No abstract

Treatment with androgen receptor pathway inhibitors (ARPIs) in prostate cancer leads to the emergence of resistant tumors characterized by lineage plasticity and differentiation toward neuroendocrine lineage. Here, we find that ARPIs induce a rapid epigenetic alteration mediated by large-scale chromatin remodeling to support activation of stem/neuronal transcriptional programs. We identify the proneuronal transcription factor ASCL1 motif to be enriched in hyper-accessible regions. ASCL1 acts as a driver of the lineage plastic, neuronal transcriptional program to support treatment resistance and neuroendocrine phenotype. Targeting ASCL1 switches the neuroendocrine lineage back to the luminal epithelial state. This effect is modulated by disruption of the polycomb repressive complex-2 through UHRF1/AMPK axis and change the chromatin architecture in favor of luminal phenotype. Our study provides insights into the epigenetic alterations induced by ARPIs, governed by ASCL1, provides a proof of principle of targeting ASCL1 to reverse neuroendocrine phenotype, support luminal conversion and re-addiction to ARPIs.

Most prostate cancer patients will progress to a castration-resistant state (CRPC) after androgen ablation therapy and despite the development of new potent anti-androgens, like enzalutamide (ENZ), which prolong survival in CRPC, ENZ-resistance (ENZR) rapidly occurs. Re-activation of the androgen receptor (AR) is a major mechanism of resistance. Interrogating our in vivo derived ENZR model, we discovered that transcription factor STAT3 not only displayed increased nuclear localization but also bound to and facilitated AR activity. We observed increased STAT3 S727 phosphorylation in ENZR cells, which has been previously reported to facilitate AR binding. Strikingly, ENZR cells were more sensitive to inhibition with STAT3 DNA-binding inhibitor galiellalactone (GPA500) compared to CRPC cells. Treatment with GPA500 suppressed AR activity and significantly reduced expression of Cyclin D1, thus reducing cell cycle progression into S phase and hindering cell proliferation. In vivo, GPA500 reduced tumor volume and serum PSA in ENZR xenografts. Lastly, the combination of ENZ and GPA500 was additive in the inhibition of AR activity and proliferation in LNCaP and CRPC cells, providing rationale for combination therapy. Overall, these results suggest that STAT3 inhibition is a rational therapeutic approach for ENZR prostate cancer, and could be valuable in CRPC in combination with ENZ.

Despite the development of effective targeted therapies and a significant understanding of carcinogenesis and cancer progression, treatment resistance is a major obstacle in achieving durable long-term control in many types of cancers. Emerging evidence supports that non-genetic mechanisms could play an underappreciated role in therapy resistance. These mechanisms include phenotypic plasticity, which is recognized as one of the hallmarks of cancer and translates to epigenetic and transcriptional control of gene expression. Alterations in the expression and activity of the epigenetic modifier enhancer of zeste homolog 2 (EZH2) support prostate cancer lineage plasticity and progression. EZH2 expression and activity is elevated in castration-resistant prostate cancer treated with androgen receptor (AR) pathway inhibitors and in treatment-resistant prostate cancer. Moreover, 17 known residues of EZH2 are phosphorylated on by multiple kinases that modulate its activity, localization, stability, and polycomb repressive complex (PRC2) assembly. In this review, we explore the contribution of EZH2 phosphorylation in regulating canonical PRC2 in a methylation-dependent manner as an epigenetic repressor and in a non-canonical manner independent of PRC2 as a transcription activator. Apart from the contribution of EZH2 phosphorylation at serine 21, threonine 350, and threonine 311 in prostate cancer progression and treatment resistance, we discuss how other EZH2 phosphorylated residues with unknown functions could contribute to prostate cancer based on their upstream regulators and potential therapeutic utility.

260 Background: Resistance to newly developed androgen receptor pathway inhibitors (ARPIs), such as Enzalutamide (ENZ), rapidly emerges and patients generally die within two years. In particular, a subset of patients who relapse following ARPI therapy exhibit lineage switching whereby tumours shed their dependence on AR signaling and emerge with neuroendocrine features. These tumours, termed treatment induced neuroendocrine prostate cancer (t-NEPC), carry an extremely poor prognosis and, to date, treatment remains decades old cytotoxic chemotherapies; therefore, targeted therapies are desperately needed. Recently our group identified the neural transcription factor BRN2 as a major clinically relevant driver of NEPC and aggressive tumor growth, both in vitro and in vivo, suggesting targeting BRN2 is a promising strategy to prevent neuroendocrine differentiation or treat NEPC. Methods: Study the effects of BRN2 inhibition using siRNA, small molecule inhibitors (BRN2i) and CRISPR K/O models. The efficacy of the small molecules was examined using reporter assays, florescence polarization assays, Biolayer Interferometry, DARTS, chromatin fractionation, RNA-seq and ChIP-seq. Pharmacokinetic studies measured stability and bioavailability of the molecules and in-vivo efficacy is to be measured in NCI-H660 xenograft model. Results: Inhibition of BRN2 drastically reduced cell proliferation in NEPC cell lines 42DENZR and NCI-H660 cell lines. Targeting BRN2 with our first-in-field small molecule inhibitors lead to downregulation several known targets in NEPC like EZH2, ASCL1, SOX2 and PEG10. Treatment with BRN2i reduced recruitment of BRN2 to the chromatin by approximately 93% within 16 hours. Moreover, these BRN2 inhibitors displayed adequate pharmacokinetic properties and reduced NEPC proliferation in vivo. Conclusions: No therapies exist for highly lethal NEPC. Hence, the described work aims to lay the pre-clinical foundation for the integration of BRN2 targeted therapies into the treatment landscape to improve survival for patients suffering from small cell prostate cancer.