Inhibition Of MDR By SiRNA In The Xenografts Of Drug-Resistant Ovarian Carcinoma In Vivo

Objective: Ovarian cancer is currently the most lethal gynecologic malignancy. Clinically, Multidrug resistance (MDR) is one of the major causes for chemotherapeutic treatment failure in ovarian cancer patients. Overexpression of P-glycoprotein (P-gp), the MDR1 gene product, confers MDR to ovarian cancer cells. RNA interference (RNAi) is a conserved biological response to double-stranded RNA, which results in sequence-specific gene silencing. We designed this experiment to study the ability of small interference RNA (siRNA) mediated RNAi to inhibit the expressions of MDR1 gene and P-gp accordingly, then to reverse its MDR in the xenografts of drug-resistant ovarian carcinoma in nude mice.Methods: Thirty nude mice with xenografts of drug-resistant human epithelial ovarian cancer cells OVCAR/AR were established subcutaneously and divided randomly into three groups, the pSN/mdr1a pSN/mdr1b and pSN/e groups. After the formation of tumor knot (0.5cm), MDR1 mRNA specific siRNA expressing plasmids pSN/mdr1a and pSN/mdr1b (0.5mg/kg) were injected into the knots. Chemotherapeutics Taxol was injected veinously with a single dose of 20mg/kg the next day. Empty vector without inserting any target gene (pSN/e) was transfected as a control. All mice were sacrificed 21 days after chemotherapy. The volumes of the tumors were measured and the tumor growth rates were calculated. The distribution of p-glycoprotein (P-gp)was detected by immunohistochemistry. In addition, MDR1 mRNA expression level of tumor tissues was quantified by real time reverse transcription polymerase chain reaction (real time RT-PCR), and Flow cytometry (FCM) was performed to assess the expression level of P-gp.Results: All nude mice were successfully inoculated and all induced tumors developed. Immunohistochemistry demonstated that the distribution levels of p-glycoprotein (P-gp) in pSN/mdr1a and pSN/mdr1b groups was lower than pSN/e group (P<0.05). The growth rates of the xenografts of the pSN/mdr1a and pSN/mdr1b groups were 164.5±15.8 % and 159.6±16.8 %, which was significantly smaller than that of the pSN/e group (P<0.01). The MDR1 mRNA and P-gp levels in the two experimental groups were both reduced significantly compared with the control group(P<0.01). There is no significant difference between the growth rates and the expressions of MDR1 mRNA and P-gp in these two siRNA-treatment groups (P>0.05).Conclusions: Our study demonstrated the proof that vector-based siRNA can be applied to activate RNAi-mediated degradation of the endogenous MDR1 mRNA in vivo, which decreases the expression of P-gp, thus to increase the chemotherapy sensitivity and inhibit the gowth of drug-resistant ovarian carcinoma xenografts.