Nicholas Bertos - Academia.edu (original) (raw)
Papers by Nicholas Bertos
PLOS Genetics, Jun 15, 2015
PubMed, 2023
Circulating tumor cells (CTCs) represent cells shed from the primary tumor or metastatic sites an... more Circulating tumor cells (CTCs) represent cells shed from the primary tumor or metastatic sites and can be used to monitor treatment response and tumor recurrence. However, CTCs circulate in extremely low numbers making in-depth analysis beyond simple enumeration challenging when collected from peripheral blood. Furthermore, tumor heterogeneity, a hallmark of many tumors, especially breast cancer, further complicates CTC characterization. To overcome this limitation, we developed a platform based on the large-scale isolation of CTCs by apheresis, allowing us to collect CTCs in large numbers, which were preserved live in liquid nitrogen for further characterization. Flow cytometry followed by cell sorting (FACS) was performed using a combination of antibodies directed against cell surface markers of white blood cells (CD45) and epithelial tumor cells (CK8). Analysis of subpopulations CD45+/- and CK8+/- by bulk RNA sequencing (RNAseq) and the CD45-/CK8 positive population by single-cell RNAseq was performed. The CD45- population was enriched using CD45 magnetic beads separation and examined by IHC for pan-cytokeratin and immunofluorescence (IF) for specific markers, including the elusive circulating cancer stem cells (CSCs). CSC-rich mammospheres were grown in vitro for further analysis and treated to examine their response to chemotherapeutic agents. Finally, mammospheres were transplanted into the mammary fat pad and bone of immunodeficient mice to examine tumor growth in vivo. This platform enables the detection and collection of CTCs in early and late-stage breast cancer patients of every subtype. Markers including CD44/24, ALDH1 and CXCR4 were identified by IF and showed high expression following mammosphere culture, which responded predictably to chemotherapeutic agents. Mammospheres were also transplanted into nude mice and induced tumors in the mammary fat pad and bone following intra-tibial transplantation. Finally, bulk RNA analysis of the FACS isolated CD45+/- and CK8+/- cells showed a clear separation of CD45- away from CD45+ populations. Single-cell RNAseq of the FACS isolated CD45-/CK8+ cells showed the presence of 4-5 clusters, confirming the high degree of heterogeneity of CTCs. Our platform for large-scale isolation of CTCs using apheresis is suitable for an in-depth analysis of the cancer phenotype and may eventually allow evaluation in real-time of the disease process to optimize cancer regimens.
ACS Biomaterials Science & Engineering, Jan 4, 2023
Methods in molecular biology, 2016
Laser capture microdissection (or LCM) allows for isolation of cells from specific tissue compart... more Laser capture microdissection (or LCM) allows for isolation of cells from specific tissue compartments, which can then be followed by DNA, RNA, and/or protein isolation and downstream characterization. Unlike other methods for cell isolation, LCM can be directed towards cells situated in specific anatomical contexts, and is therefore of significant value when investigating the tumor microenvironment, where localization is often key to function. Here, we present a summary of ways in which LCM can be utilized, as well as protocols for the isolation of tumor and tumor-associated stromal elements from frozen breast cancer samples, with a focus on preparation of samples for RNA characterization.
ACS Applied Materials & Interfaces, Jul 5, 2023
Journal of Clinical Oncology, Jun 1, 2023
Purpose: Although the murine orthologue of glycoprotein nonmetastatic B (GPNMB), Osteoactivin, pr... more Purpose: Although the murine orthologue of glycoprotein nonmetastatic B (GPNMB), Osteoactivin, promotes breast cancer metastasis in an in vivo mouse model, its importance in human breast cancer is unknown. We have examined the significance of GPNMB expression as a prognostic indicator of recurrence and assessed its potential as a novel therapeutic target in breast cancer. Experimental Design: The clinical significance of GPNMB expression in breast cancer was addressed by analyzing GPNMB levels in several published gene expression data sets and two independent tissue microarrays derived from human breast tumors. GPNMB-expressing human breast cancer cell lines were further used to validate a toxin-conjugated anti-GPNMB antibody as a novel therapeutic agent. Results: GPNMB expression correlates with shorter recurrence times and reduced overall survival of breast cancer patients. Epithelial-specific GPNMB staining is an independent prognostic indicator for breast cancer recurrence. GPNMB is highly expressed in basal and triple-negative breast cancers and is associated with increased risk of recurrence within this subtype. GPNMB expression confers a more migratory and invasive phenotype on breast cancer cells and sensitizes them to killing by CDX-011 (glembatumumab vedotin), a GPNMB-targeted antibody-drug conjugate. Conclusions: GPNMB expression is associated with the basal/triple-negative subtype and is a prognostic marker of poor outcome in patients with breast cancer. CDX-011 (glembatumumab vedotin) is a promising new targeted therapy for patients with metastatic triple-negative breast cancers, a patient population that currently lacks targeted-therapy options. Clin Cancer Res; 16(7); 2147-56. ©2010 AACR.
