Devimistat (CPI-613)

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 μM and 120 μM, 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 μM) also induces both apoptotic and non-apoptotic cell death in H460 human lung cancer and Saos-2 human sarcoma cells. ^[1]

CPI-613 (25 mg/kg) has potent anticancer activity in a human tumor xenograft model of of a pancreatic tumor cell (BxPC-3). Similarly, CPI-613 (10 mg/kg) also produces significant tumor growth inhibition of H460 human non-small cell lung carcinoma in mouse model. Besides, CPI-613 produces little or no side-effect toxicity in expected therapeutic dose ranges in large animal models and has the maximum tolerated dose of 100 mg/kg in mice. ^[1]

• E1α 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 μL lysis buffer A [455 μL Zoom 2D protein solubilizer 1, 2.5μL 1M Tris base, 5μL 100X protease inhibitor cocktail; 5μL 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, 2.5μL of dimethylacrylamide (DMA) are added and lysates are incubated for an additional 10 minutes. 5μL of 2 M DTT are added to neutralize excess DMA. Lysates are centrifuged for 15 minutes and the supernatant is recovered. Then, 40μL of lysate are mixed with 0.8μL pH 3-10 ampholytes, 0.75 μL 2 M DTT and brought up to 150μL with Zoom 2D protein solubilizer 1. 150μL 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 4.5μm 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-E1α Ab as an internal matching control or one of the three anti-phospho-E1α Abs.

• Cell lines

  HN6 cells

• Concentrations

  100 μM

• Incubation Time

  24 h

• Method

  Cells were treated with or without 100 μM CPI-613 for 24 h.

• Animal Models

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

• Dosages

  ≤25 mg/kg

• Administration

  Administered via i.p.