Relevant Degree Programs
Affiliations to Research Centres, Institutes & Clusters
Graduate Student Supervision
Doctoral Student Supervision (Jan 2008 - May 2021)
While effective, resistance to 1st generation and 2nd generation androgen receptor (AR) pathway inhibitors is inevitable, creating a need to study the mechanisms by which prostate cancer (PCa) cells become resistant to these treatments. At its core, resistance can be categorized into AR driven and non-AR driven. The research presented in this thesis explores the hypothesis that non-AR driven resistance mechanisms exploit cellular plasticity to gain a survival advantage. Previous research has demonstrated that in response to 1st generation anti-androgens, epithelial PCa cells can undergo an epithelial-mesenchymal transition (EMT); a process implicated in metastatic dissemination of cancer cells throughout the body. Augmenting prior research demonstrating LYN tyrosine kinase’s role in resistance to 1st generation anti-androgens, our research demonstrates LYN promoting both EMT and metastasis. Specifically, we discovered that signaling cascade downstream of LYN alters the sub-cellular localization and therefore stability of key EMT promoting transcription factors Slug and Snail. While LYN does contribute to both treatment resistance and metastasis in PCa, its role in EMT was a pan-cancer mechanism observed in both breast and bladder cancer. Our research also explored the increased incidence of neuroendocrine PCa (NEPC) as a resistance mechanism to more potent 2nd generation anti-androgens (enzalutamide and abiraterone). Interrogating a model of in vivo derived enzalutamide resistance, our research uncovered the crucial role of neuronal transcription factor brain derived 2 (BRN2) in the transformation of epithelial prostate cells to neuroendocrine (NE) phenotype. Importantly, we discovered that AR inhibition relives its’ suppression of BRN2; thereby increasing BRN2 expression and initiating NE differentiation of epithelial PCa cells. Moreover, targeting BRN2 not only reduced the enzalutamide-induced NE differentiation but also reduced proliferation of enzalutamide resistant PCa cells and terminal NEPC cells. Building on this data, our research outlines the discovery of small molecule BRN2 inhibitors (BRN2i) that recapitulate the functional results demonstrated by conventional knockdown techniques. These BRN2i display specificity to BRN2 and selectivity in compromising growth of NEPC cells. Altogether, this thesis demonstrates two possible mechanisms how PCa cells alter their identity as epithelial cells to more mesenchymal (LYN) and/or neuroendocrine (BRN2) and gain resistance to AR pathway inhibitors.
Prostate cancer causes morbidity and mortality for thousands of Canadian men each year. Castration remains the primary treatment for recurrent or metastatic prostate cancer. However, castration is never curative, and the cancer inevitably recurs as castration resistant prostate cancer (CRPC). Newer androgen receptor (AR) antagonists such as enzalutamide(ENZ) demonstrate significant benefit in CRPC patients, but these agents remain non-curative. Therefore, the goal of this thesis was to explore novel strategies to target ENZ-resistant prostate cancer using previously developed models of resistance. Our models suggest that similar resistance patterns to CRPC may be found in ENZ-resistance, including the upregulation of steroidogenesis and activation of survival pathways such as the PI3K/Akt pathway. Unlike inhibition of the AR pathway, we found that inhibition of different nodes of the PI3K/Akt pathway had limited efficacy as monotherapy and we therefore focused on combination strategies to target this prominent survival pathway. We found that combined Akt and MEK pathway inhibition demonstrates only moderate synergy in AR-positive models of prostate cancer, and this did not appear significantly greater in ENZ-resistant than ENZ-sensitive prostate cancer. However, we did find that blockade of Akt signaling in combination with ENZ significantly delays the development of resistance to ENZ through a very significant induction of apoptosis and cell cycle arrest. Further, co-targeting of the Akt and AR pathways appears more effective at earlier stages of prostate cancer progression, when the tumours are still castrate-sensitive. Finally, to further evaluate the combination of PI3K/Akt pathway and AR blockade, we investigated the pre-clinical rationale for the use of bromodomain inhibitors, which indirectly targets both the AR and myc transcription factors. Upregulation of myc was observed following PI3K/Akt inhibition, but overall our studies did not support the combination of bromodomain inhibitors in combination with PI3K/Akt inhibition in prostate cancer models. Taken together, our pre-clinical results highlight several treatment strategies and pitfalls in targeting ENZ-resistant prostate cancer. These studies enhance our understanding of therapeutic approaches to target resistant prostate cancer with several novel agents and combination strategies. Further evaluation in clinical trials is warranted and ongoing.
