effect on basal expression of both pro- and anti-inflammatory cytokines. Conversely, when in a state of LPS-induced inflammation both ?9-THC and CBD were able to significantly reduce levels of pro-inflammatory cytokines TNF-a, IL-1ß, and IL-6. Further investigation revealed that the observed anti-inflammatory effects of CBD but not ?9-THC were mediated through a reduction in signalling through the NF-kB transcription factor and ERK1/2 MAP kinase. Whether the endogenous cannabinoid receptors are mediating the observed findings is currently being investigated through knockdown of the cannabinoid receptors.
High Ki67 as a predictor for distant recurrence in early stage breast cancer with low oncotype Dx score
Parvaneh Fallah1, Nasser Mulla2, April Rose1, and Lawrence Panasci11Department of Oncology, Jewish General Hospital, Montréal, QC, Canada
2Taibah University, Medina, Saudi Arabia
Corresponding Author: Parvaneh Fallah, email parvaneh.fallah@mail.mcgill.ca
Abstract
Background: Ki-67 is a marker of proliferating cells. Oncotype Dx score based on the 21-gene provides prognostic and predictive information for recurrence in early stage breast cancer patients. In this retrospective study, we aimed to examine whether high Ki67 could predict the distant recurrence in early stage breast cancer with low Oncotype Dx (=<25).
Method: This retrospective study included 278 consecutive cases of hormone receptor-positive, HER2 negative (T1-2 N0 M0) breast cancer who were diagnosed between 2008 and 2015 with low oncotype Dx (=<25). Patients’ clinical outcome in terms of distant recurrence after breast surgery was determined up to December 2020 (median follow-up of 7 years). Patients were divided in to low risk (Ki67<15%) and high risk (Ki67>=15%) groups.
Methodology: Of 278 cases with average and median age of 59 and 60 respectively (age range 30 to 79 year-old), 274 (98%) were estrogen receptor (ER) > 90% and 4 (2%) were ER 75-85%.
29 (10%) patients out of 278 were low positive (<10%) for progesterone receptor (PR) and 249 (90%) were PR>10%. 271 (97%) received adjuvant anti-hormonal treatment for an average of 5 years (ranging from 6 months to 10 years).
148 (53%) patients were in Ki67 low risk and 130 (47%) were in Ki67 high risk group.
Of all cases in the study, 13 patients (4%) experienced distant metastasis that occurred in lung, liver, bone and skin. Of these 13 cases, 6 had oncotype Dx between 20 and 25. 12 out of 13 cases (92%) were in the Ki67 high risk group and only 1 (8%) belonged to the low risk category. Of 265 patients who did not have distant metastasis, 147(55%) belonged to low Ki67 and 118(45%) were in high Ki67 group.
High Ki67 patients were overrepresented in group with recurrent distant metastasis compare to group without recurrent disease (Pearson Chi-Square=51.18 with 1 degree of freedom and P=<0.001)
Conclusion: Ki67 high patients in the low risk oncotype Dx group are relapsing at a significantly higher rate suggesting that Ki67 combined with low oncotype Dx further refines the risk of distant relapse.
Development of a TaqMan Multiplex Assay for the Rapid Diagnosis of Filarial (helminth) Infections
Jackson Chen1,2,3,4, James Stewart3,4, Momar Ndao1,2,3,4,5,6,7,81Department of Microbiology and Immunology, McGill University, Montréal, QC, Canada
2Department of Physiology, McGill University, Montréal, QC, Canada
3National Reference Centre for Parasitology, The Research Institute of the McGill University Health Centre, Montréal, QC, Canada
4Infectious Diseases and Immunity in Global Health, The Research Institute of the McGill University Health Centre, Montréal, QC, Canada
5Department of Medicine, Division of Experimental Medicine, McGill University, Montréal, QC, Canada
6Department of Medicine, Division of Infectious Diseases, McGill University, Montréal, QC, Canada
7J.D. MacLean Centre for Tropical Diseases, McGill University Health Centre, Montréal, QC, Canada
8Institute of Parasitology, McGill University, Ste Anne de Bellevue, QC, Canada
Corresponding Author: Jackson Chen, email jackson.chen@mail.mcgill.ca
Abstract
Lymphatic filariasis (LF) and subcutaneous filariasis (SF) are diseases caused by parasitic helminths that constitute a serious public health issue in endemic tropical regions. LF is predominantly caused by Wuchereria bancrofti and Brugia malayi; SF is caused by Loa loa and Onchocerca volvulus. Filarial infections will persist for a lifetime if left untreated, and result in severe physical, psychological, and social impacts on the affected individual. Thereby, making filariasis the second largest cause of permanent and long-term disability worldwide. The diagnosis of filariasis is challenging due to nonspecific clinical manifestation and complete asymptomaticity in most cases. Accurate diagnosis is, however, critical as treatments differ between species and inappropriate treatment regimen can cause severe side effects including permanent blindness, and fatal encephalopathy.New, timesaving, economical, and sensitive diagnostic methods are needed, especially in locations co-endemic for all four major causative agents. Here, we describe our work on developing a TaqMan multiplex real-time PCR assay for the simultaneous detection of the four major filarial species. We demonstrated that the designed RT-PCR primers and TaqMan probes are highly specific for each species target in the singleplex assays with no cross reactivity. Preliminary findings on duplex analysis for both L. loa and O. volvulus reveal promising results with no cross reactivity thus far. However, more work is required for optimization of the L. loa and B. malayi duplex. With comparable sensitivity to established singleplex assays, this reliable and valuable tool will provide significant cost and labour savings for disease monitoring efforts in co-endemic regions.
Functional characterization of germline SMG1 mutations associated with pancreatic ductal adenocarcinoma.
Tatiana Lenko1,2, Francis Robert3, Yifan Wang1,2, Jerry Pelletier3 George Zogopoulos1,21The Research Institute of the McGill University Health Centre, Montréal, QC, Canada, 2The Goodman Cancer Research Centre of McGill University, Montréal, QC, Canada
3Department of Biochemistry, The Rosalind and Morris Goodman Cancer Research Center, McGill University, Montréal, QC, Canada
Corresponding Author: Tatiana Lenko, email tatiana.lenko@mail.mcgill.ca
Abstract
Pancreatic ductal adenocarcinoma (PDAC) has a 5-year survival of less than 10% and is predicted to become the second leading cause of cancer-related mortality by 2030. Approximately 10% of PDAC cases are hereditary. A strategy to improve outcomes of PDAC is surveillance of individuals at high risk for developing PDAC. Thus, identifying the full spectrum of PDAC susceptibility genes has important clinical implications. A recent region-based gene association study from our lab identified SMG1 as a novel PDAC susceptibility gene. We identified 14 germline variants driving the association of SMG1 with PDAC. Here, the c.10921A>G variant, a missense variant in a domain known to be sensitive to mutagenesis, was selected for characterization. Using CRISPR/Cas9 gene editing, we successfully knocked-in c.10921A>G into the HEK293T cell line. Mutants were phenotypically characterized tumorigenic qualities, using proliferation, migration, and wound-healing assays. Migration was significantly higher in the mutant cell line (p<0.05). Furthermore, evaluation of the impact of this mutation on the role of SMG1 in nonsense-mediated decay (NMD) revealed that, while baseline function of NMD is not different between mutant cell lines and wild type, the addition of an NMD inhibitor 52ahas a significantly smaller effect on mutant cell lines (p<0.05). These findings have identified functional impacts of an SMG1 mutation found to associate with PDAC. Studies are underway to further evaluate this mutation, including its metastatic potential in vivo, and to characterize additional SMG1 germline variants associated with PDAC.
Transglutaminase 1 - a novel factor regulating osteoclastogenesis?
Ebrahimi Samani S1, Kaartinen MT1,21Division of Experimental Medicine, McGill University, Montréal, QC, Canada
2Faculty of Dentistry, McGill University, Montréal, QC, Canada
Corresponding Author: Sahar Samani, email sahar.ebrahimisamani@mail.mcgill.ca
Abstract
Osteoclasts are monocyte/macrophage lineage cells capable of fusing into large multinucleated cells that dissolve and remove bone matrix as part of regular maintenance of bone mass and quality. Increased osteoclast activity can cause osteoporosis and augment fracture risk. Discovering cellular factors that regulate osteoclastogenesis can help find paths to novel therapeutics for bone loss. We and others have demonstrated that three transglutaminase (TG) family members FXIII-A, TG2, and TG1 are active in osteoclastogenesis and modulate osteoclast differentiation and fusion. Transglutaminases catalyze formation of isopeptide crosslinking in their substrate proteins. It is not known how TGs mediate osteoclastogenesis or if there is a functional overlap between the three enzymes. The in vitro and in vivo studies showed that the balanced TG2 (Tgm2) expression in osteoclasts is important for bone resorption, as its deletion in mice (Tgm2-/-) causes increased osteoclastogenesis. In this study we report that the increased osteoclastogenesis of Tgm2-/- can be completely reversed by a TG inhibitor, NC9, suggesting that another TG is responsible for the increased osteoclastogenesis. TG2 can also regulate Cathepsin D in cells which is known to proteolytically activate TG1. We show that Cathepsin D levels in Tgm2-/- osteoclast extracts are, indeed, significantly increased which coincides with significantly increased TG1 activity measured via highly specific K5 ‘Hitomi peptide’. Moreover, TG1 activity was completely blocked in cells grown with Pepstatin A, a Cathepsin D inhibitor. This provides further evidence that TG1 and Cathepsin D may be promoters of osteoclastogenesis and excellent targets to control loss of bone mass.
TITLE Transforming growth factor ß canonical pathway implications in melanoma stem cell maintenance
Julien Boudreault1, Ni Wang2, Jean-Jacques Lebrun31Division of Experimental Surgery, McGill University, Montréal, QC, Canada
2Research Institute of the McGill University Health Centre, McGill University, Montréal, QC, Canada
3Division of Experimental Medicine, McGill University, Montréal, QC, Canada
Corresponding Author: Julien Boudreault, email julien.boudreault2@mail.mcgill.ca
Abstract
Transforming Growth Factor beta (TGFß), a secreted protein including three different isoforms (TGFß1, TGFß2 and TGFß3), and other of its family members play an essential role in embryonic and developmental processes. While acting as a tumor suppressor in normal cells and early carcinomas, TGFß shifts its roles towards tumor promoter in more advanced stage cancers. However, activation of TGFß pathway has an opposite outcome in melanoma, acting in a tumor-suppressive manner. We focused on the contribution of TGFß on melanoma stem cells maintenance, primary tumor formation and metastasis. We found that TGFß treatment inhibits melanosphere formation in numerous melanoma cell lines under low-attachment condition. Additionally, CD133+ sub-population reduction, induced by TGFß, was observed in an aggressive metastatic melanoma cell line, which can be blocked in a Smad3/Smad4-specific manner. Our in vivo data demonstrated that injection of Smad2, Smad3 and Smad4 knock-out melanoma cells promoted tumorigenesis and metastasis. Our results present new evidence for the implication of the canonical axis of TGFß suppressive effect on melanoma stem cell maintenance, therefore highlighting new potential therapeutics for the treatment of this deadly cancer.
