Investigating USP2 as a mediator of therapy resistance in lethal prostate cancer

Abstract

The growth of prostate cancer (PCa) is dependent on male sex hormones, termed androgens, and the androgen receptor (AR). Therefore, implementing strategies to inhibit AR activity, collectively referred to as androgen deprivation therapy (ADT), is the key therapeutic strategy for metastatic prostate cancer. Unfortunately, ADT is never curative, and patients eventually develop a lethal therapy resistant form of the disease termed castration-resistant prostate cancer (CRPC). The AR signalling pathway is altered in CRPC and a subset of CRPC tumours may evade inhibition by ADT by progressing to a state in which tumour growth is independent of this pathway. One such AR-independent CRPC subtype is termed neuroendocrine PCa (NEPC). Understanding how tumours transition to therapy-resistant, AR-independent states is crucial for the development of new and more effective therapies. We recently undertook transcriptomic profiling of patient tumours treated ex vivo with a clinical AR antagonist, enzalutamide, as a strategy to identify therapy-mediated adaptive changes. This study identified Ubiquitin specific protease 2 (USP2) as being increased in response to enzalutamide. The role of USP2 is to remove ubiquitin groups from proteins that causes therapy resistance and cancer progression, thereby preventing these proteins from degradation and increasing their stability in cancer cells. Thus, we hypothesised that USP2 can mediate resistance to ADT by stabilising key oncoproteins, a concept that was tested in my PhD project. Increased USP2 expression in response to ADT and in therapy-resistant states was validated in multiple prostate cancer cell line models, clinical transcriptomic cohorts, and additional patient tumours. Growing androgen-dependent cell lines in the presence of enzalutamide resulted in increased USP2 expression; conversely, androgen treatment resulted in the repression of USP2 expression. Clinical datasets also revealed that USP2 expression is elevated in AR-low/negative CRPC tumours, particularly those classified as neuroendocrine PCa. These observations suggest that USP2 is consistently upregulated in response to AR-targeted therapies and may represent a previously unknown resistance factor. Supporting this, targeting USP2 in multiple castrate-resistant prostate cancer models, either pharmacologically with a USP2-specific inhibitor (ML364) or by genetic knockdown, resulted in reduced cell viability and increased cell death. In contrast, overexpression of USP2 drove the development of an aggressive, therapy-resistant neuroendocrine phenotype and conferred partial resistance to enzalutamide and a growth advantage in androgen-depleted growth conditions. Importantly, ML364 was also potently active in an in vivo model of aggressive, AR-negative prostate cancer. Mechanistically, we found that USP2 stabilises the levels of oncogenic proteins including Aurora kinase A (AURKA), Cyclin D1 and Fatty acid synthase (FAS). Interrogation of the proteome and transcriptome of PCa cells overexpressing USP2 revealed positive enrichment of neuroendocrine-associated signalling, lipid metabolism and cell cycle, and negative enrichment of interferon signalling. All these enriched pathways are important in driving growth, survival, and progression of cancer. Overall, the findings herein revealed that USP2 can promote the development of neuroendocrine prostate cancer and acts to confer resistance to standard-of-care therapies, revealing it as a bona fide therapeutic target.