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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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OSU-03012 is a potent inhibitor of recombinant PDK-1 with IC50 of 5 μM. |
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Targets
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PDK-1 |
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IC50 |
5 μM [1] |
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In Vitro
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OSU-03012 induces apoptotic death in PC-3 cells with IC50 of 5 μM and reduces the activity of immunoprecipitated p70S6K. OSU-03012 completely suppress cell growth in a diverse range of tumor cell lines at concentrations of 3–5 μm, as compared with the concentration of at least 50 μm required for celecoxib. [1] OSU-03012 promotes cell killing to a greater extent in glioma cells than in nontransformed astrocytes. OSU-03012 causes a dose-dependent induction of cell death that is not altered by p53 mutation, expression of ERBB1 VIII, or loss of phosphatase and tensin function due to a homolog deletion on chromosome 10. OSU-03012 and ionizing radiation cause an additive, caspase-independent elevation in cell killing. OSU-03012 lethality as a single agent or when combined with signaling modulators is not modified in cells lacking expression of BIM or of BAX/BAK. OSU-03012 promotes the release of cathepsin B from the lysosomal compartment and that of AIF from mitochondria. The lethality of OSU-03012 is attenuated in protein kinase R-like endoplasmic reticulum kinase-/- cells, which correlated with the reduced cleavage of BID and suppression of cathepsin B and AIF release into the cytosol. [2] OSU-03012 inhibits thyroid cancer cell (NPA, WRO, and ARO cells) proliferation, migration and induces apoptosis, which results in an increase of cells in the S phase without an increase of cells in G2. OSU-03012 is an ATP-competitive inhibitor of PAK activity and suppresses the phosphorylation of AKT in thyroid cancer cells. [3] OSU-03012 inhibits cell growth of hepatocellular carcinoma cell lines including Huh7, Hep3B and HepG2 cells with IC50 values below 1 μM. OSU-03012 does not suppress PDK1 or AKT activity or induce cellular apoptosis but induces autophagy in Huh7 cells. Moreover, accumulation of reactive oxygen species (ROS) is detected after OSU-03012 treatment. [4] A recent study shows that OSU-03012 could enhance the susceptibility of (Bcr)-Abl mutant cell lines to imatinib-induced apoptosis. [5] |
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In Vivo
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OSU-03012 suppresses tumor growth by 57.59% and increases cleaved LC3 in Huh7 tumor xenografts at 200 mg/kg. [4] OSU-03012 remarkably decreases expression of EGFR protein in the tumors by 48% compared with vehicle controls and also prevents YB-1 from binding to the EGFR promoter in MDA-MB-435/LCC6 xenografts. [6] OSU-03012 is well tolerated and inhibits the growth of HMS-97 schwannoma xenografts by 55% after oral administration. [7] |
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Clinical Trials
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Features
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OSU-03012 is a derivative of celecoxib and shows ten-fold greater antitumor activity than celecoxib but it lacks its COX2 inhibitory activity. |
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Protocol
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Kinase Assay
[1]
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| PDK-1 Kinase Assay |
This in vitro assay is performed using a PDK-1 kinase assay kit. This cell-free assay is based on the ability of recombinant PDK-1, in the presence of DMSO vehicle or OSU-03012, to activate its downstream serum- and glucocorticoid-regulated kinase which, in turn, phosphorylates the Akt/serum- and glucocorticoid-regulated kinase-specific peptide substrate RPRAATF with [γ-32P]ATP. The 32P-phosphorylated peptide substrate is then separated from the residual [γ-32P]-ATP by using P81 phosphocellulose paper and quantitated in a scintillation counter after three washes with 0.75% phosphoric acid. |
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Cell Assay
[1]
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| Cell Lines |
PC-3 cells |
| Concentrations |
0-10 μM |
| Incubation Time |
~72 hours |
| Methods |
The effect of OSU-03012 on PC-3 cell viability is assessed by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay in six replicates. Cells are grown in 10% FBS- supplemented RPMI 1640 in 96-well, flat-bottomed plates for 24 hours. They are exposed to various concentrations of OSU-03012 (0-10 μM) dissolved in DMSO (final concentration ≤0.1%) in 1% serum-containing RPMI 1640 for different time intervals (~72 hours). Controls receive DMSO vehicle at a concentration equal to that in OSU-03012-treated cells. The medium is removed and replaced by 200 μL of 0.5 mg/mL 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide in 10% FBS-containing RPMI 1640. The cells are incubated in the CO2 incubator at 37 °C for 2 hours. Supernatants are removed from the wells, and the reduced 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide dye is solubilized in 200 μL DMSO per well. Absorbance at 570 nm is determined by using a plate reader. |
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Animal Study
[4]
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| Animal Models |
Huh7 tumor xenografts in male BALB/c nude mice |
| Formulation |
Dissolved in 0.5% methylcellulose, 0.1% Tween 80 |
| Doses |
100-200 mg/kg |
| Administration |
Daily by gavage |
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References |
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[1] Zhu J, et al. Cancer Res, 2004, 64(12), 4309-4318.
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[2] Yacoub A, et al. Mol Pharmacol, 2006, 70(2), 589-603.
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[3] Porchia LM, et al. Mol Pharmacol, 2007, 72(5), 1124-1131.
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[4] Gao M, et al. Cancer Res, 2008, 68(22), 9348-9357.
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[5] Tseng PH, et al. Blood, 2005, 105(10), 4021-4027.
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[6] To K, et al. Mol Pharmacol, 2007, 72(3), 641-652.
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[7] Lee TX, et al. Eur J Cancer, 2009, 45(9), 1709-1720.
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