Prostate Cancer (PCa) is the most common male cancer and the second leading cause ofcancer‐related deaths in North America. While many gains have been made in early detectionand treatment of localized PCa, many men still die of the metastatic disease. Androgen ablationtherapy remains the most effective therapy for patients with advanced disease. While ~80% ofpatients respond initially to this treatment, most patients progress to Castrate ResistantProstate Cancer (CRPC) stage. Current literature indicates that CRPC tumours are not uniformlyhormone refractory and may remain sensitive to therapies directed against the AR axis.Therefore, several new classes of AR‐targeting agents are now in clinical development,including more potent AR antagonists (MDV3100) and inhibitors of steroidogenesis(abiraterone). Although enthusiasm for this approach remains high, prostate tumourheterogeneity and the inevitable development of resistance dictates a critical need to betterunderstand the mechanisms of resistance in which AR remain active.In the current doctoral thesis role of heat shock protein (Hsp) 27 and Lyn tyrosine kinasein regulation of AR protein expression was investigated. The central hypothesis is that increasedexpression and activity of Hsp27 and Lyn kinase stabilizes and activates AR protein and leads toprostate cancer progression through promoting prostate cancer cell survival.Three specific objectives were accomplished in this thesis. First, the relationshipbetween Hsp27 and AR was studied. To this end, it was demonstrated that expression and activity of Hsp27 via a nongenomic mechanism regulates AR protein stability, shuttling andtranscriptional activity. In the next step, the underlying molecular mechanisms through whichLyn tyrosine kinase regulates AR activity in the castrated environment was investigated. Theexperiments demonstrated that Lyn kinase regulates AR protein expression and transcriptionalactivity and that it plays a key role in PCa progression to the castrated resistant stage. Finally, anovel role for Lyn kinase in Epidermal Growth Factor‐mediated (EGF‐mediated) AR activity wasdefined.The results obtained in this thesis defined new pathways involved in regulation of ARprotein stability and justified further investigation of Hsp27 and Lyn kinase as therapeutictargets for CRPC.
Master's Student Supervision (2010 - 2020)
Potent androgen receptor pathway inhibitors such as Enzalutamide (ENZ) and Abiraterone (Abi) have become the gold standard for patients with castration resistant prostate cancer (CRPC). However, treatment resistance is inevitable and all patients eventually become insensitive to these treatments. To investigate the molecular mechanism of treatment resistance in prostate cancer, our laboratory has engineered ENZ-resistant (ENZR) CRPC cell lines, which mirror clinical observations, through serial passaging of LNCaP xenografts under ENZ. Using our RNA sequencing data, we found Receptor tyrosine kinase like orphan receptor 2 (ROR2) as one of the most upregulated receptor tyrosine kinases (RTKs) in both our in vitro cell lines and patients. The oncogenic roles of ROR2 have been elucidated in various cancers, including prostate cancer. However, the role of ROR2 in context of treatment resistance in prostate cancer is still unknown. For the first time, we generated a novel ROR2 gene signature to provide insights on its role in treatment-resistant prostate cancer using RNA sequencing data of in vitro models and patients. We successfully validated the legitimacy of ROR2 gene signature in various models. Correlation studies revealed that ROR2 activity may be ligand-independent. Further examination of ROR2 gene signature revealed that ROR2 upregulates CD274 (known as programmed death-ligand 1; PD-L1) in treatment-resistant setting. Various computational studies and in vitro experiments supported and validated the novel ROR2-CD274 axis. Together, our data reveal a novel discovery of ROR2-CD274 axis in treatment-resistant prostate cancer.
