CRISPR Screen Identifies HDAC3 as a Novel Radiosensitizing Target in Small Cell Lung Cancer

Small cell lung cancer (SCLC) is an aggressive malignancy, with most patients presenting with prognostically poor extensive-stage disease. Limited progress in standard care stresses the urgent need for novel therapies. Radiotherapy offers some survival benefit for select SCLC patients but could be enhanced with radiosensitizers. Here, we identify HDAC3 as a novel radiosensitizing target in SCLC using a CRISPR knockout screen and demonstrate its efficacy and mechanism. SBC5 cells were transduced with a custom EpiDrug sgRNA library and treated with ionizing radiation (IR). Cells were collected and sequenced at multiple timepoints to identify radiosensitizing genes. To validate this target, HDAC3 functional loss was generated by genetic knockdown (KD) and RGFP966 inhibition in several SCLC cell lines. HDAC3 KD and RGFP966 inhibition increased radiosensitivity, as shown by cell viability (dose[1]modification factor [DMF]10 = 1.14-1.69) and clonogenic assays (DMF10 = 1.16-1.41). We evaluated changes in chromatin accessibility by ATAC-seq, and IR-induced DNA damage and repair using γH2AX foci detection and double-strand break (DSB) repair assays. Cells with HDAC3 functional loss demonstrated increased chromatin accessibility which was associated with more IR-induced DNA DSBs, and impaired DSB repair function leading to persisting DNA damage. In SCLC xenograft models, HDAC3 KD or RGFP966, combined with IR, enhanced radiosensitivity and achieved significant tumor growth inhibition. Taken together, we identified HDAC3 as a novel radiosensitizing target in SCLC. Its functional loss increases the generation and persistence of IR-induced DNA DSBs, effectively sensitizing SCLC cell lines and xenografts to IR, providing a potential radiosensitization strategy to treat SCLC.