Characterising novel phosphatase enzymes important in regulating androgen receptor function in the progression of castrate resistant prostate cancer

Abstract

Prostate cancer (PC) is the most common oncogenic malignancy in men in the UK, and is expected to affect 1 in 4 men throughout their lifetime. Whilst the treatment for organ confined PC is initially very effective, no successful therapies exist for patients where the disease has progressed to an advanced stage, and is reflected by the poor 5-year survival rate of 30%. Resistance to current treatment modalities, aimed at disrupting the androgen signalling axis, renders the disease what is termed castrate resistant PC (CRPC). Crucially, the primary target in the treatment of PC, the androgen receptor (AR), remains a key driver of disease survival and differentiation throughout disease progression. Activation of the AR at the post-translational level by aberrant co-activator activity is a well-established resistance mechanism, however, the role of phosphatase enzymes on AR function represents a significant knowledge gap in AR regulation. To address this issue, phosphatase enzymes identified from a human phosphatome RNAi screen in the androgen responsive PC cell line, LNCaP, were characterised within the context of AR regulation. As such, myosin phosphatase (MLCP) was identified as a novel negative regulator of AR activity. Robust molecular biology techniques revealed that MLCP repressed AR function via indirect mechanisms involving the activation of the clinically relevant tumour suppressors RB1 and NF2. With this in mind, it was possible to identify the endogenous MLCP inhibitors, PPP1R14C and NUAK 1/2, as novel AR activators and potential therapeutic targets in both PC and CRPC. Disruption of either PPP1R14C or NUAK 1/2, via RNAi or small molecule inhibition, respectively, repressed AR transcriptional activity, characterised by reduced protein stability and impaired ligand induced nuclear translocation, culminating in reduced PC cell cycle progression, migration and proliferation, providing significant evidence for a novel, and therapeutically exploitable, AR regulatory mechanism.