Investigating aberrant signalling in enzalutamide-resistant prostate cancer

PhD ThesisProstate cancer is usually androgen-dependent and consequently, initial therapy for many patients, particularly with advanced disease, is androgen withdrawal, via anti-androgen therapeutics. Most patients respond to anti-androgen therapy in the early stages of their disease but many will develop resistance, entering a “castrate-resistant” disease state. Enzalutamide and ARN-509 have shown promise in the treatment of castration resistant prostate cancer (CRPC) patients, however response rates are just 50% and there is the inevitable development of resistance and subsequent disease progression. The aims of this study are to investigate the role of HER2/HER3 in CRPC models (Casodex-, Enzalutamide- and ARN509-resistant cell lines) and the signalling pathway(s) that can be stimulated through HER2/HER3 activation in these models. In addition, the project focusses on drug-resistant disease models, investigating the genes upregulated in a cell-line model of enzalutamide-resistance. The data showed that HER2/HER3 has a crucial role in the CRPC model cell lines, seen in the activation of both MAP kinase and PI3K/Akt pathways, which are responsible for tumour growth and metastasis. This activity is more pronounced in enzalutamide resistant- LNCaP cells. For that reason, this study aimed to interrogate the global gene expression consequences in this enzalutamide resistant- LNCaP cell model. These aims were approached using Illumina Human HT-12 arrays to detect significantly up-regulated genes and therefore could have a vital role in proliferation, migration and cell cycle. SGK1 and TROP-2 were selected from this microarray to study in more details. The data showed an increase in the expression of SGK1 in Casodex-, enzalutamide- and ARN509-resistant cell lines, compared with parental LNCaP cells. AR regulates SGK1 in both LNCaP and enzalutamide resistant- LNCaP cells. However, GR regulates SGK1 and AR target genes in enzalutamide resistant- LNCaP cells. This study indicated that GR has no effect on the AR target genes in parental LNCaP cells. SGK1 has a vital role in the proliferation, migration and cell cycle of the enzalutamide resistant- LNCaP cell line. In addition, the data from this study showed an increase in the expression of TROP-2 in enzalutamide resistant- LNCaP cells, compared with LNCaP parental cells. The results obtained from this study suggested that TROP-2 might regulates pAkt, pERK1, c-MYC and p27 signalling that are important in proliferation and cell cycle of enzalutamide resistant- LNCaP cells. In addition, TROP-2 potentially regulates the migration of ii enzalutamide resistant- LNCaP cells by its effect on the EMT process that is important in metastases. SGK1 and TROP-2 demonstrated higher protein expression in patients’ tissue samples who had relapsed after androgen withdrawal, compared to naïve patients. In conclusion, SGK1 and TROP-2 could represent either potential biomarkers of enzalutamide-resistance, or potential therapeutic targets in advanced disease.The Higher Committee for Education Development in Ira

Neutrophils are Mediators of Metastatic Prostate Cancer Progression in Bone

Bone metastatic prostate cancer (BM-PCa) significantly reduces overall patient survival and is currently incurable. Current standard immunotherapy showed promising results for PCa patients with metastatic, but less advanced, disease (i.e., fewer than 20 bone lesions) suggesting that PCa growth in bone contributes to response to immunotherapy. We found that: (1) PCa stimulates recruitment of neutrophils, the most abundant immune cell in bone, and (2) that neutrophils heavily infiltrate regions of prostate tumor in bone of BM-PCa patients. Based on these findings, we examined the impact of direct neutrophil-prostate cancer interactions on prostate cancer growth. Bone marrow neutrophils directly induced apoptosis of PCa in vitro and in vivo, such that neutrophil depletion in bone metastasis models enhanced BM-PCa growth. Neutrophil-mediated PCa killing was found to be mediated by suppression of STAT5, a transcription factor shown to promote PCa progression. However, as the tumor progressed in bone over time, neutrophils from late-stage bone tumors failed to elicit cytotoxic effector responses to PCa. These findings are the first to demonstrate that bone-resident neutrophils inhibit PCa and that BM-PCa are able to progress via evasion of neutrophil-mediated killing. Enhancing neutrophil cytotoxicity in bone may present a novel therapeutic option for bone metastatic prostate cancer

Androgen Receptor Inhibition Suppresses Anti-Tumor Neutrophil Response Against Bone Metastatic Prostate Cancer via Regulation of TβRI Expression

Bone metastatic disease of prostate cancer (PCa) is incurable and progression in bone is largely dictated by tumor-stromal interactions in the bone microenvironment. We showed previously that bone neutrophils initially inhibit bone metastatic PCa growth yet metastatic PCa becomes resistant to neutrophil response. Further, neutrophils isolated from tumor-bone lost their ability to suppress tumor growth through unknown mechanisms. With this study, our goal was to define the impact of metastatic PCa on neutrophil function throughout tumor progression and to determine the potential of neutrophils as predictive biomarkers of metastatic disease. Using patient peripheral blood polymorphonuclear neutrophils (PMNs), we identified that PCa progression dictates PMN cell surface markers and gene expression, but not cytotoxicity against PCa. Importantly, we also identified a novel phenomenon in which second generation androgen deprivation therapy (ADT) suppresses PMN cytotoxicity via increased transforming growth factor beta receptor I (TβRI). High dose testosterone and genetic or pharmacologic TβRI inhibition rescued androgen receptor-mediated neutrophil suppression and restored neutrophil anti-tumor immune response. These studies highlight the ability to leverage standard-care ADT to generate neutrophil anti-tumor responses against bone metastatic PCa