Abstract Introduction: Upon prolonged anti-androgen treatment, 15-20% of castration resistant prostate cancer (CRPC) progresses to poorly differentiated, androgen-independent neuroendocrine prostate cancer (NEPC) that express neural markers through a mechanism of lineage plasticity. Data from patient tumors have revealed a distinct DNA methylome profile between NEPC and CRPC, suggesting the relevance of DNA methylation during lineage plasticity in advanced CRPC. EZH2, a catalyzer of repressive H3K27me3 deposition is overexpressed in NEPC. EZH2 inhibition is being studied as a therapeutic strategy for advanced prostate cancer in clinical trials. Although evidence from embryonic stem cell studies suggest a possible interplay between DNA methylation and EZH2 activity, their cooperativity in driving lineage plasticity in advanced CRPC is unknown. Methods: DNA methylome and H3K27me3 deposition in patient-derived xenografts/organoids and murine model engineered with MYCN induction and Pten/Rb1 co-loss were analyzed following EZH2 or DNMT1 depletion, respectively. We also assessed the impact of EZH2/DNMT1 depletion on gene transcription. Results: Both DNA methylome and H3K27me3 profile shifted during progression of CRPC to NEPC. H3K27me3 was enriched at hypomethylated regions in NEPC compared to CRPC tumors. EZH2 loss resulted in changes at clinically relevant NEPC-associated differentially methylated regions. Neuron-related genes were hypomethylated and upregulated while, morphogenesis-related and bivalent genes were hypermethylated and downregulated upon EZH2 deletion. Conversely, DNMT1 deletion altered H3K27me3 landscape in NEPC. Neuron-related genes and bivalent genes were downregulated and enriched with H3K27me3 but were upregulated and depleted in H3K27me3 in CRPC upon DNMT1 deletion. H3K4me3 was also decreased upon DNMT1 depletion in NEPC at these regions suggesting loss of bivalency. Short-term DNMT inhibition using decitabine in NEPC xenografts resulted in similarly altered H3K27me3 deposition as noted upon DNMT deletion. Conclusions: DNMT1 and EZH2 activity overlaps and together reprograms the epigenome during lineage plasticity of advanced CRPCs. EZH2 loss was partially compensated by DNMT activity leading to further neuroendocrine differentiation while DNMT1 loss shifted H3K27me3 leading to suppression of neuron-related genes. Citation Format: Richa Singh, Varadha Balaji. Venkadakrishnan, Eddie Imada, Yasutaka Yamada, Nicholas J. Brady, Kate Dunmore, Richard Garner, Brian D. Robinson, David S. Rickman, Himisha Beltran. Cooperativity between DNA methylation and EZH2 activity drives neuroendocrine phenotype in advanced prostate cancer abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Innovations in Prostate Cancer Research and Treatment; 2026 Jan 20-22; Philadelphia PA. Philadelphia (PA): AACR; Cancer Res 2026;86 (2Suppl): Abstract nr PR035.
Singh et al. (Tue,) studied this question.