Understanding p66ShcA-dependent regulation of GPNMB in response to oxidative stress in triple-negative breast cancer?
Clark Thomson1,2, Eduardo Cepeda Canedo3,4, Josie Ursini-Siegel2,3,4, Peter M. Siegel1,2,51Goodman Cancer Research Centre, McGill University, Montréal, QC, Canada
2Department of Biochemistry, McGill University, Montréal, QC, Canada
3Lady Davis Institute for Medical Research, Montréal, QC, Canada
4Division of Experimental Medicine, McGill University, Montréal, QC, Canada
5Department of Medicine, McGill University, Montréal, QC, Canada
Corresponding Author: Clark Thomson, email clark.thomson@mail.mcgill.ca
Abstract
Background: Triple-negative breast cancer (TNBC) is an aggressive and difficult to treat subtype as it lacks specific molecular targets that can be exploited therapeutically. Data generated in our lab identified GPNMB, a cell surface protein over-expressed in TNBC, as a mediator of pro-metastatic phenotypes that correlates with poor prognosis. In response to cellular stress, GPNMB is upregulated following nuclear translocation of the MITF/TFE family of transcription factors (TFE3 and TFEB), which induce GPNMB expression. One class of stressors we have shown upregulates GPNMB are compounds that increase levels of reactive oxygen species (ROS). Furthermore, our data suggests that GPNMB upregulation in response to elevated ROS is dependent on the redox protein p66ShcA. ShcA encodes 3 isoforms (p46, p52 and p66) of which p66ShcA is longest. Importantly, p66ShcA has been shown to promote intracellular ROS formation.
Methodology: We employ various in vitro techniques and a panel of TNBC/basal-like cells to characterize expression, localization, and ROS levels of the p66ShcA-ROS-GPNMB cascade. We have shown that p66ShcA null cells are unable to induce GPNMB expression following ROS-inducing stimulus (NaAsO2) compared to wild-type cells. Unexpectedly, expression of mutant forms of p66ShcA that cannot generate mitochondrial ROS also rescue GPNMB expression. Furthermore, we demonstrate that the mechanism through which p66ShcA modulates GPNMB expression is TFE3/TFEB dependent, as depleting TFE3/TFEB by siRNA attenuates GPNMB upregulation following oxidative stress.
Conclusions: We have identified a non-mitochondrial role for p66ShcA in GPNMB induction following cellular stressors that induce ROS. Further experiments will be designed to characterize the p66ShcA-ROS-GPNMB cascade
Nonalcoholic fatty liver disease in premenopausal women with polycystic ovary syndrome: systematic review & meta-analysis
Mohamed Shengir1, Tianyan Chen2, Elena Guadagno3, Agnihotram V. Ramanakumar4, Peter Ghali2, Marc Deschenes2, Philip Wong2, Srinivasan Krishnamurthy5, Giada Sebastiani21Division of Experimental Medicine, Department of Medicine, McGill University, Montréal, QC, Canada
2Division of Gastroenterology and Hepatology, Department of Medicine, McGill University Health Centre, Montréal, QC, Canada
3Harvey E. Beardmore Division of Pediatric Surgery, McGill University Health Centre, Montréal, QC, Canada
4Research Institute, McGill University Health Centre, Montréal, QC, Canada
5Department of Obstetrics and Gynecology, McGill University Health Centre, Montréal, QC, Canada
Corresponding Author: Mohamed Shengir, email mohamed.shengir@mail.mcgill.ca
Abstract
Background & Rationale: Nonalcoholic fatty liver disease (NAFLD) and polycystic ovary syndrome (PCOS) are prevalent conditions sharing pathogenic etiologies. We performed a meta-analysis aiming to investigate the association between NAFLD and PCOS among premenopausal PCOS patients.
Methods: Relevant studies till 2019 were systematically retrieved from scientific databases. Pooled OR was calculated using random-effect model, and heterogeneity was addressed through I2. Subgroup analyses and meta-regression for various covariates were applied.
Results. 23 studies were qualified for quantitative synthesis. The pooled result showed that PCOS patients had 2.5-fold increase in the risk of NAFLD compared to controls (OR 2.49, 95% CI 2.20–2.82). In subgroup analyses comparing PCOS to controls, South American/Middle East PCOS patients had a greater risk of NAFLD (OR 3.55, 95% CI 2.27–5.55), compared to their Europe (OR 2.22, 95% CI 1.85–2.67) and Asia (OR 2.63, 95% CI 2.20–3.15) counterparts. Insulin resistance (IR) and metabolic syndrome were also frequent in PCOS group (OR 1.97, 95% CI 1.44–2.71 and OR 3.39, 95% CI 2.42–4.76, respectively). When we stratified by BMI, overweight/obese PCOS were at increased risk of NAFLD (OR 3.84, 95% CI 3.25–4.54) whereas lean ones had no association (OR 0.98, 95% CI .077–1.25). However, IR and metabolic syndrome remained significant in lean PCOS. Additionally, BMI also showed a relationship with NAFLD in meta-regression, with a regression coefficient -1.929 (95% CI -3.776 – -0.0826).
Conclusion: This meta-analysis indicates that premenopausal PCOS is associated with 2.5 risk of NAFLD, and BMI seems the main cofactor.
The Role of Memory Gamma Delta T Cells in Hypertension and Vascular Damage
Kevin Comeau1, Pierre Paradis1, and Ernesto L. Schiffrin1,21Hypertension and Vascular Research Unit, Lady Davis Institute for Medical Research, Montréal, QC, Canada
2Department of Medicine, Sir Mortimer B. Davis-Jewish General Hospital, McGill University, Montréal, QC, Canada
Corresponding Author: Kevin Comeau, email Kevin.comeau@mail.mcgill.ca
Abstract
Background: We recently demonstrated that ?d T cells participate in the pathogenesis of hypertension. Evidence also suggests that memory T cells develop during an initial hypertensive episode, sensitizing mice to develop hypertension to further mild hypertensive challenges. However, whether memory ?d T cells develop and play a role in hypertension remains unknown. Our objective is to determine if memory ?d T cells sensitize mice to develop hypertension in response to a mild hypertensive challenge.
Methods: Ten-12-week-old C57BL/6J mice were exposed or not to a hypertensive challenge (490 ng/kg/min angiotensin II (Ang II), SC) for two weeks, followed by a two-week washout period, and then infused with a subpressor dose of Ang II (140 ng/kg/min Ang II, SC) for two weeks. Blood pressure was measured via telemetry and central, effector, and resident memory ?d T cells were profiled by flow cytometry.
Methodology: Mice exposed to the first hypertensive challenge had a systolic blood pressure ~30 mm Hg higher than the sham group after the subpressor hypertensive challenge (P<0.001). After 14-days of Ang II infusion, effector memory ?d T cells increased 4.5-fold in the mesenteric artery perivascular adipose tissue (PVAT), and 2.4-fold in the mesenteric lymph nodes (mLN) (P<0.05). After repeated Ang II infusion, central memory ?d T cells decreased by 65% in the aortic PVAT, and by 23% in the mLN (P<0.05).
Conclusion: An initial exposure to a hypertensive stimulus sensitizes mice to develop hypertension to a subsequent subpressor hypertensive challenge and results in the development of memory ?d T cells.
The added value of the CAT dyspnea and respiratory sub-items as associated with poor CPET outcomes in a random sample of people with COPD and those at risk from the CanCOLD study
Saad Razzaq MSc.(c)1, Hayley Lewthwaite Ph.D.1, Dennis Jensen Ph.D.11Faculty of Education, McGill University, Montréal, QC, Canada
Corresponding Author: Saad Razzaq, email saad.razzaq@mail.mcgill.ca
Abstract
Chronic obstructive pulmonary disease (COPD) is a common disease characterized by persistent respiratory symptoms, most significantly dyspnea, and exercise intolerance [1-4]. Symptom-limited peak VO2 on cardiopulmonary exercise testing (CPET), an index of exercise capacity, has emerged as a robust independent predictor of mortality in people with COPD [3]. The COPD assessment test (CAT) is an eight-item questionnaire focused on respiratory and non-respiratory symptoms commonly used to assess COPD patients [5]. We assessed to which extent single and clustered CAT sub-items, as opposed to the CAT total score, carry additional information regarding abnormal responses to CPET. The study included 1003 adults aged =40 years from the population-based Canadian Cohort Obstructive Lung Disease (CanCOLD) study. A CAT dyspnea score of 2 and CAT respiratory symptom cluster score of 4 were found to best predict peak VO2 less than the lower limit of normal (LLN) [6]. Participants were then sub-grouped by low CAT total score (<10) or high CAT total score (10), with CAT dyspnea or CAT respiratory cluster score above (high) or below (low) the identified threshold value. A high CAT dyspnea and respiratory symptom burden within the low CAT total group identified participants with a significantly lower peak VO2 response. The high CAT dyspnea and CAT respiratory cluster also identified close to twice as many and 1.5x as many participants with peak VO2 < LLN. This study supports the use of CAT sub-items to facilitate early identification of people with or at-risk of developing COPD with impaired exercise intolerance that may be amenable to personalized pharmacological therapy.
The Clinical Utility of a Novel, Multi-parametric, Contrast-Free CMR Imaging Technique to Acquire Tissue Characterization Properties
Katerina Eyre1, Saad Razzaq1, Matthias Friedrich1,21McGill University Health Centre, Montréal, QC, Canada
2Circle Cardiovascular Imaging, Calgary, AB, Canada
Corresponding Author: Katerina Eyre, email katerina.eyre@mail.mcgill.ca
Abstract
Background: Cardiovascular Magnetic Resonance (CMR), a versatile imaging modality, provides information on cardiac morphology and function, though limited by long scan times and patient discomfort [1]. Multitasking, a multi-parametric, contrast-free technique, can resolve cardiac and respiratory motion to reduce scan times without compromising T1 mapping characterization accuracy and image quality.
Methods: Fourteen healthy volunteers (mean age = 43) underwent a cardiac MRI exam at the McGill University Health Centre (MUHC) using both the Multitasking protocol (CINE, T1 and T2 maps) and the standard protocol (bSSFP CINE, MOLLI 5:3:3 T1 and TRUFI T2 maps) on a 3T Skyra Scanner (SIEMENS, Germany).
Methodology: A paired t-test showed no significant differences in global T1 values between the Multitasking (mean = 1208, SD = 119.8) and standard protocol (mean = 1235.1, SD = 43.15). A linear mixed effects model showed that age, HR, BMI, and sex do not significantly confound global T1 value differences between protocols. The Multitasking protocol (time = 11±4 mins) was significantly shorter than the standard protocol (time = 25±10 mins) (p < 0.05). Multitasking and the bSSFP CINE AND MOLLI showed significant differences in average image quality (p < 0.05). An ease-of-use survey found that both protocols were user-friendly for experienced technologists, with only 17% of technologists repeating the Multitasking protocol compared to 67% for the standard protocol, with fewer significant artifacts during Multitasking.
