Devimistat (CPI-613)

388, 59

BxPC-3 and H460 cells are injected s.c. into the dorsal flank of CD1 nu/nu mice.

<=25 mg/kg

In vitro, CPI-613 produces the selective toxicity against several tumor cell lines including H460 human lung cancer cells and Saos-2 human sarcoma cells with EC50 of 120 uM and 120 uM, respectively. CPI-613 disrupts H460 cancer cell mitochondrial metabolism including inhibition of PDH complex activity and loss of mitochondrial membrane potential in a time- and drug dose-dependent fashion. In addition, CPI-613 (240 uM) also induces both apoptotic and non-apoptotic cell death in H460 human lung cancer and Saos-2 human sarcoma cells. [1]

E1alpha phosphorylation, Cells are seeded and grown overnight. Five dishes per test group are treated with CPI-613or solven. Cells are lysed in situ with 150 uL lysis buffer A [455 uL Zoom 2D protein solubilizer 1, 2.5uL 1M Tris base, 5uL 100X protease inhibitor cocktail, 5uL 2M DTT] and lysates from all five dishes are pooled in a 1.5 mL microfuge tube and sonicated on ice for 15 passes at 50% power. After a 10-minute incubation at room temperature, 2.5uL of dimethylacrylamide (DMA) are added and lysates are incubated for an additional 10 minutes. 5uL of 2 M DTT are added to neutralize excess DMA. Lysates are centrifuged for 15 minutes and the supernatant is recovered. Then, 40uL of lysate are mixed with 0.8uL pH 3-10 ampholytes, 0.75 uL 2 M DTT and brought up to 150uL with Zoom 2D protein solubilizer 1. 150uL of sample is loaded into IPG runner, and pH 3-10NL IPG strips are added. Strips are soaked overnight at room temperature. A step protocol is used for isoelectric focusing (250V, 20min., 450V 15min, 750 V 15 min 2000V 30 minutes). For the second dimension, strips are treated for 15 minutes in 1X loading buffer, followed by 15 minutes in 1X loading buffer plus 160 mM iodoacetatic acid. Strips are electrophoresed on NuPAGE 4-12% Bis Tris ZOOM gels. Proteins are blotted onto PVDF 4.5um membranes. Serines 232, 293 and 300 are targets of the three well-characterized PDK phosphorylation events controlling E1 activity. Equal amounts of protein from treated and mock-treated cells are loaded on gels and western transfers are probed with anti-E1alpha Ab as an internal matching control or one of the three anti-phospho-E1alpha Abs.

Devimistat (CPI-613), a lipoate analog, inhibits mitochondrial enzymes pyruvate dehydrogenase (PDH) and α-ketoglutarate dehydrogenase in NCI-H460 cell line, disrupts tumor cell mitochondrial metabolism. CPI-613 induces apoptosis in pancreatic cancer cells. Phase 2.