Abstract:

Prostate cancer remains the second leading cause of cancer-related deaths in American men. The cellular processes of the prostate, including prostate cancers, are governed by a nuclear hormone receptor known as the androgen receptor (AR). Androgen binding to AR controls the expression of thousands of target genes in prostate cells. These genes influence many important signaling pathways including cell growth, survival, metabolism, and DNA repair. Early disease interventions that include androgen deprivation therapies (ADTs) are successful; however, these therapies eventually become ineffective, and to this day, castrate resistant prostate cancer (CRPC) treatment remains an unmet clinical need. CRPC is marked by changes in signaling pathways such as cell growth, motility, and DNA repair. Understanding these changes and determining crucial regulators of these changes will provide alternative and improved therapies for CRPC. Because recent evidence has shown that androgen and DNA repair signaling are closely linked processes, we conducted a comprehensive assessment of expression level changes of DNA repair factors by androgen signaling in multiple prostate cancer cell lines, in disease progression models of prostate cancer, and using prostate cancer patient data. Using high throughput sequencing, we uncovered a previously unreported DNA repair gene set that was regulated by androgen signaling in multiple prostate cancer cell lines. Genomic analyses of these prostate cancer cell lines also provided key information related to the mutation status of genes encoding DNA repair proteins. This interrogation of prostate cancer preclinical models is likely to inform future studies in the field. DNA repair factors are not only found to be regulated by androgen-mediated transcription, but other laboratories have shown that DNA repair machinery is also required for transcriptional activities; this includes the activities of nuclear hormone receptors. As part of this study, we tested a new inhibitor that targets one such DNA repair factor. We used mirin, a small molecule inhibitor against the DNA-damage sensing protein MRE11. We found that mirin treatment of prostate cancer cells significantly inhibits the transcription of several androgen-regulated genes. Treatment with mirin also resulted in cell growth inhibition and cell cycle arrest of prostate cells. In hormone-driven cancers like breast and prostate, clinical trials recently showed that the molecular targeting of the DNA repair protein Parp1 with Olaparib improved patient outcomes in individuals with DNA repair gene mutations. Other Parp family members have emerged as central players in disease development processes like chemo-resistance and cell survival. Parp9 is one family member that along with the heterodimer partner, Dtx3L, function in the DNA damage response. In this study, we characterized the dual enzymatic nature of this Dtx3L/Parp9 heterodimer in the context of prostate cancer. Here we report the previously unknown ADP-ribosylation activity of Parp9 towards the C-terminal end of ubiquitin as well as the regulation of the E3 ubiquitin ligase activity of Dtx3L by NAD+. These activities were shown to be related to the DNA damage response, particularly the NHEJ repair pathway. The role of Dtx3L and Parp9 in prostate cells also includes a direct interaction with AR via the macrodomains of Parp9. Dtx3L/Parp9 association with AR depends on AR ADP-ribosylation by Parp7. Our data also showed that Parp7 is another apparent target of the clinically available Parp inhibitor, Olaparib. Overall, the experiments from this study show novel relationships between DNA repair signaling and androgen signaling, and provide preclinical results that will support and potentially complement current therapies for prostate cancer.

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