Conclusion: Multitasking, an easy-to-use multi-parametric technique, produces accurate contrast-free T1 mapping values faster than standard protocols. Future work should assess clinical feasibility in larger cohorts.
Identify Targetable Molecular Drivers of Chemotherapy Resistance in Gastro-Esophageal Adenocarcinoma
Iris Kong11McGill University, Montréal, QC, Canada
Corresponding Author: Iris Kong, email mingyang.kong@mail.mcgill.ca
Abstract
Introduction: Gastro-esophageal Adenocarcinoma (GEA) are one of the most lethal malignancies in the Western world. Even with the standard-of-care treatment, 40% of the patients are innately resistant and half of the initial responders develop acquired resistance.
Methods: Patient-derived organoids (PDOs) were established from primary tumor tissue and treated with chemotherapy drugs. Ex-vivo drug responses were correlated with in-patient pathological responses. The multi-omics data obtained from whole-exome (WES) and RNA sequencing of tumor samples were analyzed to identify targetable molecules. High-throughput drug screens with inhibitors against candidate molecules were performed using the liquid handler to identify novel therapeutic strategies with maximal efficacy.
Methodology: The cell viability of PDOs of chemo-resistant patients was significantly higher than those of chemo-sensitive patients after treatment. WES of tumor from patient 0588 revealed an increased copy number of EGFR. Their PDOs showed high sensitivity to an EGFR inhibitor in high-throughput drug screens. Single-cell RNA sequencing of pre- and post-treatment tumor tissue from patient 0055 revealed a broadly shift transcriptome. There were significant changes in cell composition including a drastic increase of immune cells and a decrease of epithelial cells.
Conclusions: PDOs recapitulate patient response to chemotherapy, making them a powerful tool for drug discovery. WES of tumor tissue reveals mutations that can be exploited therapeutically ex-vivo. Single-cell RNA sequencing of naïve and treated tumor tissue reveals cell state and composition changes as possible sources of chemo-resistance. Understanding the molecular basis of chemotherapy resistance is of fundamental importance for designing novel therapeutic strategies for GEA patients.
A Novel Approach of Treatment: Topical Application of an Immunomodulatory Peptide in a Murine Model of Epicutaneously-Induced Allergic Inflammation
Fatima Hubaishi1, Curtis Quan1, Annie Beauchamp1, Louis Cyr1, Jichuan Shan2, Elizabeth D. Fixman2 & Brian J. Ward11Research Institute of McGill University Health Centre, Montréal, QC, Canada
2Meakins-Christie Laboratories, Research Institute of the McGill University Health Centre, Montréal, QC, Canada
Corresponding Author: Fatima Hubaishi, email fatima.hubaishi@mail.mcgill.ca
Abstract
Atopic Dermatitis (AD) is one of the most common chronic inflammatory skin diseases that affect both children and adults. Intense itchiness and dryness are some of the clinical features of AD, leading to a continuous relapsing course of the disease. Some children with early onset of AD are likely to develop other allergic conditions such as asthma, known as atopic march. In addition to the social and financial burden of the disease, current topical treatments target only the symptoms but not the underlying immune mechanism(s) causing AD. Thus, there is great interest in developing topical therapies that inhibit the immune response to improve the lives of AD patients and to curb the atopic march. Our lab developed an immunomodulatory compound called STAT6-IP, which inhibits the development of inflammatory allergic airway responses in mouse models of asthma. I have developed a mouse model of AD using clinically relevant allergens, house dust mite (HDM) and papain. My data show that topical application of HDM or papain induces an inflammatory response in the skin and the lymph nodes near the site of allergen application. A subset of these responses is reduced by STAT6-IP applied to the skin. Thus, my data show that topical application of STAT6-IP has the potential to reduce the AD allergic response. I have more recently developed a robust model of atopic march in order to examine the effects of topical STAT6-IP delivery (to the skin) in its prevention.
Echocardiographic markers in early postnatal life to predict intervention in coarctation of aorta.
Punnanee Wutthigate1,2, Jessica Simoneau1, Gabriel Altit1,1Division of Neonatology, Montreal Children’s Hospital, McGill University, Montréal, Canada.
2Division of Neonatology, Department of Pediatrics, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
Corresponding Author: Punnanee Wutthigate, email punnanee.wutthigate@mail.mcgill.ca
Abstract
Background: Coarctation of the aorta (CoA) is challenging to diagnose in early neonatal life due to the patent ductus arteriosus (PDA). The aim of this study was to describe resource utilization, early post-natal cardiac profile and predictors for cardiac surgery.
Methods: Retrospective single-center study of those with antenatal CoA suspicion =37 weeks at birth between January 2014 and March 2020. Echocardiography (ECHO) was performed within 48 hours after birth.
Methodology: A total of 51 newborns were included (11 [22%] underwent surgery). In normal (n=40), median hospitalization was 4 [2-6] days, 38 (95%) were on intravenous fluids, 9 (23%) infants required gavage and age at full feeds was 3 days (IQR: 2-5). PDA was patent in all newborns and aortic measurements (ascending [p=0.01], proximal [p<0.001] and distal transverse [p<0.001], isthmus [0.36 [0.07] vs 0.26 [0.07] cm; p<0.001]) were significantly smaller in the CoA group. Right ventricular predominance was higher in CoA, as demonstrated by the left ventricular end-systolic eccentricity index (EI) (1.60 [0.28] vs 2.16 [0.45]; p<0.001). Isthmus and EI were highly predictive for intervention need (OR 26, 95%CI (-42 to -9); p=0.002, OR 4.5, 95%CI (1.5-7.5); p=0.003).
Conclusions: Majority of those with antenatal CoA suspicion did not require intervention but were high consumers of resources. Despite PDA patency, isthmus and EI on first post-natal ECHO were predictors for need of surgery.
Potential diagnostic value of urine microRNAs for female patients with overactive bladder
Stephanie Sirmakesyan1, Philippe Cammisotto1, Abubakr H Mossa1, Samer Shamout1, Lysanne Campeau1,21Lady Davis Institute for Medical Research, McGill University, Montréal, QC, Canada.
2Urology Department, Jewish General Hospital, Montréal, QC, Canada.
Corresponding Author: Stephanie Sirmakesyan, email stephanie.sirmakesyan@mail.mcgill.ca
Abstract
Introduction/Objective(s): MicroRNAs as biomarkers provide valuable clinical and diagnostic information. We recently reported that the urinary ratio of neurotrophin nerve growth factor (NGF) to its precursor (proNGF) was decreased in a cohort of female patients with overactive bladder (OAB). These results were linked to increased activity of the matrix metalloproteinase-9 (MMP-9), the main protease digesting NGF. We measured the miRNAs related to these proteins and those regulating the translation of their receptors p75NTR and TrkA.
Methods: Urine and blood samples from 20 control and 20 OAB female patients (50-80 years) were gathered with validated questionnaires. MicroRNAs were measured by RT-qPCR. Results were adjusted for age, renal function and insulin resistance. Receiver operating characteristics (ROC) were tested.
Methodology: MiRNAs levels controlling proNGF (miR-98-5p, let-7b-5p and let-7d-5p), the survival receptor TrkA (miR-92a-3p and 221-5p), and the markers of nerve integrity (miR-21-5p, miR-132 and miR-212-5p) were similar between groups. On the other hand, miR-491-5p, which controls the translation of MMP-9, was significantly decreased in OAB, which implies higher translation of MMP-9. Similarly, miR-592, which negatively regulates the p75NTR receptor synthesis was down-regulated in OAB. Age, renal function and insulin resistance did not affect these results. ROC curves confirmed a high sensitivity of miR-491-5p and miR-592 for diagnosis.
Conclusion(s): These results confirmed at the translational level the central role of MMP-9 and also suggest a proinflammatory state elicited by p75NTR. Measurements of miR-491-5p and miR-592 in urine could be a useful and non-invasive tool for the diagnosis of OAB syndrome in aging women.
Value of Deep Learning (ESPIRiT) for Improving the Subjective Quality of Highly Undersampled Cine Cardiovascular Magnetic Resonance (CMR) Images
Kate S. Lindsay1, Elizabeth Hillier1, Saad Razzaq1, Xucheng Zhu2, Graeme McKinnon2, Andrew J. Coristine2, Matthias G. Friedrich11McGill University, Montréal, QC, Canada
2GE Healthcare
Corresponding Author: Kate Lindsay, email Katherine.lindsay@mail.mcgill.ca
Abstract
Background: Cardiovascular magnetic resonance (CMR) scan times are reduced with novel methods that under-sample k-space. Variable density spatiotemporal (VD k-t) sampling shortens image acquisitions from 5 or more heartbeats (RR) to 3 or 1 RR, allowing for a short axis stack acquisition during a single-breath hold. A trade-off is reduced image quality. Novel deep learning (DL) reconstructions, such as DL ESPIRiT [1], can fill missing k-space data, improving perceived image quality (IQ). The aim of this study is to compare the IQ of VD k-t images acquired and reconstructed with:1 3RR undersampled (kt-ARC 3RR),2 1RR highly undersampled (kt-ARC 1RR),3 DL reconstruction (DL-ESPIRiT), and4 standard cine (FIESTA).
Methodology: Eight healthy participants and eight patients were scanned on a 3T General Electric (GE) Premier research scanner. A 4-point scale (1=poor, 2=fair, 3=good, 4=excellent) compared IQ in six-combinations of de-identified images: FIESTA, kt-ARC 3RR, kt-ARC 1RR, and DL-ESPIRiT. Statistical significance was determined using ANOVA with multiple comparisons.
Methodology: All subjects were included in the analysis. Significant differences in IQ were noted in all comparisons (see Fig. 1). DL-ESPIRiT showed significantly better IQ than both kt-ARC 3RR (P=0.0015, mean difference= 0.479±0.438) and kt-ARC 1RR (P< 0.0001, mean difference=0.958 ±0.643). FIESTA showed significantly better IQ than kt-ARC 3RR (P< 0.0001, mean difference=0.792 ±0.453), kt-ARC 1RR (P< 0.0001, mean difference=1.875±0.698), and both DL-ESPIRiT at 3RR (P=0.0118, mean difference=0.313 ±0.375) and 1RR (P< 0.0001, mean difference=0.917±0.464).
Conclusion: The subjective quality of VD k-t methods can be greatly improved by DL algorithms like ESPIRiT.
