Voltage dependent anion channels (VDAC) control the flux of most anionic respiratory substrates, ATP, ADP, and small cations, crossing the outer mitochondrial membrane. VDAC closure contributes to the partial suppression of mitochondrial metabolism that favors the Warburg phenotype of cancer cells. Recently, it has been shown that NADH binds to a specific pocket in the inner surface of VDAC1, also conserved in VDAC2 and 3, closing the channel. We hypothesized that binding of small molecules to the NADH pocket, maintain VDAC in an open configuration by preventing closure induced by NADH and possible other endogenous regulators. We screened in silico, the South Carolina Compound Collection SC ³ (~100,000 proprietary molecules), using shape-based queries of the NADH binding region of VDAC. After molecular docking of selected compounds, we physically screened candidates using mitochondrial membrane potential (ΔΨm), as an overall readout of mitochondrial metabolism. We identified SC18, as the most potent compound. SC18 bound to VDAC1, as assessed by a thermal shift assay. Short-term treatment with SC18 decreased ΔΨm in SNU-449 and HepG2 human hepatocarcinoma cells. Mitochondrial depolarization was similar in wild type, VDAC1/2, 1/3, and 2/3 double KO HepG2 cells indicating that the effect of SC18 was not VDAC isoform-dependent. In addition, SC18 decreased mitochondrial NADH and cellular ATP production; and increased basal respiration. Long-term exposure to SC18 , decreased cell proliferation as determined by wound-healing and cell viability assays. In summary, SC18 is a novel VDAC-targeting small molecule that induces mitochondrial dysfunction and inhibits cell proliferation.

The small molecule SC18 regulates mitochondrial metabolism and promotes mitochondrial dysfunction by blocking the access of endogenous regulators to the NADH-binding site of VDAC. Sustained mitochondrial dysfunction decreases cell proliferation and promotes cell death.