MED24 deficiency promotes R-loop accumulation and sensitizes to SN-38 in triple-negative breast cancer

Abstract The application of DNA topoisomerase I (TOP1) inhibitors in triple-negative breast cancer (TNBC) still needs to be optimized. The principle of synthetic lethality represents a promising strategy to optimize TNBC treatment using SN-38, a representative TOP1 inhibitor and the active metabolite of irinotecan. MED24, yet little is known about the potential function in breast cancer. Here, we identify MED24 as a key sensitizer of SN-38 through an epigenetic regulator focused CRISPR/Cas9 knockout screen conducted in TNBC cell lines. Loss of MED24 enhanced cellular sensitivity to SN-38 and suppressed colony formation compared with control cells. Silencing of MED24 led to dysregulated transcription, R-loop accumulation, genome instability, resulting in inhibited cell proliferation, particularly when combined with SN-38 treatment. Consistently, depletion of MED24 reduced tumor size and weight in vivo, particularly combined with irinotecan treatment. Moreover, high expression of MED24 was correlated with poorer prognosis of patients with TNBC. Collectively, these findings establish MED24 as a R-loop regulator that enhances SN-38 sensitivity, thereby revealing a previously uncharacterized biological function for MED24 and suggesting a potential therapeutic strategy for TNBC.