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Research Area
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Description
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Cancer |
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Biological Activity
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Description
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PHA-793887 is a novel and potent inhibitor of CDK2, CDK5 and CDK7 with IC50 of 8 nM, 5 nM and 10 nM, respectively. |
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Targets
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CDK2 |
CDK5 |
CDK7 |
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IC50 |
8 nM |
5 nM |
10 nM [1] |
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In Vitro
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[1] |
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In Vivo
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PHA-793887 (10–30 mg/kg) shows good efficacy in the human ovarian A2780, colon HCT-116, and pancreatic BX-PC3 carcinoma xenograft models. [1]PHA-793887 (20 mg/kg) is effective in xenograft models of K562 and HL60 cells, primary leukemic disseminated model, and a high-burden disseminated ALL-2 model derived from a relapsed Philadelphia-positive acute lymphoid leukemia patient. [2] |
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Clinical Trials
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A Phase I clinical trial of PHA-793887 for advanced/metastatic solid tumors has been terminated. |
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Features
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Multi-CDKs inhibitor. |
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Protocol
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Kinase Assay
[3]
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| CDK Kinase Assay |
The biochemical activity of compounds is determined by incubation with specific enzymes and substrates, followed by quantitation of the phosphorylated product. PHA-793887 (1.5 nM–10 μM) is incubated for 30?90 min at room temperature in the presence of ATP/33P-γ-ATP mix, substrate, and the specific enzyme (0.7?100 nM) in a final volume of 30 μL of kinase buffer, using 96 U bottom plates. After incubation, the reaction is stopped and the phosphorylated substrate is separated from nonincorporated radioactive ATP using SPA beads, Dowex resin, or Multiscreen phosphocellulose filter as follows: (1) For SPA Assays. The reaction is stopped by the addition of 100 μL of PBS + 32 mM EDTA + 0.1% Triton X-100 + 500 μM ATP, containing 1 mg of streptavidin-coated SPA beads. After 20 min of incubation for substrate capture, 100 μL of the reaction mixture is transferred into Optiplate 96-well plates containing 100 μL of 5 M CsCl, left to stand for 4 hours to allow stratification of beads to the top of the plate, and counted using TopCount to measure substrate-incorporated phosphate. (2) For Dowex Resin Assay. An amount of 150 μL of resin/formate, pH 3.00, is added to stop the reaction and capture unreacted 33P-γ-ATP, separating it from the phosphorylated substrate in solution. After 60 min of rest, 50 μL of supernatant is transferred to Optiplate 96-well plates. After the additon of 150 μL of Microscint 40, the radioactivity is counted in the TopCount. (3) For Multiscreen Assay. The reaction is stopped with the addition of 10 μL of EDTA (150 mM). An amount of 100 μL is transferred to a MultiScreen plate to allow substrate binding to phosphocellulose filter. Plates are then washed three times with 100 μL of H2PO4 (75 mM) filtered by a MultiScreen filtration system, and dried. After the additon of 100 μL of Microscint 0, radioactivity is counted in the TopCount.IC50 values are obtained by nonlinear regression analysis. |
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Cell Assay
[1]
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| Cell Lines |
A2780 cells |
| Concentrations |
0.1 nM – 1 μM, dissolved in DMSO |
| Incubation Time |
72 hours |
| Methods |
Cells are seeded into 96- or 384-wells plates at final concentration ranging from 1×104 to 3×104 per cm2. After 24 hours, cells are treated using serial dilution of PHA-793887. At 72 hours after the treatment, the amount of cells are evaluated using the CellTiter-Glo assay. IC50 values are calculated using a sygmoidal fitting. |
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Animal Study
[1]
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| Animal Models |
Mouse xenograft models of human ovarian A2780, colon HCT-116 and pancreatic BX-PC3 carcinoma |
| Formulation |
Dissolved in 5% dextrose solution |
| Doses |
10, 20, and 30 mg/kg |
| Administration |
Intravenous injection once daily |
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References |
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[1] Brasca MG, et al. Bioorg Med Chem, 2010, 18(5), 1844-1853.
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[2] Alzani R, et al. Exp Hematol, 2010, 38(4), 259-269.
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[3] Pevarello P, et al. J Med Chem, 2004, 47(13), 3367-3380.
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