Canadian Journal of Surgery, Feb 15, 2023
Background: It has recently been reported that mismatch repair (MMR) status and microsatellite in... more Background: It has recently been reported that mismatch repair (MMR) status and microsatellite instability (MSI) status in gastroesophageal carcinomas predict surgical, chemotherapeutic and immunotherapeutic outcomes; however, there is extensive variability in the reported incidence and clinical implications of MMR/MSI status in gastroesophaegal adenocarcinomas. We characterized a Canadian surgical patient cohort with respect to MMR status, clinicopathologic correlates and anatomic tumour location. Methods: We investigated MMR and BRAF V600E status of gastroesophaegal adenocarcinomas in patients who underwent gastrectomy or esophagectomy with extended (D2) lymphadenectomy at a single centre between 2011 and 2019. We correlated patterns of MMR expression in the overall cohort and in anatomic location-defined subgroups with treatment response and overall survival using multivariate analysis. Results: In all, 226 cases of gastroesophaegal adenocarcinoma (63 esophageal, 98 gastroesophageal junctional and 65 gastric) were included. The MMR-deficient (dMMR) immunophenotype was found in 28 tumours (12.3%) (15 junctional [15.3%], 13 gastric [20.0%] and none of the esophageal). The majority (25 [89%]) of dMMR cases showed MLH1/PMS2 loss without concurrent BRAF V600E mutation. Two MSH2/ MSH6-deficient gastric tumours and 1 MSH6-deficient junctional tumour were detected. The pathologic response to preoperative chemotherapy was comparable in the dMMR and MMR-proficient (pMMR) cohorts. However, dMMR status was associated with significantly longer median overall survival than pMMR status (5.8 yr v. 2.4 yr, hazard ratio [HR] 1.91, 95% confidence interval [CI] 1.06-3.46), particularly in junctional tumours (4.6 yr v. 1.9 yr, HR 2.97, 95% CI 1.27-6.94). Conclusion: Our study shows that MMR status has at least prognostic value, which supports the need for biomarker testing in gastroesophageal adenocarcinomas, including junctional adenocarcinomas. This highlights the clinical significance of determining the MMR status in all adenocarcinomas of the upper gastrointestinal tract. Response to induction chemotherapy, however, was not influenced by MMR status. Contexte : Selon un récent rapport, le statut du système de réparation des mésappariements (MMR, pour mismatch repair) et de l'instabilité microsatellitaire (MSI, pour microsatellite instability) dans le cancer gastro-oesophagien permet de prédire l'issue des traitements (chirurgie, chimiothérapie et immunothérapie); or, on observe une importante variabilité quant à l'incidence rapportée et quant aux répercussions cliniques du statut MMR/MSI dans les adénocarcinomes gastro-oesophagiens. Auprès d'une cohorte de patients canadiens qui ont été opérés, nous avons recueilli le statut MMR, les corrélats clinicopathologiques et la localisation de la tumeur. Méthodes : Nous avons établi le statut MMR et BRAF V600E des adénocarcinomes gastro-oesophagiens chez des personnes ayant subi une gastrectomie ou une oesophagectomie avec lymphadénectomie étendue (D2) dans 1 seul centre entre 2011 et 2019. Nous avons établi la corrélation entre les modes d'expression du MMR dans la cohorte entière et dans des sous-groupes définis selon la localisation de la lésion, la réponse au traitement et la survie globale, à l'aide d'analyses multivariées. Résultats : En tout, nous avons inclus 226 cas d'adénocarcinomes gastro-oesophagiens (63 de l'oesophage, 98 de la jonction gastro-oesophagienne et 65 de l'estomac). L'immunophénotype MMR-déficient (dMMR) a été observé dans 28 tumeurs (12,3 %) (15 de la jonction [15,3 %], 13 de l'estomac [20,0 %] et aucun de l'oesophage). La majorité (25 [89 %]) des cas dMMR étaient négatifs pour MLH1/PMS2, sans mutation BRAF V600E concomitante. Deux tumeurs gastriques négatives pour MSH2/MSH6 et 1 tumeur de la jonction négative pour MSH6 ont été détectées. La réponse anatomopathologique à la chimiothérapie préopératoire a été comparable dans les cohortes
Canadian Journal of Surgery
Background: Procedural simulation has been shown to enhance early endoscopy training. In this pro... more Background: Procedural simulation has been shown to enhance early endoscopy training. In this proof of concept study, we aimed to show that a first-person shooter (FPS) video game with a novel in-house designed modified endoscope controller shares similar constructs with real-life endoscopy. Methods: A nonfunctioning colonoscope was fashioned to a wooden platform and suspended over a sensor connected to a computer. Customized software translated the colonoscope's movements into computer input. Participants completed the first three levels on an FPS video game, Portal (Valve Corporation), first using conventional mouse and keyboard controls and then using the novel endoscope controller. Twelve expert endoscopists and 12 surgical residents with minimal endoscopy experience participated. Participants were evaluated on the basis of completion time, number of button presses, and hand motion analyses. Results: Experts outperformed novices for time to study completion (expert 944 s, novice 1515 s; p = 0.006) and number of hand movements (expert 1263.1 s, novice 2052.6 s; p = 0.004) using the novel colonoscope controller. There was no difference in number of button presses or total path length travelled. Self-reported number of past endoscopies was moderately linearly correlated with time to game completion (r =-0.493, p = 0.020) and total hand movements (r =-0.462, p = 0.030). Novices and experts did not statistically differ while using the conventional video game controls. Conclusion: Experts outperformed novices using the endoscope controller but not the conventional game controller with respect to economy of movement and completion time. This result confirms that our endoscope-controlled video game shares similar constructs with real-life endoscopy and serves as a first step toward creating a more enjoyable and cheaper alternative to commercially available endoscopy simulators. 03 Is ethnicity an appropriate measure of health care marginalization?: A systematic review and metaanalysis of the outcomes of diabetic foot ulceration in the Aboriginal population. K. Hickey, D. Pace.