QUAKING regulates microexon alternative splicing of the Rho GTPase pathway and controls microglia homeostasis
Jeesan Lee1, Oscar David Villarreal1, Xiaoru Chen1, Stéphanie Zandee2, Yoon Kow Young1, Cynthia Torok2, Nathalie Lamarche-Vane3,4, Alexandre Prat2, Serge Rivest5, David Gosselin5 and Stéphane Richard1*1Segal Cancer Center, Lady Davis Institute for Medical Research and Gerald Bronfman Department of Oncology and Departments of Biochemistry, Human Genetics and Medicine, McGill University, Montréal, QC, Canada
2Neuroimmunology Research Laboratory, Centre du Recherche du Centre Hospitalier de l'Université de Montréal (CRCHUM), Montréal, QC, Canada
3Centre for Translational Biology, Glen Site of the MUHC, Montréal, QC, Canada
4Department of Anatomy and Cell Biology, McGill University, Montréal, QC, Canada
5Department of Molecular Medicine, Faculty of Medicine, Québec City, QC, Canada
Corresponding Author: Jee-San PhD, email jeesanlee88@gmail.com
Abstract
The role of RNA-binding proteins in regulating the phagocytic and cytokine releasing functions of microglia is unknown. Herein we show that microglia-deficient for the QKI RNA-binding protein had increased proinflammatory cytokine release and defects in processing phagocytosed cargo. Splicing analysis defined a novel role for QKI in regulating microexon networks of Rho-GTPase pathways. QKI-deficient microglia had elevated levels of active RhoA and the proinflammatory cytokine release which was repressed by treating with a Rock kinase inhibitor. QKI-deficient microglia were unable to support CNS remyelination in the cuprizone mouse model and depletion of QKI at sites of remyelination increased oligodendrocyte-precursor cells apoptosis and engulfment by the microglia. The expression of QKI was reduced in preactive, chronic active and remyelinating white matter lesions of the MS patients. Overall, our findings identify QKI as an alternative splicing regulator governing a network of Rho-GTPases microexons and their implication for CNS remyelination and MS patients.
Pathogenic mechanisms of kidney defects in the mutants for PCP effector gene Fuzzy
Rhythm Sharma1, Dr. Elena Torban21Research Institute of McGill University Health Centre (RI-MUHC), Montréal, QC, Canada
2Department of Medicine, McGill University, Montréal, QC, Canada
Corresponding Author: Rhythm Sharma, email rhythm.sharma@mail.mcgill.ca
Abstract
Congenital anomalies of the kidney and urinary tract (CAKUT) are the leading cause ofend-stage renal disorder (ESRD) in children, including cystic kidney diseases caused by mutant genes that affect the primary cilia on renal epithelial cells. We discovered hypoplastic cystic kidneys in the mice mutant for the Planar Cell Polarity (PCP) effector gene, Fuzzy. Fuzzy was first discovered in Drosophila, known for the regulation of actin cytoskeleton. However, in vertebrates Fuzzy participates in ciliogenesis by controlling the trafficking of certain molecules needed for the assembly of the primary cilium. How Fuzzy regulates ciliogenesis or causes cysts in the kidney is not well understood.
Our observations show that Fuzzy might act via p190A to control ciliogenesis. p190A is a GTPase activating protein that inhibits RhoA GTPase activity and regulates actin polymerization at the base of the cilium required for proper cilia elongation. Loss of p190A gene causes shorter cilia, renal hypoplasia and glomerulocystic kidney which are phenotypically similar to the E16.5 Fuzzy-/- kidneys. We showed by CoIP that Fuzzy interacts with p190A, recruits p190A to the ciliary base and that ciliogenesis is rescued in unciliated Fuzzy-/- MEFs by inhibitors of Rho Kinase, a downstream RhoA GAP effector.By crossing p190A and Fuzzy mutant mice, we observed genetic interactions between these genes. Together, our data revealed a molecular, genetic and mechanistic link between Fuzzy and p190ArhoGAP. We propose that Fuzzy controls RhoAGTPase activity at the ciliary base by recruiting p190A to the basal body;inappropriate RhoA activation causes hypoplastic cystic kidney.
ALK4/5/7 and EGF receptors: triggering and maintaining extracellular matrix-associated gene expression during cumulus expansion
Karen F. Carvalho1,3, Hugh J. Clarke1,2,31Division of Experimental Medicine, McGill University, Montréal, QC, Canada
2Department of Obstetrics and Gynecology, McGill University, Montréal, QC, Canada
3Research Institute of the McGill University Health Centre, Montréal, QC, Canada
Corresponding Author: Karen Carvalho, email karen.carvalho@mail.mcgill.ca
Abstract
Oocytes within pre-ovulatory follicles are enclosed by several layers of tightly packed follicular cells known as the cumulus granulosa. In response to the LH surge, cumulus cells undergo a process termed expansion, in which they secrete an extracellular matrix (ECM) and become displaced away from the oocyte. In mice, expansion happens during the 12 hours preceding ovulation, and is believed to be essential for fertility. Epidermal growth factor (EGF)-like peptides, released by the mural granulosa cells in response to LH, and transforming growth factor ß family members secreted by the oocyte are both required for expansion to happen. When each pathway is active and how they cooperate remains unclear. We found that incubation of cumulus-oocyte complexes (COCs) with EGF triggered an increase in the mRNA levels of ECM-related genes within 3h, followed by a decrease over the next 6h. EGFR inhibition beginning 3 or 6h after EGF exposure had a modest impact on gene expression, leading to 2.6- and 1.7-fold decrease in Has2 mRNA levels, respectively, plus a 14 and 12% decrease in the final COC area, a measure of the expansion process. In contrast, ALK4/5/7 inhibition at the same time points lead to a respective 80- and 31-fold decrease in Has2 mRNA levels, which were accompanied by a 29 and 17% decrease in the final COC area. These results suggest that EGFR and ALK4/5/7 signaling are required to trigger initial upregulation of ECM-related genes, but ALK4/5/7 is mainly responsible for maintaining adequate levels of these genes throughout expansion.
Validation of the Thrombolysis in Myocardial Infarction Risk Score for Heart Failure in Diabetes (TRS-HFDM) in Patients with Recent Acute Coronary Syndrome: An Analysis of the EXAMINE Trial
Amir Razaghizad1, Joao Pedro Ferreira2,3, Jiayi Ni1, Faiez Zannad2, Abhinav Sharma11McGill University Health Centre Research Institute, Montréal, QC, Canada
2Université de Lorraine INSERM, Centre d'Investigations, Cliniques Plurithématique, Nancy, France
3Department of Physiology and Cardiothoracic Surgery, Cardiovascular Research and Development Unit, Faculty of Medicine, University of Porto, Portugal
Corresponding Author: Amir Razaghizad, email amir.razaghizad@mail.mcgill.ca
Abstract
Background: The Thrombolysis in Myocardial Infarction Risk Score for Heart Failure (HF) in Diabetes (TRS-HFDM) predicts the risk of HF hospitalization. However, evidence regarding its performance in diabetic patients with established cardiovascular disease is limited.
Methods: Model performance was evaluated through discrimination and calibration for the primary endpoint time to cardiovascular death or HF hospitalization. Validation data were sourced from the Examination of Cardiovascular Outcomes with Alogliptin versus Standard of Care (EXAMINE) trial (n=5,380). Variables in the TRS-HFDM include the history of HF (2 points), atrial fibrillation (1 point) and coronary artery disease (1 point), and estimated glomerular filtration rate (<60 mL/min/1.73 m2·1 point) and urine albumin-to-creatinine ratio (>300 mg/g·2 points; 30-300 mg/g·1 point).
Methodology: A primary endpoint occurred in 402 patients with prevalent coronary artery disease. Accordingly, 25% of patients were judged as intermediate risk (1 point), 30% were judged as high risk (2 points), 19% were judged as very-high risk (3 points), and 26% were judged as severe risk (= 4 points). Discrimination was robust for the primary endpoint (c-index: 0.72). However, calibration was modest as the calibration-in-the-large and calibration slopes ranged between -0.25 and -0.13 and 0.70 and 0.74 for 6- and 30-month event predictions. Performance was generally superior for hospitalization for HF alone (c-index: 0.75); the calibration-in-the-large and calibration slopes ranged between -0.18 and -0.19 and 0.82 and 0.79 for 6- and 30-month event predictions.
Conclusion: The TRS-HFDM robustly predicts and stratifies the risk of HF hospitalization. Increased implementation of the TRS-HFDM may improve patient outcomes and health service utilization.
Right Ventricle Outflow Tract Obstruction in Adults: A Systematic Review
Yu Hao Zeng, MD1, Alexander Calderone, MD1, Nicolas Rousseau-Saine, MD1, Mahsa Elmi-Sarabi, PhD1, Stéphanie Jarry, MSc1, Étienne J. Couture, MD2, Matthew P. Aldred, MBBS, FANZCA1, Jean-Francois Dorval, MD3, Yoan Lamarche, MD, MSc4,7, Lachlan F. Miles, MD5, William Beaubien-Souligny, MD6, André Y. Denault, MD, PhD1,71 Department of Anesthesiology, Montreal Heart Institute, Université de Montréal, Montréal, QC, Canada
2 Department of Anesthesiology and Department of Medicine, Division of Intensive Care Medicine, Institut Universitaire de Cardiologie et de Pneumologie, Québec, QC, Canada
3 Department of Cardiology, Montreal Heart Institute, Université de Montréal, Montréal, QC, Canada
4 Department of Cardiac Surgery, Montreal Heart Institute, Université de Montréal, Montréal, QC, Canada
5 Centre for Integrated Critical Care, The University of Melbourne, Melbourne, Victoria, Australia and Department of Anaesthesia, Austin Health, Melbourne, Australia
6 Department of Medicine, Nephrology Division, Centre Hospitalier de l’Université de Montréal, Montréal, QC, Canada
7 Critical Care Division, Montreal Heart Institute, Université de Montréal, Montréal, QC, Canada
Corresponding Author: Alexander Calderone, email alexander.calderone@mail.mcgill.ca
Abstract
Right ventricular outflow tract obstruction (RVOTO) is a cause of hemodynamic instability that can occur in several situations including cardiac surgery, lung transplantation, thoracic surgery, and in critically ill patients. The timely diagnosis of RVOTO is important because it requires
specific considerations, including the adverse effects of positive inotropes and, depending on the etiology, the requirement for urgent surgical intervention. The objective of this systematic review and meta-analysis was performed to determine the prevalence of RVOTO in adult patients and the distribution of all reported cases by etiology. Of 233 available reports, there were 229 case reports or series, four retrospective cohort studies, with one study also reporting a prospective cohort. Out of 291 reported cases of RVOTO, 61 (21%) were congenital, 56 (19%) were
iatrogenic and 174 (60%) were neither congenital nor iatrogenic (including intra-cardiac tumour). The mechanism of RVOTO was caused by an intrinsic obstruction in 169 cases (58%) and an extrinsic obstruction in 122 cases (42%). A mechanical obstruction causing RVOTO was
present in 262 cases (90%) whereas 29 cases of dynamic RVOTO (10%) were reported. In the 5 included cohorts with a total of 1122 patients, the overall prevalence was estimated to be 4.0% (1-9%). This complication, while rare, remains clinically important and therefore multicenter studies are warranted to better understand the prevalence, causes and consequences of RVOTO.
Predicting COVID-19 Outcomes using Proteomic Data
Chen-Yang Su1,2, Sirui Zhou1,3, Wonseok Jeon2, Vincenzo Forgetta1, Joelle Pineau2, J Brent Richards1,3,41Lady Davis Institute, Jewish General Hospital, McGill University, Montréal, QC, Canada.
