In cellular assays,INCB 024360 (INCB024360) selectively inhibits human IDO1 with IC50 values of approximately 10 nM, demonstrating little activity against other related enzymes such as IDO2 or tryptophan 2,3-dioxygenase (TDO). INCB 024360 also exhibits significant activity toward mouse IDO1, with an IC50 value of 52.4 nM±15.7 nM, in a similar assay using mouse IDO1-transfected HEK293/MSR cells.
Female Balb/c mice bearing CT26 tumors are treated orally twice daily for 12 d with INCB 024360 (INCB024360) at 100 mg/kg. INCB 024360 suppresses kynurenine equivalently in plasma, tumors, and lymph nodes. In naïve C57BL/6 mice, 50 mg/kg INCB024360 decreases plasma kynurenine levels within 1 hour and those levels stay at least 50% suppressed through the 8-hour time course.
To determine INCB 024360 activity against IDO in recombinant cells, HEK293/MSR cells are transiently transfected with full-length human or mouse IDO1, or mouse IDO2 cDNA, with Transit-293 transfection reagent or Lipofectamine 2000 reagents. INCB 024360 at different concentrations is added to the recovered transfected cells seeded at 2×104 cells per well in a 96-well plate (200 μL/well). The cells are incubated for 2 days, and kyn in the supernatants is measured as described in the HeLa cell assay. The tryptophan 2,3-dioxygenase (TDO) assay is performed similarly with HEK293/MSR cells transfected with a human TDO expression vector[1].
MCE has not independently confirmed the accuracy of these methods. They are for reference only.
