PURPOSE: This study investigates the synergistic therapeutic potential of a novel combination of the repurposed drug simvastatin with irinotecan chemotherapy towards glioblastoma (GBM) and the underlying molecular mechanisms. METHODS: In vitro efficacy of simvastatin and irinotecan alone and in combination against diverse GBM lines (U251MG, G34, SB28) was assessed using mechanistically distinct cell viability assays. RNA-Sequencing was performed to uncover the top pathways and genes affected by these drugs, followed by validation of promising pathways (TGF-β signaling and cell death) using targeted phosphoproteomics and in vitro genetic manipulation and functional assays. RESULTS: We observed robust in vitro synergy at nanomolar concentrations between simvastatin and irinotecan across diverse GBM lines. Notably, irinotecan alone and in combination with simvastatin reduced mRNA expression of TGF-β family members. Targeted phosphoproteomics and functional experiments further showed significant inhibition of TGF-β signaling with both treatment types. Additionally, a role for apoptosis and enrichment of caspase-independent cell death pathways (autophagy, ferroptosis) as well as immunological (interferons, complement, inflammatory responses, TNF-α) and oncogenic (K-RAS/ERK) signaling pathways were observed with the combination treatment. CONCLUSIONS: Besides the first detailed demonstration of a robust synergy between simvastatin and irinotecan against GBM lines, this study shows for the first time that both irinotecan and the combination treatment converge on inhibition of TGF-β signaling. This is notable given the lack of TGF-β inhibitors in the clinic. Collectively, this study provides preclinical data suggesting this novel drug combination be tested in patients with GBM and TGF-β driven cancers.