Annals of Oncology, May 1, 2017
Cancer Research, Apr 4, 2023
Traditionally, biobanking platforms have collected and stored non-viable biological specimens suc... more Traditionally, biobanking platforms have collected and stored non-viable biological specimens such as serum, plasma, and fresh-frozen or formalin-fixed paraffin-embedded (FFPE) tissues. These constitute key resources for clinical and contemporary genomics, transcriptomics, and proteomics studies. However, such specimens cannot be used for studies involving drug testing, high throughput target validation, and implementation for personalized medicine. Next-generation biobanking strategies rectify this issue by combining the collection of non-viable samples from patients with the propagation of viable tissue fractions as in vitro 2D and 3D cell models or in vivo xenograft models. Such cutting-edge biobanking strategies enable downstream cell-based high-throughput functional assays promoting the discovery of therapeutic targets or assessing treatment responses and resistance to treatment. Here, we provide comprehensive details of our establishment of a state-of-the-art biobanking platform for gastroesophageal adenocarcinoma (GEA) samples (n=389). Our approach opens new directions for understanding disease biology and conducting translational cancer studies. Patient-derived 2D cells (n=376), organoids (PDOs) (n=185), and xenografts (PDXs) (n= 99) included in our pipeline retain crucial features of the original human tumors, serving as valuable tools for clinical and experimental analyses in the context of precision oncology. We also discuss the entire approach of next-generation biobanking and emphasize the importance of integrating the propagation of PDOs and PDXs simultaneously. In addition, we explain each protocol optimized to propagate patient tissue-derived cell models. We validate that these models recapitulate tissue heterogeneity and are relevant preclinical models. Taken together, we described each approach used to develop one of the largest next-level biobanks for GEA. Citation Format: Mingyang Iris Kong, Sanjima Pal, Julie Berube, France Bourdeau, Betty Giannias, Nicholas Bertos, Veena Sangwan, Lorenzo Ferri. State-of-the-art biobanking of gastroesophageal adenocarcinoma samples: Integrating non-viable biospecimens with 3D and 2D cell models and their characterization, validation, and utilization of cell models for precision oncology. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5526.
In order to compare the performance of the High Pure FFPE RNA Micro Kit with that of two FFPE RNA... more In order to compare the performance of the High Pure FFPE RNA Micro Kit with that of two FFPE RNA isolation kits from other manufacturers, total RNA was isolated from sections of mouse mammary tumors with each of the three kits. Yield and integrity of the isolated RNA as well as genomic DNA (gDNA) contamination were assessed. Real-time RT-PCR of the housekeeping gene ACTB was performed to evaluate performance of the kits. In terms of yield and the A 260/280 and A 260/230 ratios, kits A and B seemed to perfom better than the High Pure FFPE RNA Micro Kit. In terms of real-time RT-PCR sensitivity, however, the High Pure FFPE RNA Micro Kit showed superior results.
