Novel Cell Cycle Regulation in Breast Cancer Treatment Resistance

Classification of breast cancer relies on the presence or absence of estrogen receptor alpha (ERα) and progesterone receptor (PR) as well as the overexpression or amplification of the Her2/neu receptor. Targeted therapies against these proteins has increased the overall 5-year survival rate of breast cancer patients. However, a subset of breast cancer patients can acquire resistance or are initially unresponsive to these therapies. Understanding the molecular pathways that can cause resistance within the various types of breast cancer is of high priority. The cell cycle regulatory factor Speedy (Spy1) has been found to be upregulated in a variety of human cancers, including invasive mammary carcinomas, as well as being downstream of two important pathways in breast cancer initiation and progression; MAPK and c-Myc. My study sought to investigate the role of Spy1 downstream of ERα and to determine its role in regulating treatment response in the presence or absence of ERα. My work defines a novel positive feedback loop whereby Spy1 activates ERK1/2 in a MEK-independent fashion. This activation was further demonstrated to increase the ligand-independent activation of ERα, correlating with a decrease in tamoxifen sensitivity. We tested our findings using an in vivo zebrafish model, demonstrating elevated levels of Spy1 alter tamoxifen sensitivity. We further demonstrate significantly high levels of Spy1 within the triple negative group of breast cancers; which correlates with decreased sensitivity to chemotherapy as well as CDK inhibitor treatment. These data could define an efficient mechanism driving proliferation and resistance in select cancers and may represent a potent drugable target

Targeting the Novel Cell Cycle Regulator, Spy1, for Treatment of Medulloblastoma

Targeting the Novel Cell Cycle Regulator, Spy1, for Treatment of Medulloblastoma Philip Habashy, Rosa M. Ferraiuolo, Dorota Lubanska, Lisa A. Porter Medulloblastoma (MB) is the most common malignant pediatric brain tumour. It occurs in 16-25% of cases, with higher incidence in children between the ages of 1 to 9 years. Current standard of care includes combination radiation, surgery and chemotherapy, this treatment relies on DNA damage to induce death of quickly growing cells. While effective for a small margin of patients the treatment is highly aggressive, is plagued with cytotoxicity and ultimately fails in many patients. One recent approach entering clinical development is the use of synthetic cyclin-dependent kinase inhibitors (CKIs). Finding new drugs and optimizing existing approaches for MB are of high importance. Our lab studies a cell cycle regulatory protein called Speedy (Spy1), which promotes cell proliferation, even during times of DNA damage produced by chemotherapeutic agents. Spy1 has been implicated in the maintenance and expansion of stem-like populations of tumour initiating cells known to be the most chemo-resistant among solid tumours. It is our hypothesis that Spy1 drives tumour initiating cells in MB and reducing the levels of Spy1 will increase sensitivity of aggressive MB to standard of care and CKI therapy. To address this hypothesis we have used patient-derived MB cells and have manipulated the levels of Spy1 using a lentiviral system. Using a high throughput platform these cells are injected into zebrafish prior to the establishment of the acquired immune system. We then determine the effect of CKI treatment on these in vivo tumours. To date our results show promise that this approach may sensitize, at least a subset of MB patients, to therapy. Our work may contribute toward optimizing the design of CKIs and the use in combination therapy. This project holds promise for improving survival and quality of life for MB patients

The cyclin-like protein Spy1/RINGO promotes mammary transformation and is elevated in human breast cancer

Abstract Background Spy1 is a novel \u27cyclin-like\u27 activator of the G1/S transition capable of enhancing cell proliferation as well as inhibiting apoptosis. Spy1 protein levels are tightly regulated during normal mammary development and forced overexpression in mammary mouse models accelerates mammary tumorigenesis. Methods Using human tissue samples, cell culture models and in vivo analysis we study the implications of Spy1 as a mediator of mammary transformation and breast cancer proliferation. Results We demonstrate that this protein can facilitate transformation in a manner dependent upon the activation of the G2/M Cdk, Cdk1, and the subsequent inhibition of the anti-apoptotic regulator FOXO1. Importantly, we show for the first time that enhanced levels of Spy1 protein are found in a large number of human breast cancers and that knockdown of Spy1 impairs breast cancer cell proliferation. Conclusions Collectively, this work supports that Spy1 is a unique activator of Cdk1 in breast cancer cells and may represent a valuable drug target and/or a prognostic marker for subsets of breast cancers

Differential expression of glucose transporters and hexokinases in prostate cancer with a neuroendocrine gene signature: A mechanistic perspective for 18 F-FDG imaging of PSMA-suppressed tumors

