The Influence Of Rapamycin In Combine With Cisplatin On The Growth Of Human Ovarian Cancer Cell

Background and objectiveOvarian cancer is hard to detect so its therapeutic effect is very poor.Thereforeovarian cancer is the leading cause of death among gynecological neoplasms, withapproximately23,000new cases diagnosed per year and over15,000fatalities in theUnited States alone. The mainly treatment of ovarian cancer is cytoreductive surgery andplatin-based chemotherapy after surgery.However the patients of ovarian cancer have apoor prognosis due to the side effect of chemotherapy and drug resistance. The5-yearsurvival rate for all ovarian cancer patients remins at less than40%for the past30years.So looking for a new and effective method is needed for the treatment of ovarian cancer.Studies have showed that disorder of the cell signal pathway is the main reason oftumor development and drug resistance, and activation of mTOR pathway is closelyassociated with tumor cells proliferation without control,tumor recurrence and drug andradiation resistance.The disorder of mTOR pathway appears in many malignancies,suchas breast cancer,leukemia,gastrointestinal stromal tumor,liver cancer and lungcancer.Mammalian target of rapamycin(mTOR) is the key molecular of thispathway,which is related to the protein synthesis and the cell cycle regulation.ThereforemTOR-targeted therapy has shown promising role in the tumor targeting therapy.Rapamycin is a macrocyclic lactone product from the soil bacteria Streptomyces hygrscopicus. It was isolate and identified as an antifungal agent.In1990s,it wasapproved by FDA as an immunosuppressive agent.Recently rapamycin has been tested aspotential anti-cancer drug as the inhibitor of mTOR.It incombination with traditionalchemotherapy drugs may improve the ability of apoptosis and strengthen the tumor cellsfor chemotherapy sensitivity.Cisplatin is a widely used chemotherapeutic agent thatexerts its cytotoxic effects by disrupting the DNA structure in cells through the formationof intrastrand adducts and interstrand cross-link,in order to inhibite the DNAreplication.It has proven to be one of the most clinically active agents for the treatment ofa variety of solid tumors,including ovarian cancer.Rapamycin can inhibite the sythesis ofDNA-damage repair proteins,and accelerate the apoptosis of cancer cells.So we expectthat rapamycin can enhance cisplatin-induced anti-proliferation and apoptosis,andstrengthen the tumor cells for cisplatin-sensitive.This study has choosen ovarian cancer cells to observe the effect of rapamycinincombination with cisplatin on the progression,apoptosis and cell cycle distribution,todisscuse the effect of target-drugs incombine with traditional chemotherapy drugs and itsmolecular mechanism.In order to explore a more effective therapy for ovarian cancer.MethodsSKOV3cells was cultured in vitro,MTT assay was applied to examine rapamycinalone or in combine with cisplatin of the inhibition rate on cell proliferation for24h、48h、72h,respectively;Meantime FCM assay was used to examine the apopotosis and the cellcycle distribution.The expression of PTEN、AKT、mTOR、p-mTOR、S6Kand Bcl-2weredeteceted by Western blot.ResultsThe results of MTT revealed that rapamycin alone could inhibit the growth ofSKOV3cells in time dependent manners, and reached the highest inhibition rate at72h,and the comparisions among different groups had statistical significance(P＜0.05）,and the joint effect of the two drugs had a significantly higher inhibition rate thanthe other groups,the differences had statistical significance(P＜0.01）.CI＜1,suggestedthat the two medicine combine applician have obvious synergic effect.The results ofFCM showed that rapamycin or cisplatin alone could induce apoptosis of SKOV3cells intime dependent manners, and reached the highest effect at72h;The rate of appotosiswas lower in the group of rapamycin administered alone for24h,and had no statistical significance(P＞0.05）between the control group.The apoptosis of rapamycin in combinewith cisplatin was higher than the other groups. Rapamycin administered alone for24hcould cause cell cycle arrest at G1phase,which had no statistical significance(P＞0.05）between the control group, S and G2phase cell population dreased, G1phase cellpopulation gradually increased as time passed by,and the jont effect of the two drugswas more stronger than any other drugs used alone,the comparisons among differentgroups had statistical significance(P＜0.05）.Westernblot showed that Rapamycin couldupregulate the expression of PTEN and decrease the expression of mTOR、p-mTOR andBcl-2,AKT was increased when treated with rapamycin for24h,then decreased as timewent on(48h,72h).The expression of S6K had no significant variation.The comparisonsbetween experiment groups and control groups had statistical significance(P＜0.05）. Theexpression of the above proteins between the group of rapamycin administered alone andthe two medicine administered in combination had no difference.Conclusion1. Rapamycin alone could have anti-cancer effect,and in combine with cisplatinshowed obvious synergistic effect,could enhance cisplatin-inducedapoptosis,strengthen the tumor cells for cisplatin-sensitive.2. Rapamycin alone or in combine with cisplatin application had time-dependentmanners, and reached the highest effect on72h.3.Rapamycin could play its anti-cancer and chemosensitive effect throughblocking the mTOR pathway, and decreasing the expression of Bcl-2.