Characterising mechanisms of aberrant mutant androgen receptor signalling in advanced prostate cancer

Abstract

Prostate cancer (PC) is the most commonly diagnosed disease in the UK which causes approximately 10,000 deaths annually. Although an initially effective response to androgen deprivation therapy (ADT) occurs in most patients, the tumour normally recurs in a more aggressive form of the disease termed castrate resistant PC (CRPC) and is largely untreatable at this stage. In many cases, disease is driven by inappropriate androgen receptor (AR) signalling. It is therefore vital to have better understanding of mechanisms that re-activate AR and promote ADT resistance in the clinic and hence better treatments for advanced tumour. Activation of AR by testosterone is crucial for prostate growth and transformation. Antiandrogens, the second most common PC therapy after surgery, antagonise ligand binding to the receptor and hence deactivate AR signalling. In 2012, enzalutamide, a more potent agents in terms of availability to block AR was approved by the FDA and ENA as a second-generation antiandrogen for clinical usage. Although it demonstrated several advantages over its pervious counterpart bicalutamide, response rates of just 50% in CRPC patients and subsequent resistance observed in responders haslimited its effectiveness. Critically,several lines of evidence from preclinical models and patient samples indicate that one particular resistant mechanism is the emergence of AR mutant(s), in part, driven by a specific AR mutation F876L that enables the compound to act as an agonist. Importantly, the same mutant was later detected in metastatic PC patient had been treated with apalutamide. Evidently, novel therapies emerging into clinical treatment of advance disease have the added challenge of being efficacious in the background of mutant AR and thus developing model systems to test this is of paramount importance. In order to enable more physiological modelling of aberrant ARF876L activity that would highlight potentially distinct mechanisms that could be exploited in future therapies, this project aimed to generate CRISPR-edited LNCaP and CWR22Rv1 cell lines expressing the enzalutamide-activated ARF876L mutant. Meanwhile, a part of project has also focused on generation of a physiologically relevant AR rescue/replacement in vitro cell line model (LNCaP- ARF876L cells) which permits ability forstudying ARF876L directly regulated gene expression profiles by effectively knockdown endogenous AR without impacting on the ectopically expressed mutant. Furthermore, by using 3 Illumina Human HT-12 arrays analysing LNCaP-ARF876L cells revealed a comprehensive transcriptomic data-set to provide an insight into how an enzalutamide-activated AR mutant can drive a distinct gene-set in advanced PC. This is important as it may enable distinct biomarker discovery in enzalutamide-resistance disease and has highlighted interplay between the ARF876L mutation and the glucocorticoid receptor. Lastly, the LNCaP- ARF876L cell lines was utilised to demonstrate that aberrantly-functioning receptor is sensitive to BET inhibitors. In all, the work hasshown that the ARF876L mutant drives a distinct transcriptional programme to the endogenous AR in LNCaP cells and the model can be utilised effectively to indicate sensitivities of the receptor to clinically-relevant compounds.