Cancer Research, Feb 15, 2017
Triple negative breast cancer (TNBC), defined as tumors lacking expression of the estrogen recept... more Triple negative breast cancer (TNBC), defined as tumors lacking expression of the estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2), are especially difficult to treat effectively. While ER+ and HER2+ breast cancer subtypes can be treated with Tamoxifen and Herceptin, respectively, there are no targeted therapies for TNBC patients. Furthermore, while only 20-30% of TNBC patients respond to chemotherapy in the neoadjuvant setting, overall outcome remains poor for non-responding patients. However, mounting evidence suggests that immune-checkpoint inhibitor immunotherapies may be especially promising for TNBC patients. We and others have shown that the presence of CD8+ T cells, a crucial component of the cytotoxic arm of the adaptive immune response, is a sign of good clinical outcome in TNBC patients. However, good outcome only correlates with CD8 +T cell invasion of the tumor parenchyma. Some patients had an accumulation of CD8+ T cells in the surrounding tumor-associated stroma, but not the tumor epithelia, and these patients responded as poorly as patients with no CD8 T cells at all. Yet how cancer associated fibroblasts (CAFs), the dominant cell type of the tumor-associated stroma, affects CD8+ T cell invasion into the tumor epithelia is still poorly understood. To identify potential stroma-dependent mechanisms which potentiate or inhibit CD8+ T cells invasion into the tumor epithelia, we performed gene expression profiling of laser-capture microdissected tumor-associated stroma (and matched epithelia) from 38 TNBC cases. Here we identify several stromal and epithelial canonical pathways as well as biomarkers that are associated with and may explain the accumulation of CD8 T cells outside of the tumor epithelia. Citation Format: Gruosso T, Gigoux M, Bertos N, Zuo D, Manem V, Monette A, Lapointe R, Haibe-Kains B, Park M. Mechanisms of CD8+ T cell immunosuppression in triple negative breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P4-03-08.
Cancer Research
Gastroesophageal adenocarcinoma (GEA) is the fastest rising cancer in North America. Over the cou... more Gastroesophageal adenocarcinoma (GEA) is the fastest rising cancer in North America. Over the course of five years, the survival rate is <20%, creating an urgent need for appropriate treatments against GEA. Currently, providing patients with peri-operative systemic docetaxel triplet-based chemotherapy (DCF or FLOT) is the most effective approach to treat GEA. Despite this, for 50% of patients that do present an initial response to therapy, the tumor returns due to pre-existing or newly acquired resistance (i.e., chemoresistance) by the cancer. Researchers have shifted their focus to the tumor microenvironment (TME) as one of the factors influencing chemoresistance in patients. The TME is composed of tumor cells, immune cells and their secreted products, as well as fibroblasts. The components of the TME have been shown to interact with one another to influence tumor growth and progression. Fibroblasts are wound-healing cells that can be transformed into cancer-associated fibroblas...
PTEN loss-of-function contributes to hyperactivation of the PI3K pathway and to drug resistance i... more PTEN loss-of-function contributes to hyperactivation of the PI3K pathway and to drug resistance in breast cancer. Unchecked PI3K pathway signaling increases activation of the mechanistic target of rapamycin complex 1 (mTORC1), which promotes tumorigenicity. Several studies have suggested that vacuolar (H+)–ATPase (V–ATPase) complex activity is regulated by PI3K signaling. In this study, we showed that loss of PTEN elevated V–ATPase activity. Enhanced V–ATPase activity was mediated by increased expression of the ATPase H+ transporting accessory protein 2 (ATP6AP2), also known as the prorenin receptor (PRR). PRR is cleaved into a secreted extracellular fragment (sPRR) and an intracellular fragment (M8.9) that remains associated with the V–ATPase complex. Reduced PTEN expression increased V–ATPase complex activity in a PRR-dependent manner. Breast cancer cell lines with reduced PTEN expression demonstrated increased PRR expression. Similarly, PRR expression became elevated upon PTEN de...
This figure contains bright-field counterparts of LysoTracker images. This figure also contains H... more This figure contains bright-field counterparts of LysoTracker images. This figure also contains HER2 immunostaining after PRR knockdown in SKBR3 cells.
Cancer Research
Introduction: Esophageal cancer causes sixth most cancer-related morbidity and mortality worldwid... more Introduction: Esophageal cancer causes sixth most cancer-related morbidity and mortality worldwide, with a survival rate of <20%. Incidence of esophageal adenocarcinoma (EA) subtype has been rising (>60%) in North America. Systemic docetaxel-based triplet chemotherapy represents the best standard of care, approx. 60% of patients showcasing innate chemo-resistance or developing acquired chemo-resistance. 3D organoids provide a robust and heterogeneous cell source but lack the stromal microenvironment. We established a high-fidelity, stromal-inclusive tumor-on-chip platform expanding on tumor heterogeneity with micro-physiological relevance and flexible complexity. Methods: A total of 8 (4 chemo-sensitive and 4 chemo-resistant) treatment naïve patient-derived organoids (PDOs) and matched fibroblasts were selected to develop syngeneic human esophageal microtissues (tumor and adjacent) on commercially available (Emulate), compartmentalized, porous polydimethylsiloxane (PDMS) membr...
