Abstract Purpose: SETD2 aberrancy, as the main reason of depleted levels of H3K36me3, plays an important role in diverse forms of cancer and is most notably in clear cell renal cell carcinoma (ccRCC). Altered chromatin modelers related to impaired H3K36me3 occupancy are linked to renal cancer aggressiveness and metastasis, where the latter dramatically drops the relative 5-year survival rate of ccRCC from 91% to 11%. Developing novel treatment targeting H3K36me3-compromised cancer cell is urgently needed for diagnostic and treatment purposes. Methods: Phenotypic and animal experiments were conducted to test the sensitivity of kidney cancer cells with impaired H3K36me3 to hypomethylating agent (HMA), PARP inhibitor (PARPi), and HMA + PARPi combination treatments. Comet assay, flow cytometry, western blot and bioinformatic analysis were performed to explore the mechanism beneath. Results: Combination treatment of 5-Aza-CdR (DAC) and Talazoparib (PARPi) generate more cytotoxicity in SETD2-altered ccRCC cell lines than in SETD2-intact cell lines, including inhibited cell growth, incomplete colony formation, more apoptosis, increased DNA damage and impaired DNA damage repair. The up-regulation of STING and transposable element (TE), as well as the loss of chromatin stability, were more remarkable in SETD2-altered ccRCC cell lines when treated with DAC + PARPi. Conclusion: Combination treatment of DAC and Talazoparib showed greater synergistic anti-tumor effect in SETD2 aberrant than in SETD2 wild-type ccRCC cells, making it a potential treatment strategy for H3K36me3-compromised cancers. Further pre-clinical studies should be conducted to make a precise medicine-based approach. Citation Format: Xinyi Zhou, Guanghou Fu, Hong-Tao Li, Hemant Gujar, Daniel J. Weisenberger, David I. Quinn, Bo Han, Gangning Liang. SETD2 aberrancy enhanced the synergetic anti-tumor effects of DNA hypomethylating agents and PARP inhibitors in aggressive clear cell renal cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3736.