2Department of Computer Science, McGill University, Montréal, QC, Canada.
3Department of Human Genetics, McGill University, Montréal, QC, Canada.
4Department of Epidemiology, Biostatistics and Occupational Health, McGill University, Montréal, QC, Canada.
Corresponding Author: Chen-Yang Su, email chen-yang.su@mail.mcgill.ca
Abstract
Background: Currently, few disease-specific therapies exist for treating severe COVID-19. There is also a need for the prediction of severe SARS-CoV-2 infection, especially as novel variants arise that escape protection of current vaccines. Determining circulating proteins that play key roles during viral infection can provide useful information to identify individuals who are at risk for severe COVID-19 outcomes, such as the need for respiratory support or death. Analyses of large-scale blood proteomic data in COVID-19 patients may help to predict the course and outcome of the disease, and to identify potential biomarkers for drug candidates.
Methods: In this study, we utilized a dataset composed of one of the largest groups of circulating protein biomarkers for COVID-19 patients collected over the course of disease progression to design an accurate, clinically relevant multiprotein machine learning model for the prediction of COVID-19 adverse outcomes. We categorize COVID-19 outcomes into two groups based on severity level and leverage the ability of L1-norm regularization combined with logistic regression (LASSO) for selecting predictive proteins for these adverse outcomes. We then externally validate the robustness of our model in an external dataset with the same proteomic profiling and clinical outcomes.
Methodology: Our best performing model achieved an area under the receiver operating characteristic (AUROC) score of 0.843 and 0.796 for predicting severe and very severe COVID-19, respectively, on the external test set.
Conclusion: Our results suggest that circulating proteins can improve prediction accuracy of severe COVID-19 outcomes, providing evidence for future clinical utility.
Understanding the Role of G3BP1 in Adaptation to Energetic Stress
Ranveer Palia1,2, David Papadopoli1,3, Ivan Topisirovic1,2,31Lady Davis Institute for Medical Research, Montréal, QC, Canada
2Department of Medicine, Division of Experimental Medicine, McGill University, Montréal, QC, Canada
3Gerald Bronfman Department of Oncology, McGill University, Montréal, QC, Canada
Corresponding Author: Ranveer Palia, email ranveer.palia@mail.mcgill.ca
Abstract
Cancer cells display metabolic plasticity in order to adapt to energetic and environmental stresses. The mitochondria provide cancer cells with the energetic and biosynthetic resources needed for increased proliferation, metastasis, and therapeutic resistance. However, the molecular mechanisms that are exploited by cancer cells to alleviate energetic stress are incompletely understood and constitute an intense area of investigation. Through the use of genome-wide CRISPR Cas9 screens, we uncovered genes whose loss confers differential susceptibility to various mitochondrial poisons. Ras GTPase-activating protein-binding protein 1 (G3BP1) emerged as a top candidate that is protective against mitochondrial inhibition in a model lymphoma cell line. This gene is known to be involved in many cellular pathways including stress granule formation, cGas/STING pathway, and viral DNA detection. However, its role in adaptation to mitochondrial dysfunction-induced energy stress is poorly understood. We aim to decipher the molecular circuitry that binds G3BP1 to the adaptation of cancer cells to energetic stress. To this end, we generated CRISPR-Cas9 mediated knockouts of G3BP1 in numerous cancer cell lines, including lymphoma (Nalm6), KRAS mutated colon cancer (HCT-116), and BRAF mutated melanoma (A375). Loss of G3BP1 impairs cellular proliferation and protein synthesis, while reducing sensitivity to phenformin, an electron transport chain inhibitor. We also observed, that the absence of G3BP1 increases mTOR signaling through the phosphorylation of ribosomal protein S6 and 4EBP1. Taken together, these data link G3BP1 to energy stress adaptation. These data could provide unprecedented insights into orchestration of signalling and mitochondria stress in cancer cells.
STAT6-IP-dependent control of type 2 innate and adaptive immunity in the lung
Haya Aldossary1, Véronique Gaudreault1, Lydia Labrie1, Jichuan Shan1, Brian J. Ward1, Elizabeth D. Fixman1.1Research Institute of McGill University Health Centre, Montréal, QC, Canada
Corresponding Author: Haya Aldossary, email haya.aldossary@mail.mcgill.ca
Abstract
Airway inflammation is one of the main characteristics of allergic airway diseases, including allergen-induced asthma. Evidence, primarily from animal studies, supports a model in which inhalation of allergens triggers epithelial cell release of innate cytokines, including IL-33, which stimulate group 2 innate lymphoid cells (ILC2) to produce large amounts of IL-13 and IL-5, which in turn amplify allergic inflammation. Published data from our labs indicate that intranasal administration of an immunomodulatory peptide, STAT6-IP, at the time of antigen priming inhibits Th2 adaptive immunity in murine models of asthma.Inhibition is thought to occur, at least in part, through inhibition of dendritic cells (DC) and ILC2 at the time of priming.
In this study, we sought to clarify how STAT6-IP affects DC responses in the lung and the lung draining mediastinal lymph nodes (MLN) induced by IL-33 and ovalbumin (OVA).
Our data show that STAT6-IP inhibited OVA/IL-33-induced recruitment to and activation of lung DC, which in turn reduced DC migration to the MLN. Because ILC2-derived IL-13 promotes DC migration to the MLN, we also quantified ILC2 responses. As expected, both total as well as IL-13-expressing ILC2 were reduced in mice treated with STAT6-IP. In addition, intranasal delivery of STAT6-IP inhibited CD4+ Th2 differentiation in the MLN. Consistent with these data, allergic inflammatory responses, including airway hyperresponsiveness, were reduced in STAT6-IP-treated mice following OVA challenge several weeks later.
Our findings provide insight into mechanisms by which STAT6-IP delivery at the time of antigen priming reduces Th2 adaptive immunity.
Investigating Mechanisms of ß-glucan-induced Trained Immunity
Alexandre Grant1,2, Maziar Divangahi1,21Department of Microbiology and Immunology, Faculty of Medicine and Health Sciences, McGill University, Montréal, QC, Canada
2Research Institute, McGill University Health Center, Montréal, QC, Canada
Corresponding Author: Alexandre Grant, email alexandre.grant@mail.mcgill.ca
Abstract
Innate immune memory, or trained immunity (TI), is characterized by epigenetic and metabolic reprogramming of innate immune cells upon primary stimulation, leading to enhanced responsiveness or protection upon secondary stimulation. Bacille Calmette-Guérin (BCG) and ß-glucan (a component of the fungal cell wall) are known inducers of TI. Our group has shown that access of BCG to the bone marrow (BM) leads to transcriptional reprogramming of hematopoietic stem cells (HSCs) and to epigenetically imprinted bone marrow-derived macrophages (BMDMs) that were more protective against Mycobacterium tuberculosis (Mtb) infection. We have also shown that ß-glucan administration reprograms HSCs and confers enhanced protection against Mtb infection. However, contrary to BCG or ß-glucan, we have recently demonstrated that virulent Mtb prevents HSC-mediated trained immunity. These studies collectively indicate that while we can target HSCs to induce TI, pathogens have evolved immune evasion mechanisms to prevent TI. However, it remains unclear how HSC reprogramming is transmitted to progeny cells to generate TI. We hypothesize that1 ß-glucan can only induce TI in HSCs and not in fully differentiated myeloid cells (e.g. macrophages), and2 ß-glucan reprogramming of HSCs is epigenetically transmitted to trained myeloid cells. To test this hypothesis, we propose to use a novel ex vivo model of TI induction in HSCs and a transgenic mouse model to conduct HSC fate mapping in TI. Elucidating the role of HSC reprogramming in TI-mediated mechanisms of host protection will aid the development of TI-based vaccines and therapeutics against important pathogens such as Mtb.
Investigating the role of central carbon metabolism in human coronavirus replication
Magan Solomon1,2, Qinghua Pan2, Chen Liang1,2,31Department of Medicine, McGill University, Montréal, QC, Canada
2Lady Davis Institute, Jewish General Hospital, Montréal, QC, Canada
3Department of Microbiology & Immunology, McGill University, Montréal, QC, Canada
Corresponding Author: Magan Solomon, email magan.solomon@mail.mcgill.ca
Abstract
Since the COVID-19 pandemic was declared in March 2020, the number of SARS-CoV-2 infections have continued to increase due to, in part, the lack of antiviral treatments that are required to prevent new infections. Identification of viral and cellular factors that are essential for human coronavirus replication is pivotal to highlight feasible targets for the development of antiviral treatments for COVID-19 and other human coronavirus infections. Viruses hijack cellular central carbon metabolism to induce large-scale alterations in these pathways enabling viruses to meet their specific metabolic demands that are essential for viral replication. Here, we use the common cold human coronavirus, HCoV-229E, as a model to investigate which cellular central carbon metabolism pathways are essential for human coronavirus replication using a pharmacological approach to target glycolysis, the citric acid cycle and oxidative phosphorylation, and carbon sources supplementation. We show that glycolysis is essential, but not sufficient to support HCoV-229E replication in Huh7 cells while a combination of glycolysis and oxidative phosphorylation is essential for HCoV-229E replication in A549 cells and enhances viral replication in Huh7 cells. We also show that in primary human airway epithelial cells, a more clinically relevant model, glycolysis is not required for HCoV-229E production. Together, the results highlight that the dependency of HCoV-229E replication on specific central metabolic pathways differs between cell types and between transformed and primary cells.
The immunomodulatory potential of excretory/secretory metabolite(s) of a parasitic worm as therapy in inflammatory bowel disease
Elizabeth Siciliani1, Mifong Tam2, Joseph F. Urban, Jr.3, Richard J. Martin4, Fernando Lopes1, Timothy G. Geary1, Mary M. Stevenson2, Armando Jardim11Institute of Parasitology, McGill University, Montréal, QC, Canada
2Department of Microbiology and Immunology, McGill University, Montréal, QC, Canada
3USDA, 4. College of Veterinary Medicine, Iowa State University, Ames, IA, USA
Corresponding Author: Elizabeth Siciliani, email elizabeth.siciliani@mail.mcgill.ca
Abstract
Helminth therapy in patients with autoimmune diseases such as multiple sclerosis and inflammatory diseases such as inflammatory bowel disease (IBD) has shown promise in ameliorating disease in clinical trials. Excretory/secretory products (ESP) released by helminths have potent immunomodulatory effects on immune cells. Rather than infecting patients with live parasites, a more rational approach is to identify the immunomodulatory molecule(s) in ESP and develop these as potential therapy. We observed that ESP from gastrointestinal nematodes including Ascaris suum suppress TNF-a secretion and induce IL-10 secretion by murine bone marrow-derived macrophages (BMDM) stimulated with LPS. In addition to proteins, metabolites in A. suum ESP were found to have similar effects on BMDM. To identify the metabolite(s) responsible for the modulatory effects, ESP was analysed by LC-MS/MS, separated by RP-HPLC, and the bioactivity of the fractions assessed on LPS-stimulated BMDM. Using this approach, a metabolite fraction (AsNP25), had TNF-a suppressing and IL-10 enhancing effects on LPS-stimulated BMDM. To further characterize AsNP25, we investigated the immunomodulatory effects on PBMC. AsNP25 suppressed TNF-a secretion and induced IL-10 secretion by human monocytes in response to LPS. The therapeutic effects of AsNP25 were investigated in DSS-induced colitis in C57BL/6 mice. Treatment with AsNP25 resulted in significantly longer colons, a lower histopathology score, and decreased TNFa mRNA expression in colon tissue compared to colitic mice treated with PBS. Together, these data indicate that non-polar metabolite(s) in A. suum ESP modulate pro-inflammatory and anti-inflammatory cytokine secretion in vitro and in vivo and ameliorate disease in DSS-induced colitis in mice.