Clinical parameters of patients from whom tissue microarrays were acquired for P-Akt staining
PLOS Genetics, Jun 15, 2015
PubMed, 2023
Circulating tumor cells (CTCs) represent cells shed from the primary tumor or metastatic sites an... more Circulating tumor cells (CTCs) represent cells shed from the primary tumor or metastatic sites and can be used to monitor treatment response and tumor recurrence. However, CTCs circulate in extremely low numbers making in-depth analysis beyond simple enumeration challenging when collected from peripheral blood. Furthermore, tumor heterogeneity, a hallmark of many tumors, especially breast cancer, further complicates CTC characterization. To overcome this limitation, we developed a platform based on the large-scale isolation of CTCs by apheresis, allowing us to collect CTCs in large numbers, which were preserved live in liquid nitrogen for further characterization. Flow cytometry followed by cell sorting (FACS) was performed using a combination of antibodies directed against cell surface markers of white blood cells (CD45) and epithelial tumor cells (CK8). Analysis of subpopulations CD45+/- and CK8+/- by bulk RNA sequencing (RNAseq) and the CD45-/CK8 positive population by single-cell RNAseq was performed. The CD45- population was enriched using CD45 magnetic beads separation and examined by IHC for pan-cytokeratin and immunofluorescence (IF) for specific markers, including the elusive circulating cancer stem cells (CSCs). CSC-rich mammospheres were grown in vitro for further analysis and treated to examine their response to chemotherapeutic agents. Finally, mammospheres were transplanted into the mammary fat pad and bone of immunodeficient mice to examine tumor growth in vivo. This platform enables the detection and collection of CTCs in early and late-stage breast cancer patients of every subtype. Markers including CD44/24, ALDH1 and CXCR4 were identified by IF and showed high expression following mammosphere culture, which responded predictably to chemotherapeutic agents. Mammospheres were also transplanted into nude mice and induced tumors in the mammary fat pad and bone following intra-tibial transplantation. Finally, bulk RNA analysis of the FACS isolated CD45+/- and CK8+/- cells showed a clear separation of CD45- away from CD45+ populations. Single-cell RNAseq of the FACS isolated CD45-/CK8+ cells showed the presence of 4-5 clusters, confirming the high degree of heterogeneity of CTCs. Our platform for large-scale isolation of CTCs using apheresis is suitable for an in-depth analysis of the cancer phenotype and may eventually allow evaluation in real-time of the disease process to optimize cancer regimens.
ACS Biomaterials Science & Engineering, Jan 4, 2023
Methods in molecular biology, 2016
Laser capture microdissection (or LCM) allows for isolation of cells from specific tissue compart... more Laser capture microdissection (or LCM) allows for isolation of cells from specific tissue compartments, which can then be followed by DNA, RNA, and/or protein isolation and downstream characterization. Unlike other methods for cell isolation, LCM can be directed towards cells situated in specific anatomical contexts, and is therefore of significant value when investigating the tumor microenvironment, where localization is often key to function. Here, we present a summary of ways in which LCM can be utilized, as well as protocols for the isolation of tumor and tumor-associated stromal elements from frozen breast cancer samples, with a focus on preparation of samples for RNA characterization.
ACS Applied Materials & Interfaces, Jul 5, 2023
Journal of Clinical Oncology, Jun 1, 2023
Purpose: Although the murine orthologue of glycoprotein nonmetastatic B (GPNMB), Osteoactivin, pr... more Purpose: Although the murine orthologue of glycoprotein nonmetastatic B (GPNMB), Osteoactivin, promotes breast cancer metastasis in an in vivo mouse model, its importance in human breast cancer is unknown. We have examined the significance of GPNMB expression as a prognostic indicator of recurrence and assessed its potential as a novel therapeutic target in breast cancer. Experimental Design: The clinical significance of GPNMB expression in breast cancer was addressed by analyzing GPNMB levels in several published gene expression data sets and two independent tissue microarrays derived from human breast tumors. GPNMB-expressing human breast cancer cell lines were further used to validate a toxin-conjugated anti-GPNMB antibody as a novel therapeutic agent. Results: GPNMB expression correlates with shorter recurrence times and reduced overall survival of breast cancer patients. Epithelial-specific GPNMB staining is an independent prognostic indicator for breast cancer recurrence. GPNMB is highly expressed in basal and triple-negative breast cancers and is associated with increased risk of recurrence within this subtype. GPNMB expression confers a more migratory and invasive phenotype on breast cancer cells and sensitizes them to killing by CDX-011 (glembatumumab vedotin), a GPNMB-targeted antibody-drug conjugate. Conclusions: GPNMB expression is associated with the basal/triple-negative subtype and is a prognostic marker of poor outcome in patients with breast cancer. CDX-011 (glembatumumab vedotin) is a promising new targeted therapy for patients with metastatic triple-negative breast cancers, a patient population that currently lacks targeted-therapy options. Clin Cancer Res; 16(7); 2147-56. ©2010 AACR.
