The Preliminary Research On Reversion Effects Of Mifepristone On Human Ovarian Carcinoma Cells (COC₁/DDR) Resistant To Cisplatin And Its Mechanism

BackgroudMultidrug resistance (MDR) is a formidable roadblock to the effective treatment of ovarian cancer by conventional chemotherapy and affects the five-year survival rate of ovarian cancer. MDR is due to the changes in gene, proteins, lipids, apoptosis, sigal transduction and the ability of cancer cells to reduce the absorbability of drug, lower intracellular drug concentration, relieve the toxin of drug and so on. Evidence suggests a correlation between cellular drug resistance and alterations in ceramide metabolism. Ceramide glycosylation, through glucosylceramide synthase (GCS), allows cellular escape from ceramide-induced programmed cell death. This glycosylation even confer cancer cell resistance to cytotoxic anticancer agents. The MDR cell has an increased capacity to convert ceramide to glucosylceramide. So some scientists have pay attention to the GCS in MDR Currently.Mifepristone (RU486), the widely used drug for the treatment of obstetric and gynecologic diseases, blocked ceramide glycosylation and increased breast cancer cell sensitivity to Adriamycin. It is only a kind of progesterone antagonist validated by clinic trial cosmically. Not only one research suggests RU486 reverses MDR through multi-mechanism, with more effec and less side effect. Presently we have not found a MDR modifying agent safe and efficacious to gynecologic cancer, so RU486 could be used in ovarian cancer chemotherapy as a resistance modifying agent if it everse resistance of ovarian cancer cells in vitro and in vivo. Objectives and SignicationBased on molecular biology and cell biology, the author investigated the expressing of GCS (glucosylceramide synthase) mRNA and protein in ovarian cisplatin-resiatant cell line COCi/DDP and its parent ciaplatin-sensitive cell line COC₁ The author also attempts to investigate the reversioneffect of a MDR modulator mifepristone (RU486) on COC₁/DDP in vitro andin vivo, explore the changes in the expressing of GCS mRNA and protein.SThe aim of the research is to elucidate the importance and affection of GCSin MDR, provide possibility of mifepristone used as resistance modifyingagent, and to provide new ideals and clues to seek new effective methods fortherapy and reversing the MDR in ovarian cancer as well.Methodsl.MDR cell line COC₁/DDP was treated with RU486 in different conce-ntration for 24h. The alterations of chemosensitivity to cisplatin (DDP) wereevaluated by MTT (the tetrazolium dye) assay. A xenografted cisplatinresistant mouse of ovarian cancer was developed. The change of tumorvolumes and its pathological appearances were compared before and aftertreatment.2.GCS mRNA in cell line COC₁, COC₁/DDP (before and after the treatmentof RU486) and xenografted were isolated, changes of GCS expressing werestudied by RT-PCR. The expresses of GCS protein was detected by Westerblot.Results1.RU486 had no effect on the viability of COCi/DDP at the consentrationbetween 1.25μmol/L and 10μmol/LP = 0.951⁰.395), but could increase thesensitivity of COC₁/DDP to DDP. These modulations of MDR were contentdependent.2.In vivo, the greater inhibitory rates of mifepristone+DDP regimen (70.48%)were shown when compared with DDP alone regimen (21.55%) and RU486regimen (8.98%) (P<0.01).3.In the combined group, pathological appearances were significant differentas compared with that of the control group. Morphological analysis showedthat the majority of the xenografted cells revealed necrosis and denaturalizat-ion, endoplast enlarging like bubbles, chromatin condensing and so on.4.Compared to the parent cells, the resistant cells(COC₁/DDP)have higherGCS expressing at the mRNA and protein level. The level of GCS mRNAwere 1.37.33 in COC₁/DDP, higher than that in COC, 0.28?.08 (P<0.01). 5.RU486 inhibited the expresses of GCS mRNA and protein.Treated by 1.25, 5 and 10μmol/L RU486, the expressing of GCS mRNA were 0.96?.14 (P<0.05), 0.51?.10(P<0.01 )and 0.32?.04(P<0.0...