An Mps1/TTK inhibitor
NMS-P715 is an inhibitor of the spindle assembly checkpoint kinase Mps1/TTK (IC50 = 0.182 ?M).1 It is selective for Mps1/TTK over a panel of 59 additional kinases (IC50s = >5 ?M for all). NMS-P715 (1 ?M) accelerates mitosis, reducing the length of mitosis by approximately 3-fold, in U2OS cells. It induces aneuploidy and apoptosis in A2780 ovarian cancer cells when used at a concentration of 1 ?M, as well as inhibits proliferation in a panel of 127 cancer cell lines, including colon, breast, renal, and melanoma cells, with IC50 values ranging from 0.192 to 10 ?M. NMS-P715 (90 mg/kg per day) reduces tumor growth in an A2780 mouse xenograft model.
1.Colombo, R., Caldarelli, M., Mennecozzi, M., et al.Targeting the mitotic checkpoint for cancer therapy with NMS-P715, an inhibitor of MPS1 kinaseCancer Res.70(24)10255-10264(2010)
Kinase experiment: | The potency of the compound towards MPS1 and 60 additional kinases belonging to kinase selectivity screening (KSS) panel is determined using either a strong anion exchanger based assay or P81 Multiscreen plate. MPS1 activity is measured using 5 nM of MPS1 recombinant protein in 50 mM HEPES pH 7.5, 2.5 mM MgCl2, 1 mM MnCl2, 1 mM DTT, 3 μM NaVO3, 2 mM β-glycerophosphate, 0.2 mg/mL BSA, 200 μM P38-βtide substrate-peptide (KRQADEEMTGYVATRWYRAE) and 8 μM ATP with 1.5 nM 33P-γ-ATP. The assay is run in a robotized format, 10 serial 1:3 compounds dilutions (including NMS-P715, from 30 μM to 1.5 nM) are tested and IC50 determined[1]. |
Cell experiment: | Cells lines are seeded in 384 well-plates in the appropiate complete medium and treated with compounds (NMS-P715, etc.) dissolved in 0.1% DMSO 24 hours after seeding. The cells are incubated at 37°C and 5% CO2 and after 72 hours the plates are processed using CellTiter-Glo assay. Inhibitory activity is evaluated comparing treated versus control data using Assay Explorer software. IC50 of proliferation is calculated using sigmoidal interpolation curve fitting. Activity Ratio is calculated as the ratio of the single cell line IC50 and the IC50 average of all the cell lines tested[1]. |
Animal experiment: | Mice[1]Athymic nu-nu mice, 5-6 weeks of age (20-22 g) are used in the assay. A2780 ovary carcinoma and A375 melanoma cells are transplanted s.c. into female nu-nu mice. Mice bearing a palpable tumor (100-200 mm3) are selected and randomized into control and treated groups. Treatment starts one day after randomization. NMS-P715 is typically administered by oral administration at doses of 90-100 mg/kg daily for more than seven days. Each group includs 8 animals. Tumor dimension is measured regularly by calipers during the experiments and tumor mass is calculated[1]. |
参考文献: [1]. Colombo R, et al. Targeting the mitotic checkpoint for cancer therapy with NMS-P715, an inhibitor of MPS1 kinase. Cancer Res. 2010 Dec 15;70(24):10255-64. | |
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