Canadian Journal of Surgery, Feb 15, 2023
Background: It has recently been reported that mismatch repair (MMR) status and microsatellite in... more Background: It has recently been reported that mismatch repair (MMR) status and microsatellite instability (MSI) status in gastroesophageal carcinomas predict surgical, chemotherapeutic and immunotherapeutic outcomes; however, there is extensive variability in the reported incidence and clinical implications of MMR/MSI status in gastroesophaegal adenocarcinomas. We characterized a Canadian surgical patient cohort with respect to MMR status, clinicopathologic correlates and anatomic tumour location. Methods: We investigated MMR and BRAF V600E status of gastroesophaegal adenocarcinomas in patients who underwent gastrectomy or esophagectomy with extended (D2) lymphadenectomy at a single centre between 2011 and 2019. We correlated patterns of MMR expression in the overall cohort and in anatomic location-defined subgroups with treatment response and overall survival using multivariate analysis. Results: In all, 226 cases of gastroesophaegal adenocarcinoma (63 esophageal, 98 gastroesophageal junctional and 65 gastric) were included. The MMR-deficient (dMMR) immunophenotype was found in 28 tumours (12.3%) (15 junctional [15.3%], 13 gastric [20.0%] and none of the esophageal). The majority (25 [89%]) of dMMR cases showed MLH1/PMS2 loss without concurrent BRAF V600E mutation. Two MSH2/ MSH6-deficient gastric tumours and 1 MSH6-deficient junctional tumour were detected. The pathologic response to preoperative chemotherapy was comparable in the dMMR and MMR-proficient (pMMR) cohorts. However, dMMR status was associated with significantly longer median overall survival than pMMR status (5.8 yr v. 2.4 yr, hazard ratio [HR] 1.91, 95% confidence interval [CI] 1.06-3.46), particularly in junctional tumours (4.6 yr v. 1.9 yr, HR 2.97, 95% CI 1.27-6.94). Conclusion: Our study shows that MMR status has at least prognostic value, which supports the need for biomarker testing in gastroesophageal adenocarcinomas, including junctional adenocarcinomas. This highlights the clinical significance of determining the MMR status in all adenocarcinomas of the upper gastrointestinal tract. Response to induction chemotherapy, however, was not influenced by MMR status. Contexte : Selon un récent rapport, le statut du système de réparation des mésappariements (MMR, pour mismatch repair) et de l'instabilité microsatellitaire (MSI, pour microsatellite instability) dans le cancer gastro-oesophagien permet de prédire l'issue des traitements (chirurgie, chimiothérapie et immunothérapie); or, on observe une importante variabilité quant à l'incidence rapportée et quant aux répercussions cliniques du statut MMR/MSI dans les adénocarcinomes gastro-oesophagiens. Auprès d'une cohorte de patients canadiens qui ont été opérés, nous avons recueilli le statut MMR, les corrélats clinicopathologiques et la localisation de la tumeur. Méthodes : Nous avons établi le statut MMR et BRAF V600E des adénocarcinomes gastro-oesophagiens chez des personnes ayant subi une gastrectomie ou une oesophagectomie avec lymphadénectomie étendue (D2) dans 1 seul centre entre 2011 et 2019. Nous avons établi la corrélation entre les modes d'expression du MMR dans la cohorte entière et dans des sous-groupes définis selon la localisation de la lésion, la réponse au traitement et la survie globale, à l'aide d'analyses multivariées. Résultats : En tout, nous avons inclus 226 cas d'adénocarcinomes gastro-oesophagiens (63 de l'oesophage, 98 de la jonction gastro-oesophagienne et 65 de l'estomac). L'immunophénotype MMR-déficient (dMMR) a été observé dans 28 tumeurs (12,3 %) (15 de la jonction [15,3 %], 13 de l'estomac [20,0 %] et aucun de l'oesophage). La majorité (25 [89 %]) des cas dMMR étaient négatifs pour MLH1/PMS2, sans mutation BRAF V600E concomitante. Deux tumeurs gastriques négatives pour MSH2/MSH6 et 1 tumeur de la jonction négative pour MSH6 ont été détectées. La réponse anatomopathologique à la chimiothérapie préopératoire a été comparable dans les cohortes
Canadian Journal of Surgery
Background: Procedural simulation has been shown to enhance early endoscopy training. In this pro... more Background: Procedural simulation has been shown to enhance early endoscopy training. In this proof of concept study, we aimed to show that a first-person shooter (FPS) video game with a novel in-house designed modified endoscope controller shares similar constructs with real-life endoscopy. Methods: A nonfunctioning colonoscope was fashioned to a wooden platform and suspended over a sensor connected to a computer. Customized software translated the colonoscope's movements into computer input. Participants completed the first three levels on an FPS video game, Portal (Valve Corporation), first using conventional mouse and keyboard controls and then using the novel endoscope controller. Twelve expert endoscopists and 12 surgical residents with minimal endoscopy experience participated. Participants were evaluated on the basis of completion time, number of button presses, and hand motion analyses. Results: Experts outperformed novices for time to study completion (expert 944 s, novice 1515 s; p = 0.006) and number of hand movements (expert 1263.1 s, novice 2052.6 s; p = 0.004) using the novel colonoscope controller. There was no difference in number of button presses or total path length travelled. Self-reported number of past endoscopies was moderately linearly correlated with time to game completion (r =-0.493, p = 0.020) and total hand movements (r =-0.462, p = 0.030). Novices and experts did not statistically differ while using the conventional video game controls. Conclusion: Experts outperformed novices using the endoscope controller but not the conventional game controller with respect to economy of movement and completion time. This result confirms that our endoscope-controlled video game shares similar constructs with real-life endoscopy and serves as a first step toward creating a more enjoyable and cheaper alternative to commercially available endoscopy simulators. 03 Is ethnicity an appropriate measure of health care marginalization?: A systematic review and metaanalysis of the outcomes of diabetic foot ulceration in the Aboriginal population. K. Hickey, D. Pace.
