Updated project metadata. Mitochondria support cancer cell proliferation by producing ATP and supplying metabolic intermediates needed for macromolecule synthesis. Voltage-Dependent Anion Channels (VDACs), particularly isoform 1 (VDAC1), regulate mitochondrial metabolite flux and serve as metabolic gatekeepers. We recently identified SC18, a small-molecule inhibitor targeting the NADH-binding pocket of VDAC1, which reduces mitochondrial membrane potential, NADH, and ATP levels, and synergizes with FDA-approved hepatocarcinoma drugs. In this study, we used structural biology, electrophysiology, proteomics, and metabolic assays to show that SC18 binds cooperatively within the NADH-binding pocket, enhancing VDAC closure in the presence of NADH. This impairs mitochondrial uptake of key substrates and initiates compensatory metabolic changes, ultimately disrupting mitochondrial bioenergetics and cancer cell viability. Our findings provide a mechanistic basis for targeting VDAC1 to disrupt cancer metabolism.