Traf2- and Nck-interacting kinase (TNIK) is a serine/threonine kinase upregulated and amplified in pancreatic and gastric cancer respectively. TNIK has also been identified as a potential therapeutic target of colorectal cancer. However, the role of TNIK in prostate cancer (PCa) has not been investigated. Interrogating public human PCa patient data, we found that TNIK expression is associated with an aggressive form of PCa termed neuroendocrine prostate cancer (NEPC). Treatment-induced NEPC can arise as a consequence of strong selective pressure from androgen receptor (AR) pathway inhibition. Clinically, TNIK expression is positively correlated with neuroendocrine (NE) markers and inversely correlated with androgen regulated genes. In agreement, our in vitro studies reveal that TNIK expression is increased under AR pathway inhibition. We found that TNIK is transcriptionally repressed by androgen via direct binding of the AR at the TNIK locus. Through gain of function studies, we demonstrated that TNIK is not required for NE differentiation. Likewise, loss of function studies using siRNA or small molecule inhibitors targeting TNIK did not have significant effect on the growth of Enzalutamide-resistant cells with NE phenotype in vitro. Overall, our results indicate that TNIK may serve as a possible biomarker for NEPC.
Despite advances in therapeutics, castration resistant prostate cancer (CRPC) continues to be a major problem due to ongoing androgen receptor (AR) activity driving disease progression. This lack of control of AR inhibition in drug resistant CRPC may reflect a shift in the natural history of classically “AR-driven” disease to a “AR non-driven” phenotype, characterized by low circulating levels of prostate specific antigen (PSA) despite high metastatic burden in soft tissues. This hypothesis is supported by recent data from the Stand up to Cancer-Prostate Cancer Foundation East Coast and West Coast Dream Team (SU2C) indicating up to 27% of CRPC patients resistant to AR pathway inhibitors, including ENZ or ABI, develop an AR non-driven disease. Notably, our pre-clinical model of ENZ-R also reflects the clinical distribution of disease heterogeneity. We have shown that while 75% of ENZ-R tumors recur in vivo with AR re-activation and rising PSA, 25% of tumors show downregulation of canonical AR target genes and maintain an AR non-driven phenotype. Analysis of ENZ-R cells using RNAseq revealed that ENZ-R cells with downregulation of canonical pathway and NE phenotype up-regulate the paternally expressed gene 10 (PEG10). PEG10 is a retrotransposon-derived gene that was recently reported to be upregulated in neuroendocrine prostate cancer (NEPC) and regulates cell survival and metastasis in NEPC. However the underlying mechanisms by which PEG10 is regulated in ENZ-R and NEPC remain unexplored.We found that AR negatively regulates PEG10 and that activated AR directly binds to the PEG10 promoter and suppresses its expression while ENZ inhibits AR activity which allows for the NE differentiation to commence in CRPC and ENZ-R cell lines. In the next step, we investigated whether PEG10 is a potential target of NEPC progression in CRPC and ENZ-R cell lines. Through our experiments, we demonstrated that targeting PEG10 reduces cell proliferation in vitro, while knocking down PEG10 in vivo attenuates tumor growth. The results justify PEG10 as potential therapeutic target for NEPC.
Heat shock protein 27 (Hsp27) is a molecular chaperone highly and ubiquitously expressed in aggressive cancers where it controls a variety of pro-tumorigenic signaling pathways. Using gene expression profiling in prostate cancer cells with loss of Hsp27 function, we identified for the first time that Hsp27 regulates target genes in signaling pathways dependent on YAP and TAZ. Suppression of these transcriptional co-activators occurs via their phosphorylation and cytoplasmic retention by the Hippo tumor suppressor pathway. Mechanistic studies revealed that Hsp27 expression is associated with reduced YAP phosphorylation and enhanced transcription of YAP/TAZ target genes. Examination of the core components of the Hippo kinase cascade revealed that Hsp27 facilitates the proteasomal degradation of the core Hippo kinase, MST1, leading to reduced phosphorylation/activity of other main kinases responsible for YAP phosphorylation/inactivation, LATS1 and MOB1. Importantly, our data from cell lines was supported by data from human tumors; clinically, high expression of Hsp27 correlates with increased expression of YAP target genes in prostate cancer as well as reduced phosphorylation of YAP in lung and invasive breast cancer clinical samples. Together, our data reveal a novel mechanism by which Hsp27 regulates the Hippo tumor suppressor pathway, providing further rationale to target Hsp27 in multiple cancers.