Screening Of Multidrug Resistance Reversal Agents And Shikonin Anti-mdr-tumor Activity

Multidrug resistance(MDR) is an obstacle to succeed in chemotherapy, and is a significant problem frustrating and depressing oncologists and cancer patients. There are a variety of mechanisms that result in MDR, including decrease of drug accumulation by tumour cells as a result of the overexpression of transporter proteins such as Pgp, MRP, LRP and MXR, the enhancement of detoxification of the drug in the cell, the alteration of drug targets, and so on. The high level expression of Pgp as an ATP denpendent efflux pump is thought to be the most typical mechanism.In this thesis, over 30 naturally occuring compounds were assayed for their activities in reversing cancer multidrug resistance:1. The MDR cell subline K562/ADR eatablished by exposing to doxorubicin was confirmed to be resistant to a wide spectrum of anticancer durg including daunomycin, doxorubicin, epirubicin, etoposide, homoharringtonine, vincristine, paclitaxel, and mitomycin c. The mechanism underlying the drug resistance acquired by the cell investigated is overexpression of P-glycoprotein. Immunochemical studies using monoclonal antibodies against P-glycoprotein indicated the overexpression of this drug pump, which explained the cause of the drug resistance by this cell line.2. Using the MDR cell model K562/ADR, we carried out primary and secondary screening for the sample library composed of 30 Chinese herb monomers, and found that Andrographolide had the property of reveral MDR to some degree, but its activity was limited. Otherwise, Tanshinone I, Cryptotanshinone, Dihydrotanshinone and Shikonin had cytotoxic activity, and Shikonin was the best one. Honokiol had reversal activity and cytotoxicity, both of which were not strong.3. Study on Shikonin: we found that Shikonin isolated from the roots of the Chinese herb Lithospermum erythrorhizon possessed extraordinary capabilityof anti-MDR- cancer cells. After the cells were treated with shikonin, cells under went apopotosis, as menifested by the degradation of genomic DNA as 'ladder' in agarose gel electrophoresis, hypodiploid peak as analyzed by flow cytometry, morphological change under optical microscope, and the activation of Caspase-3. Particularly important is that shikonin has the same efficacies in killing the MDR cells and the drug sensitive cells. Furthermore, shikonin is much more potent in killing MDR cell K562/ADR than doxorubicin, indicating that the shikonin may have the potential in overcoming cancer MDR. On the other hand, cellular pharmakokinetic studies indicated that shikonin was not the substrate of P-glycoprotein, therefore explaining why shikonin could efficiently kill MDR cancer cells. Shikonin in the cells shikonin was quickly transformed to another compound, which is not pumped out of cells by P-glycoprotein.In conclusion, by drug screening, we discovered that shikonin is an component which is potent in the induction of apoptosis in the cancer cell lines K562 and K562/ADR; Shikonin is more potent than doxorubicin in killing the MDR cell K562/ADR; Shikonin is not the substrate of P-glycoprotein, indicating its potential application in cancer MDR therapy.