The role of CTCF in the context of skeletal muscle
Yuqing Yang1, Solène Jamet2, Colin Crist2,3, Michael Witcher1,31Division of Experimental Medicine, McGill University and Lady Davis Institute, Montréal, QC, Canada
2Department of Human Genetics, McGill University and Lady Davis Institute, Montréal, QC, Canada
3Corresponding Author
Corresponding Author: Yuqing Yang, email yuqing.yang3@mail.mcgill.ca
Abstract
CCCTC-binding factor (CTCF) is an indispensable protein in vertebrates and complete Ctcf deficiency in mice leads to early implantation lethality. As a transcription factor and an insulator, CTCF has multivalent interactions with DNA and proteins and regulates epigenetic activities. It has been shown that CTCF involves in the myogenic differentiation program. However, the role of CTCF in skeletal muscle in vivo has not been clearly elucidated. Our novel study will focus on the mechanism of CTCF in skeletal muscle by studying the Ctcf deletion mice model.
Here, we generated a mice model HSACre/+; Ctcffl/fl. They exhibit progressive weight loss and skeletal muscle wasting, but can develop normally and be fertile. Our preliminary results show that both tibialis anterior muscle and diaphragm muscle of these mice are characterized by newly formed myofibers, which indicates the presence of muscle injury. And we will analyze the transcriptional changes and epigenetic changes due to the Ctcf deletion in skeletal muscle by RNA-seq and ChIP-seq. This research is working towards a better understanding of CTCF mechanisms in the differentiation process of skeletal muscle and providing more ideas for treating skeletal muscle wasting.
THX-B compound decreases the activity of matrix metalloproteinase-9 and increases secretion of nerve growth factor by mouse urothelial cells in culture
Aalya Hamouda1, Stephanie Sirmakesyan1, Philippe Cammisotto1, Uri Saragovi1, Lysanne Campeau1,21Lady Davis Institute for Medical Research, McGill University, Montréal, QC, Canada
2Urology Department, Jewish General Hospital, Montréal, QC, Canada
Corresponding Author: Aalya Hamouda, email aalya.hamouda@mail.mcgill.ca
Abstract
Introduction: Low levels of Nerve Growth Factor (NGF) have been observed in the urine of patients with overactive bladder (OAB) and of type 1 diabetic rodents. This was linked to high activity of metalloproteinase-9 (MMP-9). Moreover, THX-B, an inhibitor of the proinflammatory receptor p75NTR, restored normal levels of NGF in type 1 diabetic mice, suggesting an involvement of p75NTR in MMP-9 expression. We examined the in vitro effect of THX-B on MMP-9 in bladder cells.
Methodology: Primary cultures of urothelial cells (UCs) and smooth muscle cells (SMCs) are grown from mice (C57BL/6) bladders. NGF and MMP-9 expressions are assessed by RT-qPCR, immunohistochemistry, ELISA and enzymatic assay.
Methodology: NGF and MMP-9 mRNAs were expressed in UCs and SMCs at similar levels. Microscopy confirmed the localization of both proteins in cells. UCs and SMCs were both major sources of NGF. MMP-9 protein content was 7 times higher in SMCs than in UCs. However, secretion of active MMP-9 was 40 times higher in UC medium. Incubation with THX-B abolished the synthesis and secretion of MMP-9 and doubled the NGF concentration in UC medium. THX-B had little effect on NGF and MMP-9 in SMC medium.
Conclusion: Bladder cells secrete NGF, proNGF and MMP-9. Further, UCs appear to be the primary target for THX-B. These results are in accordance with our previous publications on OAB patients and type 1 diabetic rodents. To this extent, THX-B may be a therapeutic tool for improving symptoms of OAB through primarily targeting MMP-9 in the urothelium.
Investigating Nuclear Functions of MNK1
Omar Moussa1,2, Sathyen Prabhu1,2, Laura Quiron3, Jean-Francois Coté (3,5,6,7,8), Wilson H. Miller Jr. (1,2,4), Sonia V. del Rincon (1,2,4).1Division of Experimental Medicine, McGill University, Montréal, QC, Canada
2Lady Davis Institute, Jewish General Hospital, Montréal, QC, Canada
3Institut de Recherches Cliniques de Montréal (IRCM), Montréal, QC, Canada
4McGill Centre for Translational Research in Cancer, McGill University, Montréal, QC, Canada
5Molecular Biology Programs, Université de Montréal, Montréal, QC, Canada
6Department of Medicine, Université de Montréal, Montréal, QC, Canada
7Department of Biochemistry and Molecular Medicine, Université de Montréal, Montréal, QC, Canada
8Department of Anatomy and Cell Biology, McGill University, Montréal, QC, Canada
Corresponding Author: Omar Moussa, email omar.moussa@mail.mcgill.ca
Abstract
Melanoma is the deadliest form of skin cancer due to its high propensity to metastasize. Recent advances in immunotherapy and targeted therapies have revolutionized the treatment of advanced melanoma. However, major challenges remain including the development of resistance and the occurrence of untreatable subtypes. Thus, novel therapeutic approaches are required to further ameliorate the overall survival of patients with metastatic melanoma. MAPK-interacting protein kinases (MNK1/2) are serine/threonine kinases known to phosphorylate eukaryotic initiation factor 4E (eIF4E). Interestingly, deregulation of the MNK1/2-eIF4E axis has been shown to promote oncogenicity and metastasis. Although, MNK1/2 inhibitors are in phase 2 clinical trials for solid tumours, large gaps in our knowledge remain concerning the underlying molecular mechanisms by which MNK1/2 exert pro-carcinogenic functions. In our clinical characterization of MNK1/2 activity in patient-derived tumours, we noted that MNK1 is present both in the cytoplasm and nucleus, however little is known concerning the role of nuclear MNK1. Therefore, to investigate the role of MNK1 in the nucleus we generated an in vitro experimental model system. We used MNK1 knocked-out D4M.3a murine melanoma cells wherein we re-expressed a1 MNK1 harbouring a mutated nuclear export signal of MNK1 (exclusively nuclear) or2 MNK1 with its nuclear localization signal mutated (exclusively cytoplasmic). We will use these cell lines in in vitro and in vivo experiments to characterize the impact of these MNK1-mutant expressing cell lines on invasion and metastasis. Finally, to identify nuclear substrates of MNK1, we designed a mass spectrometry workflow incorporating proximity-dependent biotin identification (BioID).
The McGill MD-PhD Program: 35 Years Later
Joan Miguel Romero1*, Mark Sorin1*, Matthew Dankner1, Heather Whittaker1, Mark Eisenberg11Faculty of Medicine, McGill University, Montréal, QC, Canada
*Authors contributed equally to this work
Corresponding Author: Joan Miguel Romero, email joan.romero@mail.mcgill.ca
Abstract
Background: MD-PhD programs aim to train future physician-scientists in both research and medicine. In this study, we assessed the career trajectories of alumni of the McGill MD-PhD program since its inception.
Methods: Information was collected using an online survey consisting of 64 questions. Alumni of the program were contacted by email and phone.
Methodology: Survey response rate was 85% (40/47). The average number of post-graduate training years was 7.0 ± 2.1 (mean ± SD).The median number of publications produced by alumni from their PhD research was 5 (for those having started their graduate training between 1985-1994), 3 (1995-2004), and 8.5 (2005-2014). As of 2021, 47 have completed their MD-PhD training, with a current enrollment of 21 spread across medical and graduate training. Of alumni having finished training, 67% (22/33) are in academia, 24% (8/33) are in non-academic clinical practice, and the remaining work in international development (1/33), industry (1/33), or the military (1/33). With respect to protected research time, 45% of alumni dedicate at least 25% of their time to research, with 73% of these dedicating at least 50%. The most frequent research they conduct is translational (55%, 18/33), followed by patient oriented (39%, 13/33), basic science (27%, 9/33) and health services (21%, 7/33). Eighty-two percent (32/39) of graduates report program satisfaction scores of 4 or 5 out of 5.
Conclusion: The MD-PhD program at McGill has grown since its inception in 1985, producing a large proportion of physicians remaining in academic practice. Half of the alumni engaged in research pursue translational research.
Cannabinoids Activate AhR to Protect Against Lung Inflammation
Emily Wilson1,2, Hussein Trabulsi1, David Eidelman1,3, Carolyn Baglole1,2,3,41Research Institute of the McGill University Health Centre, Montréal, QC, Canada
2Department of Pharmacology and Therapeutics, McGill University, Montréal, QC, Canada
3Department of Medicine, McGill University, Montréal, QC, Canada
4Department of Pathology, McGill University, Montréal, QC, Canada.
Corresponding Author: Emily Wilson, email emilywilson460@gmail.com
Abstract
Background: In 2018 non-medical cannabis was legalized in Canada despite the unknown consequences of cannabis products on lung health. This is relevant as inhaling cannabis smoke is the most common way cannabis is consumed. Tetrahydrocannabinol (THC) and cannabidiol (CBD) are the most well-known cannabinoids in Cannabis sativa and have anti-inflammatory properties. Cannabinoids classically interact with cannabinoid receptors CB1 and CB2. However, there is little expression of these receptors in human or mouse structural lung cells. Cannabinoids may modulate lung immunity through the aryl hydrocarbon receptor (AhR); the activation of which is protective against lung inflammation from cigarette smoke. Through our preclinical models we assess the AhR-dependent effects of cannabis products on lung inflammation.
Hypothesis: Cannabinoids protect against lung inflammation via AhR activation.
Methods: Ahr+/- mice and Ahr-/- mice were exposed to cannabis smoke from high THC and high CBD cannabis strains. Inhalation exposures were be done using the SCIREQ® inExposeTM system equipped with a single cigarette chamber. Pulmonary immune cell infiltration, cytokines, lung mRNA and protein were analyzed for inflammatory markers.
Methodology: Smoke from high THC and high CBD cannabis induces AhR activation. High CBD cannabis smoke activates the AhR significantly more than high THC cannabis smoke. High THC cannabis smoke induces neutrophil infiltration to the lungs in an AhR-dependent manner. Smoke from both cannabis strains alters inflammatory markers compared to air-exposed control mice.
