SUMMARYAchaete-scute homologue 1 (ASCL1) is a transcriptional factor related to small cell lung cancer (SCLC) development. Because ASCL1 inhibition suppresses cancer development, targeting ASCL1 would be effective for the treatment of SCLC. We focused on the natural product rocaglamide, a synthetic compound, and identified indole rocaglamide (−)-4k as a strong inhibitor of ASCL1. Elucidation of the underlying mechanism revealed that translation of ASCL1 is inhibited through the interaction of (−)-4k with eukaryotic initiation factor 4A and the 5’UTR of ASCL1 mRNA like a molecular glue. Also, (−)-4k induced S and G2/M arrest in SCLC cells via suppression of POLA2, which is downstream of ASCL1. Deletion of POLA2 caused DNA replication stress, which led phosphorylated check-point kinases to phosphorylate CDK1, causing S and G2/M arrest in SCLC cells. This is the first report of such a strong ASCL1 inhibitor and elucidation of cytotoxicity mechanism after inhibition of ASCL1. This study found that inhibiting the translation of ASCL1 using rocaglamide is effective for suppressing SCLC.