Annals of Oncology, May 1, 2017
Cancer Research, Apr 4, 2023
Traditionally, biobanking platforms have collected and stored non-viable biological specimens suc... more Traditionally, biobanking platforms have collected and stored non-viable biological specimens such as serum, plasma, and fresh-frozen or formalin-fixed paraffin-embedded (FFPE) tissues. These constitute key resources for clinical and contemporary genomics, transcriptomics, and proteomics studies. However, such specimens cannot be used for studies involving drug testing, high throughput target validation, and implementation for personalized medicine. Next-generation biobanking strategies rectify this issue by combining the collection of non-viable samples from patients with the propagation of viable tissue fractions as in vitro 2D and 3D cell models or in vivo xenograft models. Such cutting-edge biobanking strategies enable downstream cell-based high-throughput functional assays promoting the discovery of therapeutic targets or assessing treatment responses and resistance to treatment. Here, we provide comprehensive details of our establishment of a state-of-the-art biobanking platform for gastroesophageal adenocarcinoma (GEA) samples (n=389). Our approach opens new directions for understanding disease biology and conducting translational cancer studies. Patient-derived 2D cells (n=376), organoids (PDOs) (n=185), and xenografts (PDXs) (n= 99) included in our pipeline retain crucial features of the original human tumors, serving as valuable tools for clinical and experimental analyses in the context of precision oncology. We also discuss the entire approach of next-generation biobanking and emphasize the importance of integrating the propagation of PDOs and PDXs simultaneously. In addition, we explain each protocol optimized to propagate patient tissue-derived cell models. We validate that these models recapitulate tissue heterogeneity and are relevant preclinical models. Taken together, we described each approach used to develop one of the largest next-level biobanks for GEA. Citation Format: Mingyang Iris Kong, Sanjima Pal, Julie Berube, France Bourdeau, Betty Giannias, Nicholas Bertos, Veena Sangwan, Lorenzo Ferri. State-of-the-art biobanking of gastroesophageal adenocarcinoma samples: Integrating non-viable biospecimens with 3D and 2D cell models and their characterization, validation, and utilization of cell models for precision oncology. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5526.
In order to compare the performance of the High Pure FFPE RNA Micro Kit with that of two FFPE RNA... more In order to compare the performance of the High Pure FFPE RNA Micro Kit with that of two FFPE RNA isolation kits from other manufacturers, total RNA was isolated from sections of mouse mammary tumors with each of the three kits. Yield and integrity of the isolated RNA as well as genomic DNA (gDNA) contamination were assessed. Real-time RT-PCR of the housekeeping gene ACTB was performed to evaluate performance of the kits. In terms of yield and the A 260/280 and A 260/230 ratios, kits A and B seemed to perfom better than the High Pure FFPE RNA Micro Kit. In terms of real-time RT-PCR sensitivity, however, the High Pure FFPE RNA Micro Kit showed superior results.
