Abstract / Summary
Abstract Galunisertib (LY2157299), a selective TGF‑βRI/ALK5 inhibitor with limited single‑agent Phase II efficacy, was used as a pharmacological probe to ask whether sustained Galunisertib exposure reorganizes cellular dependencies toward an OXPHOS-associated state. A genome‑wide CRISPR‑Cas9 loss‑of‑function chemogenomic screen was performed in Galunisertib‑treated NALM‑6 pre‑B leukemia cells and analyzed with CRANKS scoring, Hallmark gene set enrichment analysis, STRING network mapping, subcellular compartment annotation, and MitoCarta3.0/MitoPathways3.0 cross‑referencing. Oxidative phosphorylation (OXPHOS) was the only Hallmark gene set significantly enriched after correction for multiple testing, and mitochondrial compartments, mitochondrial gene expression and mitochondrial translation were enriched among genes whose loss improved fitness under treatment. Prioritized hits resolved into a layered architecture in which a chromatin-associated transcriptional module, anchored by the high-confidence sensitizer TCF4, is coupled to post-transcriptional regulation of mRNA metabolism and to genes maintaining membrane and ionic homeostasis, converging on mitochondrial gene expression and translation as the terminal layer. Resistance-associated genes defined two routes out of this state: altered chromatin remodeling, and loss of mitochondrial translation or iron-sulfur cluster biogenesis. These dependencies derive from a single high-dose chemogenomic screen and await confirmation by individual gene perturbation or by direct metabolic measurement. Within that limit, the screen defines a reorganization of cellular dependencies under sustained Galunisertib exposure and nominates chromatin, post-transcriptional and OXPHOS-directed combinations as priority candidates for validation.