Discovery and Biological Characterization of PRMT5:MEP50 Protein-Protein Interaction Inhibitors.

Protein arginine methyltransferase 5 (PRMT5) is a master epigenetic regulator and an extensively validated therapeutic target in multiple cancers. Notably, PRMT5 is the only PRMT that requires an obligate cofactor, methylosome protein 50 (MEP50), to function. We developed compound 17, a novel small-molecule PRMT5:MEP50 protein-protein interaction (PPI) inhibitor, after initial virtual screen hit identification and analogue refinement. Molecular docking indicated that compound 17 targets PRMT5:MEP50 PPI by displacing the MEP50 W54 burial into a hydrophobic pocket of the PRMT5 TIM barrel. In vitro analysis indicates IC50 < 500 nM for prostate and lung cancer cells with selective, specific inhibition of PRMT5:MEP50 substrate methylation and target gene expression, and RNA-seq analysis suggests that compound 17 may dysregulate TGF-ß signaling. Compound 17 provides a proof of concept in targeting PRMT5:MEP50 PPI, as opposed to catalytic targeting, as a novel mechanism of action and supports further preclinical development of inhibitors in this class.

Targeting Protein Arginine Methyltransferase 5 Suppresses Radiation-induced Neuroendocrine Differentiation and Sensitizes Prostate Cancer Cells to Radiation.

Prostate cancer remains the second leading cause of cancer death among American men. Radiotherapy is a potentially curative treatment for localized prostate cancer, and failure to control localized disease contributes to the majority of prostate cancer deaths. Neuroendocrine differentiation (NED) in prostate cancer, a process by which prostate adenocarcinoma cells transdifferentiate into neuroendocrine-like (NE-like) cells, is an emerging mechanism of resistance to cancer therapies and contributes to disease progression. NED also occurs in response to treatment to promote the development of treatment-induced neuroendocrine prostate cancer (NEPC), a highly aggressive and terminal stage disease. We previously demonstrated that by mimicking clinical radiotherapy protocol, fractionated ionizing radiation (FIR) induces prostate cancer cells to undergo NED in vitro and in vivo. Here, we performed transcriptomic analysis and confirmed that FIR-induced NE-like cells share some features of clinical NEPC, suggesting that FIR-induced NED represents a clinically relevant model. Furthermore, we demonstrated that protein arginine methyltransferase 5 (PRMT5), a master epigenetic regulator of the DNA damage response and a putative oncogene in prostate cancer, along with its cofactors pICln and MEP50, mediate FIR-induced NED. Knockdown of PRMT5, pICln, or MEP50 during FIR-induced NED and sensitized prostate cancer cells to radiation. Significantly, PRMT5 knockdown in prostate cancer xenograft tumors in mice during FIR prevented NED, enhanced tumor killing, significantly reduced and delayed tumor recurrence, and prolonged overall survival. Collectively, our results demonstrate that PRMT5 promotes FIR-induced NED and suggests that targeting PRMT5 may be a novel and effective radiosensitization approach for prostate cancer radiotherapy.

PRMT5: a putative oncogene and therapeutic target in prostate cancer.

Protein arginine methyltransferase 5 (PRMT5) was discovered two decades ago. The first decade focused on the biochemical characterization of PRMT5 as a regulator of many cellular processes in a healthy organism. However, over the past decade, evidence has accumulated to suggest that PRMT5 may function as an oncogene in multiple cancers via both epigenetic and non-epigenetic mechanisms. In this review, we focus on recent progress made in prostate cancer, including the role of PRMT5 in the androgen receptor (AR) expression and signaling and DNA damage response, particularly DNA double-strand break repair. We also discuss how PRMT5-interacting proteins that are considered PRMT5 cofactors may cooperate with PRMT5 to regulate PRMT5 activity and target gene expression, and how PRMT5 can interact with other epigenetic regulators implicated in prostate cancer development and progression. Finally, we suggest that targeting PRMT5 may be employed to develop multiple therapeutic approaches to enhance the treatment of prostate cancer.

Protein Arginine Methyltransferase 5 Promotes pICln-Dependent Androgen Receptor Transcription in Castration-Resistant Prostate Cancer.

The majority of advanced prostate cancer therapies aim to inhibit androgen receptor (AR) signaling. However, AR reactivation inevitably drives disease progression to castration-resistant prostate cancer (CRPC). Here we demonstrate that protein arginine methyltransferase 5 (PRMT5) functions as an epigenetic activator of AR transcription in CRPC, requiring cooperation with a methylosome subunit pICln. In vitro and in xenograft tumors in mice, targeting PRMT5 or pICln suppressed growth of CRPC cells. Full-length AR and AR-V7 transcription activation required both PRMT5 and pICln but not MEP50. This activation of transcription was accompanied by PRMT5-mediated symmetric dimethylation of H4R3 at the proximal AR promoter. Further, knockdown of PRMT5 abolished the binding of pICln (but not vice versa) to the AR proximal promoter region, suggesting that PRMT5 recruits pICln to the AR promoter to activate AR transcription. Differential gene expression analysis in 22Rv1 cells confirmed that PRMT5 and pICln both regulate the androgen signaling pathway. In addition, PRMT5 and pICln protein expression positively correlated with AR and AR-V7 protein expression in CRPC tissues and their expression was highly correlated at the mRNA level across multiple publicly available CRPC datasets. Our results suggest that targeting PRMT5 or pICln may be explored as a novel therapy for CRPC treatment by suppressing expression of AR and AR splice variants to circumvent AR reactivation. SIGNIFICANCE: This study provides evidence that targeting PRMT5 can eliminate expression of AR and can be explored as a novel therapeutic approach to treat metastatic hormone-naïve and castration-resistant prostate cancer.

PRMT5 Cooperates with pICln to Function as a Master Epigenetic Activator of DNA Double-Strand Break Repair Genes.

DNA double-strand break (DSB) repair is critical for cell survival and genome integrity. Upon recognition of DSBs, repair proteins are transiently upregulated to facilitate repair through homologous recombination (HR) or non-homologous end joining (NHEJ). We present evidence that PRMT5 cooperates with pICln to function as a master epigenetic activator of DNA damage response (DDR) genes involved in HR, NHEJ, and G2 arrest (including RAD51, BRCA1, and BRCA2) to upregulate gene expression upon DNA damage. Contrary to the predominant role of PRMT5 as an epigenetic repressor, our results demonstrate that PRMT5 and pICln can activate gene expression, potentially independent of PRMT5's obligate cofactor MEP50. Targeting PRMT5 or pICln hinders repair of DSBs in multiple cancer cell lines, and both PRMT5 and pICln expression positively correlates with DDR genes across 32 clinical cancer datasets. Thus, targeting PRMT5 or pICln may be explored in combination with radiation or chemotherapy for cancer treatment.