Cancer Research, Feb 15, 2017
Triple negative breast cancer (TNBC), defined as tumors lacking expression of the estrogen recept... more Triple negative breast cancer (TNBC), defined as tumors lacking expression of the estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2), are especially difficult to treat effectively. While ER+ and HER2+ breast cancer subtypes can be treated with Tamoxifen and Herceptin, respectively, there are no targeted therapies for TNBC patients. Furthermore, while only 20-30% of TNBC patients respond to chemotherapy in the neoadjuvant setting, overall outcome remains poor for non-responding patients. However, mounting evidence suggests that immune-checkpoint inhibitor immunotherapies may be especially promising for TNBC patients. We and others have shown that the presence of CD8+ T cells, a crucial component of the cytotoxic arm of the adaptive immune response, is a sign of good clinical outcome in TNBC patients. However, good outcome only correlates with CD8 +T cell invasion of the tumor parenchyma. Some patients had an accumulation of CD8+ T cells in the surrounding tumor-associated stroma, but not the tumor epithelia, and these patients responded as poorly as patients with no CD8 T cells at all. Yet how cancer associated fibroblasts (CAFs), the dominant cell type of the tumor-associated stroma, affects CD8+ T cell invasion into the tumor epithelia is still poorly understood. To identify potential stroma-dependent mechanisms which potentiate or inhibit CD8+ T cells invasion into the tumor epithelia, we performed gene expression profiling of laser-capture microdissected tumor-associated stroma (and matched epithelia) from 38 TNBC cases. Here we identify several stromal and epithelial canonical pathways as well as biomarkers that are associated with and may explain the accumulation of CD8 T cells outside of the tumor epithelia. Citation Format: Gruosso T, Gigoux M, Bertos N, Zuo D, Manem V, Monette A, Lapointe R, Haibe-Kains B, Park M. Mechanisms of CD8+ T cell immunosuppression in triple negative breast cancer [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P4-03-08.
Cancer Research
Gastroesophageal adenocarcinoma (GEA) is the fastest rising cancer in North America. Over the cou... more Gastroesophageal adenocarcinoma (GEA) is the fastest rising cancer in North America. Over the course of five years, the survival rate is <20%, creating an urgent need for appropriate treatments against GEA. Currently, providing patients with peri-operative systemic docetaxel triplet-based chemotherapy (DCF or FLOT) is the most effective approach to treat GEA. Despite this, for 50% of patients that do present an initial response to therapy, the tumor returns due to pre-existing or newly acquired resistance (i.e., chemoresistance) by the cancer. Researchers have shifted their focus to the tumor microenvironment (TME) as one of the factors influencing chemoresistance in patients. The TME is composed of tumor cells, immune cells and their secreted products, as well as fibroblasts. The components of the TME have been shown to interact with one another to influence tumor growth and progression. Fibroblasts are wound-healing cells that can be transformed into cancer-associated fibroblas...
PTEN loss-of-function contributes to hyperactivation of the PI3K pathway and to drug resistance i... more PTEN loss-of-function contributes to hyperactivation of the PI3K pathway and to drug resistance in breast cancer. Unchecked PI3K pathway signaling increases activation of the mechanistic target of rapamycin complex 1 (mTORC1), which promotes tumorigenicity. Several studies have suggested that vacuolar (H+)–ATPase (V–ATPase) complex activity is regulated by PI3K signaling. In this study, we showed that loss of PTEN elevated V–ATPase activity. Enhanced V–ATPase activity was mediated by increased expression of the ATPase H+ transporting accessory protein 2 (ATP6AP2), also known as the prorenin receptor (PRR). PRR is cleaved into a secreted extracellular fragment (sPRR) and an intracellular fragment (M8.9) that remains associated with the V–ATPase complex. Reduced PTEN expression increased V–ATPase complex activity in a PRR-dependent manner. Breast cancer cell lines with reduced PTEN expression demonstrated increased PRR expression. Similarly, PRR expression became elevated upon PTEN de...
This figure contains bright-field counterparts of LysoTracker images. This figure also contains H... more This figure contains bright-field counterparts of LysoTracker images. This figure also contains HER2 immunostaining after PRR knockdown in SKBR3 cells.
Cancer Research
Introduction: Esophageal cancer causes sixth most cancer-related morbidity and mortality worldwid... more Introduction: Esophageal cancer causes sixth most cancer-related morbidity and mortality worldwide, with a survival rate of <20%. Incidence of esophageal adenocarcinoma (EA) subtype has been rising (>60%) in North America. Systemic docetaxel-based triplet chemotherapy represents the best standard of care, approx. 60% of patients showcasing innate chemo-resistance or developing acquired chemo-resistance. 3D organoids provide a robust and heterogeneous cell source but lack the stromal microenvironment. We established a high-fidelity, stromal-inclusive tumor-on-chip platform expanding on tumor heterogeneity with micro-physiological relevance and flexible complexity. Methods: A total of 8 (4 chemo-sensitive and 4 chemo-resistant) treatment naïve patient-derived organoids (PDOs) and matched fibroblasts were selected to develop syngeneic human esophageal microtissues (tumor and adjacent) on commercially available (Emulate), compartmentalized, porous polydimethylsiloxane (PDMS) membr...
Clinical parameters of patients from whom tissue microarrays were acquired for P-Akt staining