Although the incidence of de novo neuroendocrine prostate cancer (PC) is rare, recent data suggest that low expression of prostatespecific membrane antigen (PSMA) is associated with a spectrum of neuroendocrine hallmarks and androgen receptor (AR) suppression in PC. Previous clinical reports indicate that PCs with a phenotype similar to neuroendocrine tumors can be more amenable to imaging by 18F-FDG than by PSMA-targeting radioligands. In this study, we evaluated the association between neuroendocrine gene signature and 18F-FDG uptake-associated genes including glucose transporters (GLUTs) and hexokinases, with the goal of providing a genomic signature to explain the reported 18F-FDG avidity of PSMA suppressed tumors. Methods: Data-mining approaches, cell lines, and patient-derived xenograft models were used to study the levels of 14 members of the SLC2A family (encoding GLUT proteins), 4 members of the hexokinase family (genes HK1-HK3 and GCK), and PSMA (FOLH1 gene) after AR inhibition and in correlation with neuroendocrine hallmarks. Also, we characterize a neuroendocrine-like PC (NELPC) subset among a cohort of primary and metastatic PC samples with no neuroendocrine histopathology. We measured glucose uptake in a neuroendocrine-induced in vitro model and a zebrafish model by nonradioactive imaging of glucose uptake using a fluorescent glucose bioprobe, GB2-Cy3. Results: This work demonstrated that a neuroendocrine gene signature associates with differential expression of genes encoding GLUT and hexokinase proteins. In NELPC, elevated expression of GCK (encoding glucokinase protein) and decreased expression of SLC2A12 correlated with earlier biochemical recurrence. In tumors treated with AR inhibitors, high expression of GCK and low expression of SLC2A12 correlated with neuroendocrine histopathology and PSMA gene suppression. GLUT12 suppression and upregulation of glucokinase were observed in neuroendocrine- induced PC cell lines and patient-derived xenograft models. A higher glucose uptake was confirmed in low-PSMA tumors using a GB2-Cy3 probe in a zebrafish model. Conclusion: A neuroendocrine gene signature in neuroendocrine PC and NELPC associates with a distinct transcriptional profile of GLUTs and hexokinases. PSMA suppression correlates with GLUT12 suppression and glucokinase upregulation. Alteration of 18F-FDG uptake-associated genes correlated positively with higher glucose uptake in AR- and PSMA-suppressed tumors. Zebrafish xenograft tumor models are an accurate and efficient preclinical method for monitoring nonradioactive glucose uptake

The Multifaceted Roles of the Tumor Susceptibility Gene 101 (TSG101) in Normal Development and Disease

The multidomain protein encoded by the Tumor Susceptibility Gene 101 (TSG101) is ubiquitously expressed and is suggested to function in diverse intracellular processes. In this review, we provide a succinct overview of the main structural features of the protein and their suggested roles in molecular and cellular functions. We then summarize, in more detail, key findings from studies using genetically engineered animal models that demonstrate essential functions of TSG101 in cell proliferation and survival, normal tissue homeostasis, and tumorigenesis. Despite studies on cell lines that provide insight into the molecular underpinnings by which TSG101 might function as a negative growth regulator, a biologically significant role of TSG101 as a tumor suppressor has yet to be confirmed using genuine in vivo cancer models. More recent observations from several cancer research teams suggest that TSG101 might function as an oncoprotein. A potential role of post-translational mechanisms that control the expression of the TSG101 protein in cancer is being discussed. In the final section of the review, we summarize critical issues that need to be addressed to gain a better understanding of biologically significant roles of TSG101 in cancer

A retrospective single center study investigating the clinical significance of grade in triple negative breast cancer .

Background: Triple negative breast cancer (TNBC) is a heterogenous cancer type which lacks the receptors for estrogen (ER), progesterone (PR), and human epidermal growth factor receptor (HER-2) proteins. Comparatively, HER-2 positive cancers currently have a 7 year disease-free survival rate of 93% while in triple negative breast cancers, it can be as low as 77%. Purpose: While histological tumor grade or the degree of similarity to normal cells is an important prognostic factor (overall outcome), there is limited information on its predictive value (effect of specific therapeutic intervention). This project aimed to investigate the predictive value of grade in triple negative breast cancer for clinical decision making regarding treatment. Experimental Design: We reviewed 305 patient charts of triple negative breast cancer patients from 2004-2017 at Windsor Regional Cancer Center and the significance of grade with respect to oncological variables, survival-time, and time to relapse were explored. Results: The overall survival rates were 90.12%, 64.4%, and 77.2%, for grade 1, 2, 3 respectively. Comparing only between grade 2 and grade 3, we found that after five years, grade 2 patients had a 5.5-fold increased risk of death (HR = 5.5; 95% CI 1.2-25.6) and 2-folds higher risk of relapse (HR= 1.9; 95% CI 1.1-3.2). Grade 3 does significantly better than grade 2 in time to relapse with relapse rates of 70%, 55.6 %, and 75.6%, respectively for grades 1, 2, and 3 (P= 0.04). Conclusion: Grade can be shown to have positive predictive value in determining relapse with grade 2 showing poorest disease-free survival and faster time to relapse after the 5-year mark with implications in stratifying patients by grade in future clinical trials as further research elucidates more information about molecular differences between grades, as an explanation for these findings