Edinburgh Napier University

  Background: Neuroendocrine differentiation is commonplace in prostate tumors treated with androgen deprivation therapy and is associated with castrate-resistant disease progression and poor prognosis. The molecular mechanisms driving neuroendocrine differentiation and the role of neuroendocrine-like cells in supporting growth and increased aggression of castrate-resistant prostate cancer remains unclear. This study aims to assess the expression of key genes and proteins relating to neuroendocrine differentiation and investigate the potential involvement of γ-aminobutyric acid (GABA) in paracrine maintenance of prostate cancer growth and neuroendocrine phenotype.


Method: Androgen-sensitive LNCaP prostate cancer cells were cultured in charcoal-stripped RPMI 1640 media to simulate androgen deprivation. Gene and protein expression was analysed via quantitative PCR, immunoblotting and confocal microscopy. Morphological changes were assessed using light microscopy.


Results: Androgen deprived LNCaP cells showed loss of detectable prostate specific antigen (PSA) expression and upregulation of canonical neurogenesis promoter achaete-scute homolog-1 (hASH1). Neuron specific enolase (NSE), was upregulated by androgen deprivation and LNCaP cells acquired a neuroendocrine-like morphology, evidenced by neurite-like protrusions. Neurogenesis repressor, RE1-Silencing Transcription (REST) factor expression was unchanged by androgen deprivation, however PTOV1 and GAD1 were upregulated. Vimentin and MMP-9 expression increased after androgen deprivation, suggesting neuroendocrine-like cells promote a more aggressive phenotype. Supplementing charcoal-stripped media with 1 nM synthetic androgen (R1881) prevented neuroendocrine-like changes in morphology. Androgen deprivation associated increases in PTOV1, NSE and MMP-9 expression were also attenuated by 1 nM R1881 and PSA expression was maintained. Whilst GAD1 gene expression was upregulated, GAD65/67 protein expression was unchanged, suggesting GABA synthesis by neuroendocrine-like cells may occur via an alternative pathway.


Conclusion: Androgen deprivation induces LNCaP cells to become neuroendocrine-like via upregulation of PTOV1 and hASH1. Identifying molecular drivers of neuroendocrine differentiation and their role in prostate cancer progression will significantly enhance our understanding of how neuroendocrine signaling may support highly aggressive castrate resistant growth.