Abstract 4351: Clinical indenoisoquinolines strongly inhibit the global <i>MYC</i> transcriptional pathway

MYC, one of the most validated driver cancer oncogenes, is overexpressed in a majority of human cancers. However, the MYC protein is intrinsically disordered and considered “undruggable”. The MYC gene promoter forms G-quadruplex, a globular non-B DNA secondary structure. MYC G-quadruplex (MycG4) is a transcription repressor that silences MYC expression upon stabilization by small molecules. Targeting MycG4 for transcriptional inhibition is a promising approach for cancer therapeutics. Indenoisoquinolines were designed as topoisomerase I inhibitors with better physicochemical and biological properties than the clinical camptothecin drugs, with three compounds in phase I/II clinical trials. Recently, we found that indenoisoquinolines can bind and stabilize MycG4 and downregulate MYC expression in cancer cells, which significantly correlates with anticancer activities. Herein, we investigate the effects of the three clinical indenoisoquinolines on MYC downregulation and MYC gene pathways, with comparison to other well-known G4-ligands and the topoisomerase I inhibitor camptothecin (CPT). The clinical indenoisoquinolines strongly bind and stabilize MycG4, whereas CPT shows little binding and stabilization. We assessed global changes in gene expression by RNA sequencing in the MYC-dependent B-lymphoma Raji cells. We treated the tumor cells with the three clinical indenoisoquinolines (LMPs), other G4-ligands, and CPT, at subtoxic doses across various timepoints. Our enrichment analysis on gene ontology and pathway identified the primary cellular responses and pathways altered with MYC expression modulation. We also evaluated the treatment-induced effects on all hallmark biological pathways from the Molecular Signature Database using Gene Set Enrichment Analysis. The RNA-seq results indicate that the MYC pathway is the major cellular response of the clinical LMP compounds. We further examined the treatment-induced suppression of MYC-regulated genes and observed significant and durable inhibition of MYC-regulated gene pathways. Notably, we found that the clinical LMPs, especially LMP400, exhibited significantly stronger MYC suppression compared to other G4-ligands. We speculate that dual MYC-targeting and topoisomerase I inhibition is critical to the exceptional MYC inhibition and anticancer efficacy of indenoisoquinolines. In conclusion, we demonstrate that the clinical indenoisoquinolines effectively and persistently inhibit the global MYC transcriptional pathway, serving as a major mechanism of action. Because MYC is a common and essential oncogene, our study provides critical mechanistic understanding and a molecular basis for the future clinical developments and therapeutic applications of indenoisoquinolines in MYC-dependent cancers. Citation Format: Yichen Han, Sheng Liu, Mark Cushman, Jun Wan, Danzhou Yang. Clinical indenoisoquinolines strongly inhibit the global MYC transcriptional pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4351.