MON-321 - Identification and Characterization of SRRM4-Regulated Microexons in Neuroendocrine Prostate Cancer

Neuroendocrine prostate cancer (NEPC) is a highly advanced and lethal form of prostate cancer (PCa) that is not responsive to androgen deprivation therapy. Progression to NEPC is driven by the transdifferentiation of prostate luminal cells into neuroendocrine (NE) cells that support androgen-independent tumor growth through paracrine signaling. As a form of cellular reprogramming, the luminal-to-NE transition is thought to be orchestrated by transcriptional and post-transcriptional mechanisms. While many transcriptional drivers of NE differentiation are known, the potential contribution of alternative splicing to NEPC development has not yet been fully characterized. To investigate potential splicing dysregulation in NEPC, we analyzed RNA-sequencing datasets from castration-resistant PCa (CRPC) and NEPC patient samples and identified differential microexon inclusion events in NEPC tumors. These microexons are mostly in-frame and are also enriched in neural tissues, implicating the role of neural-like splicing in shaping pathways involved in NEPC development. Differential expression analysis of splicing factors and differential splicing analysis of splicing factor overexpression models then showed that these microexons are largely regulated by the splicing factor SRRM4, suggesting that a program of SRRM4-regulated microexons is activated during NE transformation. Gene ontology analysis of the microexon-containing genes indicated their involvement in vesicular trafficking, migratory behavior, and tumor adaptation – processes that contribute to the maintenance and progression of NEPC. To determine the functional role of SRRM4 in NEPC, we generated lentiviral-mediated knockdown and conditional overexpression lines of SRRM4 in 22Rv1, a cell model of CRPC with NE features. SRRM4 knockdown resulted in reduced inclusion of NEPC-associated microexons in an isoform-specific RT-qPCR analysis and decreased expression of genes associated with the NE phenotype. Further phenotypic characterization of the SRRM4-edited cell lines is currently underway to elucidate the role of SRRM4 in mediating a NE phenotype in PCa. Taken together, our data will contribute to our understanding of the role of alternative splicing in the development of NEPC, and identify potential targets for NEPC therapy.

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