CCT245737 

Cytotoxicity is determined as the drug concentration that gives 50% inhibition of tumor cell proliferation (GI₅₀) using a 96 h Sulforhodamine B (SRB) assay. Inhibition of intracellular CHK1 activity is measured using a cell based ELISA for the abrogation of an VP-16 induced G2 checkpoint (mitosis induction assay, MIA). The IC₅₀ for G2 checkpoint abrogation (MIA) is determined in the presence of nocodazole using UCN01 as a positive control. The activity index (AI) is used as a measure of the compounds ability to induce mitosis relative to its toxicity (i.e., ratio of MIA IC₅₀: 96 h SRB GI₅₀). Routine potentiation studies are carried out using a fixed concentration (GI₅₀) of either LY 188011 or SN38 in combination with a range of CCT245737 concentrations to determine the combination GI₅₀ of CCT245737. The ability of CCT245737 to enhance LY 188011 or SN38 cell killing is expressed as a potentiation index (PI) equal to the ratio of the GI₅₀ for CCT245737 alone versus the combination GI₅₀ for CCT245737. PI values > 1 indicate potentiation of the genotoxic activity. In addition, a series of experiments is carried out using fixed, non- or minimally toxic concentrations of CCT245737 (≤GI₂₀) with a range of different concentrations of LY 188011 or SN38 to determine the extent to which CCT245737 enhances drug cytotoxicity compared with the genotoxic agent alone, i.e. conventional PI (ratio GI₅₀ genotoxic alone: GI₅₀ genotoxic combined with non-toxic CCT245737 concentration, Con PI)^[2].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Human HT29 colorectal carcinoma cells are injected s.c into the flanks of female NCr athymic mice 6-8 weeks of age. Dosing commenced 5 days after transplantation when tumors reach a mean diameter of 5.5 mm. LY 188011 (100 mg/kg i.v.) is dosed in saline on days 0, 7 and 14 and compounds 4 (CCT245737) and 41 (150 mg/kg p.o.) in 10% DMSO 20% PEG 400, 5% Tween 80, 65% water, 24 and 48 h after each dose of LY 188011. Tumors are measured and body weights recorded three times weekly. Animals are culled on an individual basis when tumors reach a predetermined humane endpoint (mean diameter <15 mm)^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Osborne JD, et al. Multiparameter Lead Optimization to Give an Oral Checkpoint Kinase 1 (CHK1) Inhibitor Clinical Candidate: (R)-5-((4-((Morpholin-2-ylmethyl)amino)-5-(trifluoromethyl)pyridin-2-yl)amino)pyrazine-2-carbonitrile (CCT245737). J Med Chem. 201  [Content Brief]

  [2]. Walton MI, et al. The clinical development candidate CCT245737 is an orally active CHK1 inhibitor with preclinical activity in RAS mutant NSCLC and Eμ-MYC driven B-cell lymphoma.  [Content Brief]