Ganoderma Lucidum, Dong Ling Grass Extract Reverse Mcf 7 / Adm Cells Multi-drug Resistance Of Breast Cancer Research

Objective:Chemotherapy plays an important role in clinical therapy of human breast cancer. However, multidrug resistance (MDR) of tumor impact the effect of chemotherapy, even have become the main cause of chemotherapy failure. Therefore, searching MDR reversal agent to enhance the efficacy of chemotherapy is urgently required problem in cancer treatment. At present, there have many researches on multidrug resistance reversal agents. There are many MDR agents have been found, such as calcium channel blocker, calcium channel antagonist, immune-modulators and so on. However, most of the reversal agents are difficult to apply in clinical because of side effects. Chinese traditional drug has a long history, the toxic and side effects were relatively smaller and the cost was lower. Therefore, development of safe and effective MDR reversal agent from Chinese traditional drug is a unique advantage. Chinese medicine Ganoderma Lucidum and Rabdosia rubescens are two kinds of famous Chinese herbal medicine, which are used for the treatment of cancer for long times, due to their low toxic and good anti-tumor efficacy. In our research, we focused on the MDR reversal effect of Ganoderma Lucidum and Rabdosia rubescens extracts on human breast cancer MCF-7/ADM cells, the mechanism of its MDR reversing effect is also explored. It can explain the mechanisms of Ganoderma lucidum and Rabdosia rubescens as a supplementary drug in cancer treatment, and provide a theoretical basis for its further development.Methods:The cross-resistance assays of drug human breast cancer MCF-7/ADM cells to adriamycin, cisplatin and vincristine were determined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) method. Apply MTT method to measure direct cell toxicity effect on MCF-7/ADM cells by different active chemical extracts of Ganoderma Lucidum and Rabdosia rubescens, in order to choose their non-cytotoxic doses. To observe the reverse effect of different active chemical extracts of Ganoderma Lucidum and Rabdosia rubescens which has screened for nontoxicant concentration to MCF-7/ADM cells by MTT assay. The MCF-7/ADM intracellular accumulation of adriamycin and Rh123and expression of P-gp were examined by flow cytometry. The gene expression of MDR1mRNA was determined by RT-PCR. Results:(1) When MTT assay was performed, it was found that MCF-7/ADM cells were resistant not only to adriamycin but also to multiple anticancer drugs, such as Cisplatin (DDP) and Vincristine (VCR), and drug resistance reversal fold was19.23,2.14and2.40, separately.(2) Cell toxicity experiment results showed that Ganoderma Lucidum had non-cytotoxic on MCF-7/ADM cells. Inhibitory rates of40μg/mL Ganoderma Lucidum and20μg/mL Rabdosia rubescens for MCF-7/ADM cells was under20%, so this concentration was regarded as non-cytotoxic dose.(3) When adriamycin was used in combination with different active chemical extracts of Ganoderma Lucidum and Rabdosia rubescens at non-cytotoxic concentration on MCF-7/ADM cells, they can reverse MDR of MCF-7/ADM cells in dose-dependent manner, and drug resistance reversal fold is1.78,3.56,2.11,3.47,1.87,4.02,2.16, and4.60, respectively.(4) Adriamycin and Rh123accumulation experiment indicated that different active chemical extracts of Ganoderma Lucidum and Rabdosia rubescens could concentration-dependent enhance the accumulation of intracellular adriamycin and Rh123, and inhibit the efflux of Rh123. In contrast, they had a little affect the accumulation of intracellular adriamycin and Rh123on sensitive MCF-7cells.(5) Different active chemical extracts of Ganoderma Lucidum and Rabdosia rubescens could down regulate expression levels of P-gp in MCF-7/ADM cells by flow cytometry.(6) Different active chemical extracts of Ganoderma Lucidum and Rabdosia rubescens could down regulate the over-expression of MDR1in a concentration-dependent manner by RT-PCR.(7) The HPLC fingerprint chromatograms of Ganoderma Lucidum’s, ethyl acetate and butanol sites are very similar, and it is twelve characteristic peaks. On the basis of ergosterol standards, this is peak12in the fingerprint chromatograms. Quantitative assay results showed that the ergosterol in Ganoderma Lucidum’s, ethyl acetate sites1.70%and in butanol sites1.91%. The HPLC fingerprint chromatograms of Rabdosia rubescens’s ethyl acetate and chloroform sites are also very similar, and it is eleven characteristic peaks. On the basis of Oridonin standards, this is peak6in the fingerprint chromatograms. Quantitative assay results showed that the Oridonin in Rabdosia rubescens’s ethyl acetate sites15.46%and in chloroform sites13.84%. Conclusions:The extracts from Ganoderma Lucidum and Rabdosia rubescens had the revesal effect significantly, and its mechanism may be relevant with inhibition of the function of P-gp and expression of MDR1gene. Within a certain drug concentration, they might reverse multidrug-resistance by inhibiting the efflux function of P-gp and increased the drug concentration in MCF-7/ADM cells. The effective componnents of Ganoderma Lucidum and Rabdosiae Rubescentis could reverse multidrug resistance is not only ergosterol and oridonin, there are other components.