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Molecular mechanism and relevance of microRNAs in DSB repair pathway choice


? DESCRIPTION (provided by applicant): A dividing cell constantly exposed to environmental and endogenous DNA damaging agents can transform into a tumor due to incorrect repair. Conversely, radiation and chemotherapeutic agents eradicate tumors by inducing irreparable DNA damage. Cancer cells often develop resistance to therapy by manipulating the DNA repair machinery. Therefore we need to have an in-depth understanding of DNA repair and the factors involved, both for gaining insight into the cause of cancer and to enhance the efficacy of cancer therapy. Double stranded DNA breaks (DSBs) are critical for cell health as a single unrepaired DSB is sufficient for inducing apoptosis. Two major mechanistically distinct pathways, homologous recombination (HR) and non-homologous end joining (NHEJ) have evolved to repair DSBs. A decisive factor in the choice between DSB repair pathways is in the competition between DNA end protection (necessary for NHEJ) and DNA end resection (necessary for HR). DSB end resection must be restricted to S/G2 phases of the cell cycle, as HR requires the presence of an intact sister chromatid. Depletion of factors such as 53BP1 allows DNA end resection in the G1 phase, thereby impairing DSB repair and causing genomic instability. Likewise, loss of the HR protein, BRCA1 allows the error-prone NHEJ pathway to dominate throughout the cell cycle potentially leading to tumorigenesis. However, loss of BRCA1 provides a therapeutic opportunity as these tumors are exquisitely sensitive to inhibitors of poly (ADP-ribose) polymerase (PARP), and are also susceptible to platinum-based drugs. Surprisingly, loss of 53BP, or associated factors (RIF1, PTIP) in BRCA1 mutant tumors make them resistant to PARP inhibitors with restoration of HR. Therefore factors that regulate optimal expression of HR and NHEJ proteins are crucial for pathway choice, and may have significant relevance in cancer therapy. We discovered that a new class of gene expression regulators, microRNA (miRNA)s down-modulate DSB repair proteins, and influence specific repair pathways. MiRNAs are abundant small non-coding RNAs that typically dampen gene expression and are aberrantly expressed in tumors. We recently observed that miRNA-mediated suppression of the HR pathway specifically in the G1 phase maybe critical for genomic stability. Building on these observations we conducted independent functional screens to identify candidate miRNAs that regulate the HR-pathway (miR-1231, miR-876-3p, miR-221*and miR-185*) and the NHEJ pathway (miR-142-5p, miR-502, miR-622, and miR-597). Here we will investigate the hypothesis that miRNAs regulate the choice of DSB repair pathways by maintaining the cell cycle phase specific expression of HR and NHEJ factors. We will test the idea that a miRNA-induced, modest decrease in specific DSB repair factors may differentially impact the repair of DSBs in euchromatin versus heterochromatin/ intergenic loci. Finally we will investigate clinical relevance of the candidate miRNAs in primary ovarian tumors and lymphomas, that is, correlate miRNA expression with response to PARP inhibitors/platinum based therapy and overall prognosis.

Funded by the NIH National Center for Advancing Translational Sciences through its Clinical and Translational Science Awards Program, grant number UL1TR002541.