Conclusions: In vivo cannabis smoke exposures implicates the potential for C. sativa, especially high CBD strains, to protect against lung inflammation through the AhR.
Combining ATR inhibition with Carboplatin in chemoresistant TNBC conditionally reprogrammed cells and patient-derived xenograft
Juliet Guay1, Catherine Chabot1, Cédric Darini1, Marguerite Buchanan1, Adriana Aguilar-Mahecha1, Tim Kong2, Connie Yang2, Sidong Huang2, Mark Basik11Lady Davis Institute, Montréal, QC, Canada
2Rosalind & Morris Goodman Cancer Research Centre, Montréal, QC, Canada
Corresponding Author: Juliet Guay, email juliet.guay@mail.mcgill.ca
Abstract
Triple negative breast cancer patients mostly rely on chemotherapy1. In addition to the standard of care, they are often prescribed the DNA crosslinking agent carboplatin. Unfortunately, the majority are resistant or develop resistance to chemotherapy. Overcoming chemotherapy resistance in TNBC is a major clinical unmet need2.
We used conditional reprogramming to establish five patient-derived TNBC cell lines (CRCs) from patient-derived xenografts (PDXs) generated from chemotherapy resistant tumors. All cell lines were resistant to carboplatin and we performed shRNA high throughput screens to identify genetic vulnerabilities that could resensitize these cells to Carboplatin. We identified ataxia telangiectasia and Rad3-related protein (ATR) as a target.
We used AZD6738 and BAY1895344 to pharmacologically validate this hit. Combination with BAY1895344 was more synergistic than the combination with AZD6738. We observed a loss of ATR protein by western blot when the cells were exposed to the combination with BAY1895344. Pharmacological inhibition or knockdown of Chk1 did not recapitulate the synergy observed with BAY1895344-Carboplatin, suggesting the synergy observed is independent of Chk1 activity. Finally, our in vivo results demonstrated that BAY1895344-Carboplatin combination significantly delayed tumor growth compared to carboplatin treatment alone and prolonged survival in animal models.
We highlighted the efficacy of ATR inhibition to resensitize TNBC to carboplatin, both in vitro and in vivo. Our data shed light on a potentially novel mechanism of action of the BAY1895344 drug involving ATR expression. These studies highlight the relevance of primary tumor cells derived from freshly biobanked tumor tissues in providing novel therapeutic opportunities in breast cancer.
Molecular Mechanism of Calcium-Activated Potassium Channel Activation
Segura Emilie1,2, Marsolais Mireille2, Sauvé Rémy2, Parent Lucie1,21Institut de Cardiologie de Montréal, Montréal, QC, Canada
2Département de pharmacologie et physiologie, Université de Montréal, QC, Canada
Corresponding Author: Emilie Segura, email emilie.segura@umontreal.ca
Abstract
Small-conductance calcium-activated potassium channels SK4 belong to the family of 6-TM ion channels. Ca2+ ions near the channel pore open SK4 via constitutively-bound calmodulin (CaM). Their high submicromolar Ca2+ sensitivity makes SK4 a potential pharmacological target for the treatment of heart failure. 3D structures from cryo-EM show that SK4 and CaM adopt a tetrameric topology in a 4:4 ratio. C-terminal cytoplasmic helices HA and HB in SK4 anchor the C-lobe of CaM in the channel closed state. Activation proceeds when the free hanging N-lobe of CaM moves toward the S45A helix of the opposite subunit of SK4. Analysis of the structural model 6CNO.pdb suggests that the interaction between SK4 HA helice residues and CaM C-lobe residues that could underlie constitutive CaM binding. We carried biochemical and path-clamp assays to test this hypothesis. Co-expression of SK4 mutant with CaM mutant impaired the co-immunoprecipitation of SK4 with CaM and led to a significate decrease (p < 0.001) in the peak current densities when compared to the co-expression of both wild type proteins (41 ± 14 pA/pF, n=5). Noteworthy, this effect required the co-expression of both substituted proteins. We are currently investigating the role of a neighboring salt bridge between SK4 and CaM. Altogether, our results identified a hotspot for channel activity that we will exploit in the pharmacological control of SK4 in heart failure.
Patient-Specific Differences in Response to Doxorubicin-Induced Injury in iPSC-Derived Cardiomyocytes
Ida Derish1, Jeremy Zwaig1, Kashif Khan1, Renzo Cecere1,21Department of Experimental Surgery, McGill University, Montréal, QC, Canada
2Department of Cardiac Surgery, McGill University, Montréal, QC, Canada
Corresponding Author: Ida Derish, email ida.derish@mail.mcgill.ca
Abstract
Heart failure (HF) is an irreversible cardiovascular disease which remains a major cause of death and disability worldwide, as well as burden on the healthcare system. Terminally differentiated cardiomyocytes (CMs) are unable to proliferate and repair the heart tissue, once it has been damaged. Unfortunately, pharmacological development yields high rates of unforeseen side effects, notably cardiotoxicity and great variability between patient reactions, due to a lack of a representative heart model which could account for the diversity in a population. Doxorubicin (DOX) is a widely prescribed drug for the treatment of solid and haematologic malignancies, which causes cumulative and dose-dependent cardiotoxicity, to varying degrees between patients. Our preliminary study explores the usage of induced pluripotent stem cell (iPSC)-derived heart cells as a model to monitor patient differences in response to DOX injury. We generated iPSC-CMs from patient's blood (n=3 patients with cardiomyopathies, n=2 healthy donors), and we will treat iPSC-CMs with identical doses of DOX. Finally, we will monitor their response to the drug via changes in 1) metabolic activity, 2) beating rate and 3) viability. Since the generated CMs all contain the patient’s genetic code, these cells should hypothetically reflect their donor’s distinct cardiovascular cell reactions to therapies outside the patient’s body. As such, this study provides a glance into the predictive quality of utilizing patient-specific in vitro heart models as a tool to screen for personalized therapies. This highlights the necessity to uphold a higher standard of care for patients with a propensity for HF.
Integration of smoking cessation by telephone counseling into the McGill Lung Cancer Screening Trial
Ankita Ghatak1, Nicole Ezer2,31Department of Experimental Medicine, McGill University, Montréal, QC, Canada
2Department of Medicine, Faculty of Medicine and Health Sciences, McGill University, Montréal, QC, Canada
3Department of Medicine, Division of Respiratory Medicine, McGill University Health Centre, Montréal, QC, Canada
Corresponding Author: Ankita Ghatak, email ankita.ghatak@mail.mcgill.ca
Abstract
It has been widely established that the greatest risk factor for lung cancer is smoking and subsequently, smoking cessation clearly and significantly reduces lung cancer risk, and when combined with lung cancer screening, provides a significant mortality benefit. How best to implement smoking cessation interventions within lung cancer screening is not known, and what the impact is of phone counselling for smoking cessation in lung cancer screening populations is unclear.
This study compares the effectiveness of telephone counselling for smoking cessation in lung cancer screening eligible participants from the McGill Lung Cancer Screening Trial, as compared to healthcare worker referred participants and self-referred participants contacted by the Ligne J’arrete.
We performed a multivariate logistic regression to study the effect of referral group on 6 month quit rates, adjusted for variables such as age, gender, education, time to first cigarette, baseline cigarette use per day. Results showed a six month quit rate of 11% amongst lung cancer referred patients – significantly lower than other referral groups and comparable to similar studies with no intervention. Our model showed that lung cancer screening referred participants were significantly less likely to quit than healthcare worker referred and self-referred participants, even after adjustment.
Our study indicates that smoking cessation interventions by phone counselling may have poor efficacy and that contrary to current practices, lung cancer screening referred populations differ significantly from healthcare worker referred populations and suggests that smoking cessation services and lung cancer screening programs accommodate this difference in an effort to increase quit rates.
Extracellular Vesicles: A Potential Window into the Etiology of Major Depressive Disorder
Pascal Ibrahim1,2, Prakroothi Danthi2, Jean-Francois Theroux2, Corina Nagy2,3, Gustavo Turecki2,31Integrated Program in Neuroscience, McGill University, Montréal, QC, Canada
2McGill Group for Suicide Studies, Douglas Mental Health University Institute, Verdun, QC, Canada
3Department of Psychiatry, McGill University, Montréal, QC, Canada
Corresponding Author: Pascal Ibrahim, email pascal.ibrahim@mail.mcgill.ca
Abstract
Background: Major Depressive Disorder (MDD) is one of the leading causes of disability worldwide. Environmental factors are thought to play a role in disease development via epigenetic mechanisms. MicroRNA’s (miRNA) are well known epigenetic regulators that are disrupted in MDD and are packaged into extracellular vesicles (EVs). EVs have emerged as means of intercellular communication. They are thought to transfer miRNA and other molecules, such as proteins, between cells, altering gene expression in recipients. Therefore, we hypothesize that EV cargo from the anterior cingulate cortex will have a disease specific profile that could mediate disease development in MDD subjects compared to healthy controls.
Methods: EVs were isolated from post-mortem human brain tissue from the anterior cingulate cortex of using size exclusion chromatography. RNA was extracted and a small-RNA library was constructed and sequenced using the Illumina Platform. Proteins were also extracted and profiled using LC-MS/MS. Differential expression analysis was then performed.
Methodology: Western blots showed little to no contamination with cellular debris, along with enrichment of the exosomal marker CD9. TEM images showed the typical cup-shaped morphology with sizes mostly between 30 and 200 nm. Preliminary differential analyses revealed that both the miRNA and proteomic profiles of the EVs are dysregulated in MDD.
Conclusion: This will be the first study to profile brain-derived EV miRNA and protein in the context of depression. This could provide novel mechanistic insights into the pathophysiology of MDD, which could be a starting point for the development of targeted therapeutic strategies and prevention measures.
iPSC Secretome in the Rescue of Cardiomyocytes After Doxorubicin-Induced Injury
Zwaig J1, Derish I1, Khan K1, Cecere R1,21Department of Experimental Surgery, Faculty of Medicine, McGill University, Montréal, QC, Canada
2Department of Cardiac Surgery, Faculty of Medicine, McGill University, Montréal, QC, Canada
Corresponding Author: Jeremy Zwaig, email jeremy.zwaig@mail.mcgill.ca
Abstract
Myocardial infarction (MI) often leads to heart failure or death due to the cardiac tissue’s inability to regenerate after injury. Many attempts have been made to implant stem cells into infarcted hearts to induce recovery, yielding insignificant results. Now, the focus has shifted towards cell-free therapies to induce cardiac repair after MI. This study aims to evaluate the secretome of induced pluripotent stem cells (SiPSCs) generated from heart failure patients and healthy individuals as a potential treatment for doxorubicin (DOX)-injured primary cardiomyocytes (CMs). We hypothesize that the SiPSCs from both diseased patients (SiPSC-DP) and healthy controls (SiPSC-HC) will significantly increase CM metabolic activity and viability and decrease CM hypertrophy after DOX-induced injury. CD34+ cells were isolated from diseased patients (n=3) and a control patient’s (n=2) blood, reprogrammed into iPSCs, and secretome was collected. CMs were treated with DOX, which mimics a heart failure phenotype in CMs by significantly increasing cellular hypertrophy and cell death and decreasing metabolic activity. DOX-injured CMs were treated with i) SiPSC-DP, ii) SiPSC-HC, or iii) mesenchymal stem cell secretome. AlamarBlue, CrystalViolet, and actin immunostaining assays were performed to assess changes in CM metabolism, viability, and hypertrophy, after treatment with secretome. We will elucidate whether stem cells generated from various sources make effective secretome to treat cardiac tissue after injury, which is a step towards using secretome as a cardiac repair therapy. If SiPSC-DP is as effective as SiPSC-HC, then our future direction will be to evaluate the effect of patient-specific iPSC secretome on DOX-injured iPSC-derived cardiomyocytes.
