Abstract Epigenetic alteration contributes to tumor initiation, progression and invasion. Epigenetic machinery has become a major focus for the new development of molecular targeted cancer therapeutics. H3K4 trimethylation (H3K4me3) is an epigenetic mark which exist surrounding the transcription start site and have been presumed to be associated with cancer stem like cells. JARID1 family is demethylase of H3K4me2/me3. Especially, JARID1a and JARID1b have been suggested to have oncogenic properties in several cancers, including lung cancer. Here we examined the effects of a JARID1 family inhibitor against human lung cancer cell lines. 2-4(4-methylphenyl)-1,2-benzisothiazol-3(2H)-one (PBIT) was screened from more than 15000 small molecules as a JARID1 family inhibitor by Sayegh et al. First, we confirmed that PBIT increased H3K4me3 levels in time-dependent and dose-dependent manners in non-small cell lung cancer cell lines. PBIT inhibited cell growth in a dose-dependent manner with the IC50 value of about 10 µM to 30 µM. Next, we detected drug-tolerant persisters (DTPs) from each cell line by exposing them to very high concentration of the drug that is effective to each of the cell lines (gefitinib 10µ for PC9, PC3 and HCC827; AZD9291 5µM for H1975; CDDP 100µM for H226; paclitaxel 500nM for H460, A549 and H1299). DTPs detected were less than 1% in each cell line after 9-day exposure. These small subpopulations showed increased expression of JARID1a and/or JARID1b. Likewise, side population cells, which are considered to contain cancer stem like cells, showed increased expression of JARID1a and decreased H3K4me3 levels. PBIT in combination with anticancer drugs reduced colony formation of DTPs from all cell lines whose DTPs formed colonies. The DTPs showed decreased H3K4me3 levels and increased expression of cancer stem cell marker Oct-4, while these changes were abrogated by subsequent PBIT treatment. These results indicate that PBIT, a JARID1 family inhibitor, recovers sensitivity of drug-resistant lung cancer cells. Citation Format: Shin Ariga, Ichiro Kinoshita, Junko Kikuchi, Yasushi Shimizu, Hirotoshi Dosaka-Akita. JARID1 family inhibitor recovers sensitivity of drug-tolerant subpopulation in lung cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1393.