| Lung cancer is the most common cancer all over the world,which is also one of the main causes of death among all diseases.Only 15 percent of patients can survive more than five years.Among these,most of lungcancer patients suffer none-small lung cancer.Target therapy is one of the hottest topics in the field of cancer research.Tumorigenesis is quite a complex process with many genes involved.Therefore,single target therapy gradually shows disadvantages.Correspondingly,system biology,which calls for multi-level and multi-target therapy,is accepted by people.The emergence of combinatorial chemistry,high-throughput screening. and landmarks of drug discovery,has accelerated the discovery of novel anti-cancer drugs.Thiazolidinedione derivatives,important kinds of heterocyclic compounds,are important molecular probes for cancer cell lines.We synthesized an optimized combinatorial library,which composed 170 kinds of compounds.The anti-cancer activity was also detected by single compound screen and 10 kinds of active compounds against two kinds of none-small cell lung cancer cell lines(H460 and H460/taxR)were identified.However,the method of in vitro screen that uses one compound at a time utilizes only inadequate information oncombinatorial chemistry.Therefore,the method of identifying active structures by compound mixture screen might better show the advantages of combinatorial chemistry.OBJECTIVES:1.Test the feasibility of the approach that analyzes active compounds through investigation of the cancer cell inhibition by thiazolidinone compound mixtures. 2.Analyze interactions of compounds in the mixtures and effects on the anti-cancer activity of compound mixtures.3.Through the analysis of in vitro screen results,the active mixtures could be selected for future study as multi-therapy drugs.4.Through the study of the inhibition of P-glycoprotein by thiazolidinone derivatives,the way that identifying the drug targets could be paved.METHODS:Based on the theory of combinatorial chemistry and the data collected from single compound screen,we chose 98 kinds of compounds from the thiazolidinone library.According to their structure-activity relationships, we divided these compounds into five groups.1.The morphological changes are observed after treatment by the compound mixtures for 72h.2.The compounds were screened against two non-small cell cancer cell lines(H460 cells and H460/taxR cells)with SRB assay and the possible active compounds are identified.3.Mixtures with potent inhibition were selcted to be screened with normal human fibroblast(NHFB).4.Selected several groups of mixtures to study dose response and GI50s. Molecular probes involved in the mixtures are also being studied,and their GI50s were compared with the GI50s of the corresponding mixtures.5.According to the results of pharmacophore models,three possible P-glycoprotein inhibitors were chosen to be studied against both sensitive cancer cell line H460 and paclipaclitaxol resistant cell line H460/taxR.RESULTS:1.From the observation of the morphology of H460 cells and H460/taxR ells.the effects of mixtures on cell proliferation are quite different. The mixtures with four active compounds show the most potent cancer cell inhibition,while the mixtures with four inactive compounds show the least potent inhibition.2.Through screening against both H460 cells and H460/taxR cells,the percentage growth of two kinds of cell lines could be obtained atter the treatment with compound mixtures for 72h.The H460 cell screen results showed that the mean values of percentage growth are as following:the group of mixtures with four kinds of active compounds was 19%,with four inactive compounds was 65%,and the group with both active compounds and inactive compounds was 42%.Similarly, the results of H460/taxR cell screen showed that the mean values of percentage growth are as following:the group with four kinds of active compounds is 5%,with four kinds of inactive compounds is 72%, and the group with both active compounds and inactive compounds is 35%.3.Two mixtures,m56 and m91,were chosen to be studied on their dose-response characteristics.Compounds 38,45,187,264,274,284. and 344 were also studied and their GI50s were calculated. 4.Compound 25,27 and 48 were mixed with paclitaxol and their inhibition of P-glycoprotein was studied.Compounds 25,27 and 48 didn't show potent inhibition to H460 cells and H460/taxR cells were treated for 72h.However,when mixed with paclitaxol,the inhibition of paclitaxol for H460/taxR cells changed a lot;while the inhibition of paclitaxol for H460 cells changed little,which makes the growth of H460/taxR cells about 20 percent lower.CONCLUSIONS:1.The data of mean percentage growth were ranked and the potent mixtures were selected.The structures of mixtures were analyzed,and the frequencies of each group of potent mixtures were counted.The groups show more often in the potent mixtures(the frequency >50%) were identified as active groups.The results were consistent with single compound screening results.Therefore,the method identifying active compounds from the in vitro screening of compound mixtures was feasible.2.The cancer cell inhibition of mixtures was relevant to the amount of active compounds.The more active compounds in a mixture,the more potent inhibition could be observed.Thus,although there has some interactions existing among the compounds,it is possible to analyze the active structures from the screening results.3.Mixtures with potent inhibition of cancer cells,and having little effects on normal human cells could be selected for the study of the possible multi-target drugs.4.The inhibition of the compound mixtures was not only determined by the most potent compound,but also by the ability of binding to the target.5.Compounds 25,27 and 48 could inhibit P-glycoprotein.
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