Gene tests are not one-size-fits-all: Direct-to-consumer genetic tests and the need for patient recruitment ethics
Cassandra Haley11Centre of Genomics and Policy, McGill University, Montreal, QC, Canada
Corresponding Author: Cassandra Haley, email cassandra.haley@mail.mcgill.ca
Abstract
Direct-to-consumer (DTC) genetic tests, a multi-million-dollar industry, raise many concerns of scientific validity and clinical utility because DTC tests algorithms are designed and maintained by internal, often nondiverse, datasets. Therefore, there exists a disparity in the return of results to people of colour that derives from the nondiverse datasets used to create the proprietary DTC test algorithms. This directly results from the systematic underrepresentation of people of colour in genome-wide association studies. In this paper, I argue that patient recruitment ethics must be applied to DTC test development due to the diversity of customers purchasing tests, and the ethical imperative for fair subject selection in research. Maintaining a diversity of sampling is standard practice for ethical research; this application of the principle of justice is mentioned in seminal research ethics policy documents including the Belmont report and the Declaration of Helsinki. Many within the field of genetics have noted that the genome-wide association studies, which inform polygenic risk scores and DTC test results, lack diversity in sample data. As well, it is well known that there are measurable differences in the genetic landscape for a variety of diseases DTC tests purport to cover, including cancers, sickle-cell disease, and Alzheimer’s disease. Therefore, there is a need to increase the external validity of DTC tests to diverse customer populations. In order to increase diversity in research, researchers must consider the nuanced reasons behind minority underrepresentation, including structural barriers and cultural hesitance, and foster community-based participatory research to grow trust with marginalized populations.
Targeting the pro-metastatic function of CAFs in breast cancer: the promise of inhibiting the MNK1/2-eIF4E signaling axis
Julian Smith-Voudouris1, Margarita Bartish1, Samuel Preston1, Wilson H. Miller1,2, Sonia del Rincon1,21Division of Experimental Medicine, McGill University, Montréal, QC, Canada
2Department of Oncology, Segal Cancer Centre, Lady Davis Institute, Jewish General Hospital, McGill University, Montréal, QC, Canada
Corresponding Author: Julian Smith-Voudouris, email julian.smith-voudouris@mail.mcgill.ca
Abstract
Background: Dysfunctional translational control, specifically involving the MNK1/2-eIF4E axis, is common in breast cancer. Phosphorylation of eIF4E on serine 209 by MNK1/2 in malignant cells causes increased translation of a subset of pro-survival and invasion mRNAs. However, the specific role of phospho-eIF4E in non-malignant cells, such as cancer-associated-fibroblasts (CAFs) is unknown. My laboratory has shown through preliminary experiments that phospho-eIF4E deficient mammary gland fibroblasts repress 4T1 cell mediated lung metastasis independent of primary tumor growth. Yet, which fibroblast functions are modulated by phospho-eIF4E remains unknown. This study aims to uncover this by providing a phenotypic comparison of wild-type (WT) vs. phospho-eIF4E-deficient mammary gland fibroblasts.
Methods: Primary fibroblasts were harvested from the mammary glands of WT and phospho-eIF4E-deficient female mice and plated on cell culture dishes. A variety of in vitro analyses were used to compare well studied fibroblast properties and functions such as proliferation, migration, contractility, and promotion of invasion. Initial characterizations were performed on non-manipulated “baseline” fibroblasts.
Methodology: There are a similar number of fibroblasts present in the mammary glands of wild-type and phospho-eIF4E-deficient mice. In vitro analysis of baseline fibroblasts showed that phospho-eIF4E does not play a significant role in this state. Fibroblasts were shown to proliferate, migrate, contract collagen gel, and induce 4T1 cell invasion similarly regardless of phospho-eIF4E status.
Conclusion: We have shown that phospho-eIF4E does not play a role in the normal functions of mammary gland fibroblasts in a baseline state. Future work aims to explore potential differences once fibroblasts adopt a CAF phenotype.
Investigating Epigenetic Programs Promoting Triple-Negative Breast Cancer Metastasis
Xiaoting You1, Josie Ursini-Siegel1, Michael Witcher11Departments of Oncology and Experimental Medicine, The Lady Davis Institute of the Jewish General Hospital, McGill University, Montréal, QC, Canada
Corresponding Author: Xiaoting You, email xiaoting.you@mail.mcgill.ca
Abstract
Background: Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype, with high relapse rate and early metastatic events. Recent evidence suggests that epigenetic mechanisms play a central role in promoting TNBC metastasis. In a previous study from the Ursini-Siegel lab, overexpression of the oncogenic signaling adapter protein p66ShcA was shown to be a requirement for 4T1, a TNBC cell line, to metastasize toward the liver and lung. However, p66ShcA overexpression alone is not sufficient to trigger metastatic events, suggesting the cooperation of additional pro-metastatic pathways.
Methods: 4T1 TNBC cells, derived from the primary tumor, or from derivative lines that metastasize specifically to the liver, lung or bone are used in this project. We propose that comparing transcriptomic and epigenetic programming between the metastatic and primary cell lines will reveal pro-metastatic mechanisms. We have carried out RNA-sequencing on the cell lines, revealing differentially expressed targets and pathways. ChIP-qPCR was performed to profile histone modifications at the p66shcA gene to discern mechanisms that might lead to its overexpression. We have also submitted DNA-methylation samples for microarray.
Results: RNA-sequencing revealed repression of pro-inflammation and cell adhesion pathways in the metastatic cell lines. We propose that epigenetic therapies with a "Sting agonist", to promote inflammation, will work in synergy to reduce metastasis. ChIP-qPCR revealed that key activating marks (H3K27ac, H3K4me1), repressive mark H3K27me3 and H3K4me3 may be important in regulating metastasis.
Conclusion: This study provides insights regarding the pro-metastatic mechanisms in TNBC. We will further investigate treatment options targeting metastasis.
The Role of Sex Hormones in Plaque Instability in Men and Women with Severe Carotid Atherosclerosis
Diana Di Iorio, B.Sc1, Karina Gasbarrino, PhD1, Huaien Zheng, PhD1, Stella S. Daskalopoulou, MD, PhD11Vascular Health Unit, Research Institute of McGill University Health Centre, Department of Medicine, Faculty of Medicine, McGill University, Montréal, QC, Canada
Corresponding Author: Diana Di Iorio, email diana.diiorio2@mail.mcgill.ca
Abstract
Background: Carotid atherosclerotic plaques can be stable or unstable; the latter more likely to rupture resulting in strokes. Men develop more unstable carotid plaques than women, yet women have increased mortality rates post-stroke. Sex hormones influence the vasculature differently in men and women; until menopause estrogen protects women against cardiovascular disease. We hypothesize that sex hormones and their receptors may affect plaque instability.
Methods:
uid chromatography mass spectrometry, circulating testosterone, estradiol, androstenedione, and dehydroepiandrosterone were measured in blood samples from patients undergoing a carotid endarterectomy. Plaques were classified into 4 groups (n=20/group): women stable/unstable and men stable/unstable. Immunohistochemistry was performed on plaques to quantify expression of estrogen receptor alpha (ER-a), estrogen receptor beta (ER-b), G protein-coupled estrogen receptor, and androgen receptor (AR). Receptor mRNA expression was assessed in the plaque and in an in vitro model of in-plaque inflammation using qRT-PCR.
Methodology: There were no differences in circulating sex hormone levels between patient groups. However, men had significantly greater ER-a, ER-b, and AR expression in unstable vs stable plaques (p<0.05) and compared to women (p<0.05). Unstable plaques in men demonstrated less AR mRNA expression compared to stable plaques (p<0.05) and to women irrespective of plaque type (p<0.05). AR mRNA expression decreased significantly (p<0.05) upon differentiation of monocytes to macrophages, and macrophages to foam cells.
Conclusion: Our preliminary findings indicate a possible association between sex hormone receptor expression and plaque instability. With further investigations, our ongoing work may lead to hormone-specific therapies aimed at stabilizing plaques and reducing the incidence of stroke.
Determining mechanisms of resistance to Eribulin in TNBC using novel patient-derived models
Kathryn Bozek1,2, Marguerite Buchanan1, Cathy Lan1, Cédric Darini1, Adriana Aguilar1, Mark Basik1,2,3,41Cancer Genomics and Translational Research Laboratory, Lady Davis Institute/Segal Cancer Centre, Montréal, QC, Canada
2Division of Experimental Medicine, McGill University, Montréal, QC, Canada
3Departments of Medicine, McGill University, Montréal, QC, Canada
4Department of Oncology, McGill University, Montréal, QC, Canada
Corresponding Author: Katie Bozek, email kathryn.bozek@mail.mcgill.ca
Abstract
Eribulin is a chemotherapeutic agent approved for patients with advanced or metastatic breast cancer who have received taxane- and anthracycline-based chemotherapy. Unfortunately, many patients do not respond initially or inevitably develop resistance to Eribulin. We hypothesize that we can determine mechanisms of resistance and identify potential therapeutic targets for Eribulin-resistant Triple-Negative Breast Cancer (TNBC) using patient-derived xenografts (PDXs). We have developed 6 models of acquired-resistant TNBC via the treatment of PDX-bearing NSG mice with Eribulin until the emergence of resistance. For one PDX, we have a matched patient recurrent tumour-derived PDX which became resistant to Eribulin after the patient received carbo/taxol. We also have 3 models of intrinsic resistance and several sensitive models. All models are undergoing RNA-Seq and Copy Number Analysis. By RNA-Seq, we observed the loss of expression of the lncRNA XIST, the main player in X-chromosome inactivation, in two acquired resistant models. Transcript analysis revealed that XIST exon4 expression was also lost in three of our acquired resistant models, suggesting a potential link between XIST inhibition and Eribulin resistance. We are generating conditionally reprogrammed cells for 3 of these matched sensitive-resistant PDXs for validation studies. In our resistant cells, we will re-express XIST via both genetic introduction of the lncRNA and by targeting XIST negative-regulators to determine if XIST re-expression re-sensitizes cells to Eribulin. We will likewise validate these results in the matched PDXs. This study should uncover mechanisms of resistance to Eribulin and has the potential to identify novel therapeutic avenues for patients with Eribulin-resistant TNBC.
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