Antibodies
CP-466722
- 貨號 Cat. No.
- S2245
- 規格 Size
- 5~200mg
- 品牌 Brand
- Selleckchem
Description
CP-466722
Catalog No.S22452 Product Citations
CP-466722 is a potent and reversible ATM inhibitor, does not affect ATR and inhibits PI3K or PIKK family members in cells.
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Distributor Tel: +886-2-2827-1197jimmy@new.abscience.com.tw
Biological Activity
| Description | CP-466722 is a potent and reversible ATM inhibitor, does not affect ATR and inhibits PI3K or PIKK family members in cells. | |||||
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| Targets | ATM | |||||
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| In vitro | In vitro, CP-466722 is identified as a potential inhibitor to decrease the activity of purified ATM kinase to phosphorylate GST-p53(1–101) substrate. In addition, CP-466722 also shows the inhibitory activities against abl and src kinases. [1] In HeLa cells, CP-466722 at doses of 6μM, results in the inhibition in ATM-dependent phosphorylation by reversibly inhibiting ionizing radiation (IR)-induced ATM kinase activity. Besides, ATM-dependent p53 induction is also inhibited by CP-466722 in MCF-7 human breast cancer cells and primary and immortalized diploid human fibroblasts. [1] In response to IR, CP-466722 increased proportion of cells with G2/M DNA content and reduces proportion of cells with G1-phase DNA content in HeLa cells. [1] Transient exposure to CP-466722 for a period of 4 hours sensitizes HeLa cells to IR without affecting cell plating nor cell viability. [1] | |||||
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Protocol(Only for Reference)
Kinase Assay: [1]
| In vitro kinase assays [1] | To screen for small molecule inhibitors of ATM kinase activity, an in vitro kinase assay is adapted, and an ELISA assay develops which measured the phosphorylation status of the ATM downstream target p53. Recombinant GST-p53(1-101) and full-length Flag-tagged ATM & ATR are purified for use in the ELISA and in vitro kinase assays. Briefly, Nunc 96 well Maxisorp plates are coated overnight (4 °C) with 2μg of purified, recombinant GST-p53(1-101) in PBS. All subsequent incubations are performed at room temperature. The plates are washed (0.05%v/v-Tween/PBS) before addition of purified recombinant full-length ATM kinase (30 ng–60 ng) in a final volume of 80μL of reaction buffer (20 mM HEPES, 50 mM NaCl, 10 mM MgCl2, 10 mM MnCl2, 1 mM DTT and 1 μM ATP) in the presence or absence of CP-466722. CP-466722 (10μM) is added to plates in duplicate and the kinase assay is incubated (90 minutes). Plates are washed (0.05%v/v-Tween/PBS), blocked (1hour, 1%w/v-BSA/PBS) and rinsed before anti-Phospho(Ser15)-p53 antibody (1:1000/PBS) is added to the plates and incubated (1hour). To reduce non-specific binding plates are washed (0.05%v/v-Tween/PBS) prior to incubation (1hour) with HRP-conjugated goat anti-rabbit IgG secondary antibody (1:5000/PBS). Secondary antibody that is linked to the phosphorylated GST-p53(1–101) protein is detected with TMB substrate reagent. Plates are developed (15 minutes–30 minutes) and the reaction is stopped (1 M H2SO4 final concentration) before absorbance is determined (λ450nm). CP-466722 that inhibits ATM kinase activity in ELISA assays, are characterized with respect to inhibition of ATM/ATR kinases using in vitro kinase assays. Western blotting using the anti-Phospho(Ser15)-p53 antibody is used as a readout of ATM/ATR inhibition. Extended analysis of CP466722 (10 μM) against a commercially available panel of kinases is performed by Upstate. |
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Cell Assay: [1]
| Cell lines | HeLa and A-T |
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| Concentrations | 0-6 μM |
| Incubation Time | 4 hours |
| Method | HeLa or A-T (GM02052 expressing hTERT) cells are plated in triplicate and incubated for 24 hours. Cells are pre-treated: DMSO, CP466722 or KU55933 prior to IR (0-10Gy). Cells are incubated for 4 hours following IR before media is removed, cells washed (PBS), trypsinsed, counted and re-plated (2000 cells/plate, 10 cm plates) in the absence of drug and incubated for 10 days. Prior to colony counting, cells are washed (PBS), stained (PBS, 0.0037%v/v-formaldehyde, 0.1%w/v-crystal violet), rinsed (dH2O) and dried. Defined populations (>50 cells) are counted as one surviving colony, data are calculated as percentage surviving colonies relative to control plates +/− SE. |
References
Chemical Information
| Molecular Weight (MW) | 349.35 |
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| Formula | C17H15N7O2 |
| CAS No. | 1080622-86-1 |
| Storage | 3 years -20℃Powder |
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| 6 months-80℃in DMSO | |
| Syonnyms | CP466722 |
| Solubility (25°C) * | In vitro | DMSO | 0.28 mg/mL (<1 mM) |
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| Water | <1 mg/mL (<1 mM) | ||
| Ethanol | <1 mg/mL (<1 mM) | ||
| * <1 mg/ml means slightly soluble or insoluble. * Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations. | |||
| Chemical Name | 2-(6,7-dimethoxyquinazolin-4-yl)-5-(pyridin-2-yl)-2H-1,2,4-triazol-3-amine |
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Product Citations (2)
Breast Cancer Stem Cell-Like Cells Are More Sensitive to Ionizing Radiation than Non-Stem Cells: Role of ATM. [Kim SY, et al. PLoS One 2012;7(11):e50423]
PubMed: 23185620
ATM inhibitor KU-55933 increases the TMZ responsiveness of only inherently TMZ sensitive GBM cells. [Nadkarni A, et al. J Neurooncol 2012;110(3):349-57]
PubMed: 23054561
