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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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PF-03814735 is a novel, potent and reversible inhibitor of both Aurora A and Aurora B with IC50 of 0.8 nM and 5 nM, respectively. |
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Targets
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Aurora A |
Aurora B |
Flt1 |
FAK |
TrkA |
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IC50 |
0.8 nM |
5 nM |
10 nM |
22 nM |
30 nM [1] |
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In Vitro
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In intact cells, the inhibitory activity of PF-03814735 on the Aurora1 and Aurora2 kinases reduces levels of phospho-Aurora1 (Thr 232, a sensitive marker of Aurora1 activity, with IC50 ~ 20 nM), phosphohistone H3 (with IC50 ~ 50 nM), and phospho-Aurora2 (with IC50 ~150 nM). PF-03814735 produces a block in cytokinesis, resulting in inhibition of cell proliferation and the formation of polyploid multinucleated cells. [1] A recent research indicates small cell lung cancer (SCLC) and, to a lesser extent, colon cancer lines are very sensitive to PF-03814735. The status of the Myc gene family and retinoblastoma pathway members significantly correlates with the efficacy of PF-03814735. [1] |
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In Vivo
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Once-daily oral dosing of ≥20 mg/kg of PF-03814735 for 10 days to mice bearing HCT-116 xenografts resulted in statistically significant and dose-dependent tumor growth inhibition of ≥50% relative to vehicle-treated mice. The inhibition is associated with a reduction in phosphorylated histone H3 levels. Significant single-agent antitumor efficacy is observed in five additional xenograft tumor models, including A2780 ovarian carcinoma, MDA-MB-231 breast carcinoma, colo-205 and SW620 colorectal carcinomas, and HL-60 acute promyelocytic leukemia. [1] In vivo experiments with two SCLC xenograft models confirms the sensitivity of Myc gene-driven models to PF-03814735 and a possible schedule dependence of MYC/c-Myc-driven tumors. [1] |
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Clinical Trials
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Features
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Combination Therapy
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Description
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The combination of PF-03814735 and docetaxel in xenograft mouse tumor models shows additive tumor growth inhibition. [1] |
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Protocol
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Kinase Assay
[1]
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| Recombinant Kinase Assays |
Aurora1 and Aurora2 proteins are produced as full-length His-tag recombinant proteins expressed in insect cells. For the Aurora2 kinase assay, phosphorylation of the substrate peptide by recombinant Aurora2 protein is assessed by a Z'-LYTE assay at 3 to 300 μM ATP and various concentrations of PF-03814735 over 60 minutes, at a substrate peptide concentration of 2 μM (biotinylated LRRWSLG, ×4). Phosphorylation is linear over this time for all conditions. For the Aurora1 kinase assay, phosphorylation of the substrate peptide by recombinant Aurora1 protein is assessed by a scintillation proximity assay in a 96-well plate format in which the incorporation of 33P into the peptide substrate (biotinylated LRRWSLG, ×4) is measured by capturing the peptide on a streptavidin scintillation proximity assay bead. |
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Cell Assay
[1]
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| Cell Lines |
HCT-116 cell |
| Concentrations |
300 nM |
| Incubation Time |
4, 8, 12, 24, 48, or 72 hours |
| Methods |
Cell lines are grown in appropriate media and evaluated after 48 h of exposure to either PF-03814735 or vehicle, followed by cell number determination in a Coulter Counter. Proliferation (as measured by an increase in cell number) is expressed as a percent of untreated controls. To evaluate the PF-03814735 exposure time required for antiproliferative activity, HL-60 cell cultures are cultured in RPMI medium supplemented with 15% heat-inactivated fetal bovine serum and exposed to various PF-03814735 concentrations for 4, 8, 12, 24, and 48 hours, followed by a washout step and incubation with growth media without PF-03814735 for the remainder of the 72-h assay period. Continuous exposure to PF-03814735 for 72 hours is also evaluated. Cell counts are determined by a Coulter Counter. |
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Animal Study
[1]
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| Animal Models |
HCT116 tumors are implanted s.c. on the right flank of nude mice. |
| Formulation |
PF-03814735 is formulated as a solution in cremophor EL [cremophor/ethanol/0.9% saline (12.5%/12.5%/75%)]. |
| Doses |
10, 20, 30 mg/kg |
| Administration |
Administered orally |
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References |
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[1] Jani JP, et al, Mol Cancer Ther, 2010, 9(4), 883-894.
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[2] Hook KE, et al, Mol Cancer Ther, 2012, 11(3), 710-719.
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