Effects Of Sirolimus Combined With Carboplatin On Ovarian Cancer Skov-3 Cells

Research background:Recently years,the incidence of ovarian cancer increases year by year.And with poor prognosis and high mortality rate,the ovarian cancer’s5-year survival rate is only 20%[1].The early stage of ovarian cancerlacks of obvious symptoms,so itsdiagnosis rate is low,two-thirds of the patientsin the treatment is late,unable to do radical surgery.So with the constantly exploring for the treatment,chemotherapy is becoming an indispensable part of the treatment ofovarian cancer.Different from traditional medicines,Platinum-like substancesresulting DNA replication disorder and directly inhibit the division of cancer cells,.This effect is not specific,but the proliferation of tumorcells is more activity than normal cells,so the cancer cells are more sensitive to platinum.A large number of investigations show that platinum has become an important part of chemotherapy regimens in our country,which is widely used in the treatment of ovarian cancer、small cell lung cancer and other malignant tumor,and nearly 80% of the patients with ovarian canceris sensitive toplatinum.Carboplatin as the second generation of platinum family of anti-tumor drugs,water solubility and stability are better than cisplatin,.Carboplatin as the second generation of platinum family of anti-tumor drugs,ts water solubility and stability are greatly superior to cisplatin.Under the condition of same curative effect,itsadverse reactions are less than cisplatin,so in the clinical treatment of some cancer,carboplatin ingradually replace cisplatin,becomes an important chemotherapeutic drugs.But with the extensive application of platinum,its drug resistance is becoming an problome which can not be ignored.Sirolimus is a macrolide antibiotic,not only has an antibacterial effect,but also can play the role of immunosuppressive agents.For chronic kidney disease,rapamycin can reduce proteinuria,and reduce the interstitial fibrosis of renal tubule.In the field of cardiovascular disease,the use of rapamycin drug-coated stents in the treatment of coronary artery restenosis also has obtained the remarkable result.In the 90 s,scientists found that rapamycin have antitumor effect.At the year of 1984,Eng C P and his colleaguescarried outfurther experiments which could prove that rapamycin can inhibit the growth of tumor cells.Experiments aiso showed that compared with other antitumor drugs such as fluorouracil,rapamycin can inhibit the growth of any stage of the tumor cells with lowest toxicity.Recentiy years,rapamycin is used in the treatment of kidney cancer,breast cancer step by step,and the related experiments also prove that it can also improve the effect of other drugs.Objective:To monitor the proliferation,apoptosis and cell cycle arrest of the ovarian cancer Skov-3 cell under the drug Sirolimus and Carboplatin,which added alone or in combination.The confirms the inhibitory effect of sirolimus on ovarian cancer cells and its role in improving the efficacy of carboplatin chemotherapy,so as to provide new ideas for the treatment of ovarian cancer.Methods:(1)MTT assay was used to detect the effects of sirolimus and carboplatinon the proliferation of ovarian cancer Skov-3 cells.(2)Annexin V / PI double staining was used to detect the effects of sirolimus and carboplatin on the apoptosis of ovarian cancer Skov-3 cells.(3)Flow cytometry was used to detect the effects of sirolimus and carboplatin on the cell cycle distribution of ovarian cancer cells.(4)RT-PCR was respectivelyused to detect the expression of Bcl-2 / Bax gene in sirolimus and carboplatin.Results:(1)MTT assay showed that the proliferation of ovarian cancer Skov-3 cells was inhibited by sirolimus,carboplatin,and the combination of the two groups.The inhibitory rates were in the order of sirolimus combined with carboplatin group,carboplatin group and sirolimus group.The inhibitory effect was positively correlated with the time of action.The differences were statistically significant.(P<0.05).(2)Annexin V / PI double staining showed that the use of sirolimus and carboplatin could promote the apoptosis of ovarian cancer Skov-3 cells,and the effect was as follows: sirolimus carboplatin,Combined effect group,carboplatin group and sirolimus group,the effect effect and the time of action were positively correlated,the differences were statistically significant.(P<0.05)(3)Flow cytometry(FCM)PI staining showed that the ratio of G1 phase cells in ovarian cancer Skov-3 cells was increased by single and simultaneous sirolimus and carboplatin,and the effect was from strong to weak: Sirolimus carboplatin combination group,carboplatin group,sirolimus group,the effect and the role of time was positively correlated,the difference was statistically significant(P <0.05).(4)The results of RT-PCR showed that the expression of Bax gene and the expression of Bcl-2 gene were inhibited by using sirolimus and carboplatin in the Skov-3 cells.The effect was in the order of sirolimus carboplatin combined with carboplatin group and sirolimus group,and the effect was positively correlated with the time of action.The differences were statistically significant.(P <0.05).Conclusion:Sirolimus and carboplatin can inhibit the proliferation of ovarian cancer Skov-3 cells,induce apoptosis,block the progress of cell cycle and decrease the expression of Bcl-2 / Bax,and when the two are combined Anti-tumor effect is best.