Despite advances in treatment of prostate cancer, metastatic castration-resistant prostate cancer (mCRPC) remains fatal: in the U.S. alone, over 30,000 patients die yearly from mCRPC. The developmental mechanism of mCRPC is unknown. The goal of this project is to explain the role in DNA repair of a previously unstudied gene, FAM35A. I have evidence that FAM35A is a novel tumor/metastasis suppressor in prostate cancer, and that its depletion in BRCA1-deficient cancer confers resistance to camptothecin. Based on my data, I predict that FAM35A is involved in development of resistance in mCRPC. The mechanism of FAM35A involvement in metastasis and resistance is completely uncharacterized; however, clarification of FAM35A's specific role in DNA repair is of great import to effective cancer treatment.
Specific Aims : The overall hypothesis is that FAM35A participates in end-joining (EJ).
Aim 1. Determine the function of FAM35A in double strand break (DSB) repair a) Confirm FAM35A participation in EJ by random plasmid integration and EJ5-GFP reporter assay. Expected results: Results from these studies should confirm FAM35A participation in EJ. b) Determine the specific pathway of EJ involving FAM35A by comparing the sensitivity of alt-NHEJ mutant cells infected with Fam35a shRNAs vs control after treatment with DNA damaging agents. Expected results: Fam35a shRNA depleted Polq-/- MEF is more sensitive to DNA damage than control.
Aim 2. Mechanism of chemoresistance in FAM35A-depleted BRCA1-deficient cells a) Monitor the response to DNA damaging agents in FAM35A-depleted BRCA1-inactivated cells. Method: Colony-forming assay. Expected results: FAM35A depletion in BRCA1 mutant cells will confer resistance to PARPi. Furthermore, FAM35A depletion in LNCaP cells (prostate cancer cell line) will also generate resistance to camptothecin and PARPi. b) Determine whether FAM35A inhibits resection by counting RAD51 foci and monitoring ssDNA formation. Method: Immunohistochemistry and fluorescence activated cell sorting. Expected results: FAM35A functions alongside 53BP1, RIF1 and REV7 to inhibit resection of the 5' ends of DSBs. In total, my preliminary findings support FAM35A as a new target for cancer treatment and substantiate its involvement in the EJ pathway of DNA repair. Results from these (and future) studies are expected to advance mCRPC etiology, elucidate acquisition of metastasis and resistance in PCa and, most importantly, to facilitate discovery of therapeutic and prognostic targets for both chemotherapy and radiation therapy.
Despite advances in treatment of prostate cancer, metastatic castration-resistant prostate cancer (mCRPC) remains fatal in the U.S. alone, over 30,000 patients die yearly from mCRPC. Our preliminary findings support FAM35A as a new target for cancer treatment and substantiate its involvement in the end-joining pathway of DNA repair. Results from these (and future) studies are expected to advance mCRPC etiology, elucidate acquisition of metastasis and resistance in PCa and, most importantly, to facilitate discovery of therapeutic and prognostic targets for both chemotherapy and radiation therapy.
| Tomida, Junya; Takata, Kei-Ichi; Bhetawal, Sarita et al. (2018) FAM35A associates with REV7 and modulates DNA damage responses of normal and BRCA1-defective cells. EMBO J 37: |
