| 包装: | 250mg |
| 市场价: | 347元 |
Topoisomerase I cleavable complex assay | Topoisomerase I was isolated from calf thymus and was devoid of topoisomerase II. All reactions were carried out in 10 mL volumes of reaction buffer (50 mM Tris-HCl, pH 7.5, 100 mM KCl, 0.5 mM EDTA, and 30 μg/mL BSA) in microtiter plates. Camptothecin was dissolved in DMSO at 10 mg/mL and serially diluted in 96-well microtiter plates to which the 32P end-labeled pBR322 DNA and topoisomerase enzyme were added. The reaction mixture was incubated at room temperature for 30 mins and then the reaction stopped by adding 2 mL of a mixture of sodium dodecyl sulfate and proteinase K (1.6% and 0.14 mg/mL final concentrations, respectively). The plates were heated at 50℃ for 30 mins, 10 mL of standard stop mixture containing 0.45 N NaOH was added in order to generate single-stranded DNA, and the samples were electrophoresed in 1.5% agarose gels in TBE buffer. Gels were blotted on nitrocellulose paper, dried, and exposed to X-ray film. The units of cleavage were calculated from the autoradiographs and plotted against the log drug concentration. The IC50 values were then obtained. |
Cell lines | HCT116 and RKO colorectal cancer (CRC) cells |
Preparation method | Limited solubility in DMSO. General tips for obtaining a higher concentration: Please warm the tube at 37℃ for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below -20℃ for several months. |
Reaction Conditions | 20 and 50 nM; 72 hrs |
Applications | The low doses of Camptothecin for HCT116 and RKO CRC cells were 20 nM and 50 nM, respectively, both of which induced the least detectable cell death. Low-dose Camptothecin induced autophagy via AMPK-TSC2-mTOR pathway and premature senescence by ATM-Chk2-p53-p21 pathway. |
Animal models | Nude mice bearing xenografts of CASE, SW48, DOY, SPA, and CLO cells |
Dosage form | 0 ~ 8 mg/kg; i.m. or i.v.; twice a week |
Applications | In mice xenografts of various tumors, including colon, lung, breast, stomach and ovary tumors, Camptothecin treatment (8 mg/kg) exhibited complete growth inhibition and regression. |
Other notes | Please test the solubility of all compounds indoor, and the actual solubility may slightly differ with the theoretical value. This is caused by an experimental system error and it is normal. |
| 文献引用 | |
| 产品描述 | Camptothecin is a selective inhibitor of topoisomerase I with IC50 value of 679 nM [1]. Camptothecin could induce cell death in SMMC-7721, MCF-7, and HCT-116 tumor cells. Camptothecin has been reported to induce autophagy via AMPK-TSC2-mTOR pathway, at the mean time, induce premature senescence by ATM-Chk2-p53-p21 pathway [2]. Studies indicated that camptothecin had anti-tumor activity in (non-small cell lung cancer) NSCLC xenografts. Camptothecin -induced DNA damage has revealed to cause phosphorylation of H2AX (γH2AX). Previous studies have also demonstrated to reduce the expression of Top1 in cancer cells treated with camptothecins [3]. References: |
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