Establishment Of Human Prostate Cancer Abiraterone-resistant Cell Line And The Study Of Drug-resisitance Mechanism

Objective To establish the advanced resistant cell model of abiraterone in the prostate cancer,describe its biological characteristics and explore the potential molecular mechanism of the acquired drug resistance of abiraterone,and provide reference for reversing the drug resistance to abiraterone and develop new drugs.Methods DU145 / AA were developed from human prostate cancer parental cell line DU145,by exposing DU145 to gradually increasing concentration of abiraterone in vitro.DU145 / AA and DU145 / S were used as the research object: observed the morphological characteristics of the two cell lines under the optical microscope;The resistance of DU145 / AA to DU145 / S was detected by CCK-8 method.;CCK-8 method was used to detect the drug sensitivity of both cells to abiraterone and draw the cell growth curve,the cell population doubling time was calculated at the same time.The cell cycle of the two cell lines was detected by flow cytometry.The expression of drug resistance candidate genes in two cell lines was analyzed by fluorescence quantitative PCR.Result 1.After 5 months of induction,we successfully established the human prostate cancer abiraterone resistant cell lines DU145 / AA,DU145/AA and DU145 are homologous.2.Cytology morphology: campare to DU145/S,the DU145/AA has stronger refraction,its vacuolization and grainy became more obviously,has more disordered arrangement.3.CCK-8 method showed that the IC50 of DU145/S and DU145/AA to abiraterone were(9.091±0.422)u M and(35.95±2.57)u M,DU145/AA resistance index RI = 3.955,the difference was statistically significant(P<0.01).The growth curve of DU145/S and DU145/AA were drew by CCK-8 method.The population doubling time of DU145/S and DU145/AA was(37.12 ± 2.25)h and(44.76±2.42)h respectively,DU145/AA was 1.2 times longer than DU145/S and the growth rate was significantly slower(P<0.05).4.The results of flow cytometry showed that DU145/AA had a significant increase in G0/G1 cells(P<0.01),G2/M phase cells were decreased(P<0.05),S phase cells were decreased(P<0.01).5.The expression of MDR1 was up-regulated in DU145/AA(P<0.05),and there was no significant difference in the expression of MRP in the multidrug-resistant gene(P<0.05);The expression of Vimentin in EMT-related gene was not significantly different(P<0.01).The expression of E-cadherin and N-cadherin was up-regulated(P<0.01).The expression of NSE in neuroendocrine-related genes was not significantly different,but Syn and Cg A expression was significantly up-regulated(P<0.01),There were no significant differences in the expression of AR-FL,AR-V7,ARv567 es and GR;In the related enzymes of androgen synthesis pathway,the expression of CYP17A1,AKR1C3 and HSD17B3 were significantly up-regulated(P<0.01),but there was no significant difference in the expression of CYP11A1,SRD5A1 and HSD3B2.Conclusion 1.Compared with DU145/S,DU145 / AA has a mild resistance to abiraterone,and its cell morphology and biological characteristics have changed significantly.2.The process of acquired resistance to abiraterone is complicated,involving multiple genes and multiple access.The up-regulation of neuroendocrine-related genes and enzymes of androgen synthesis pathway may play a more important role in the process.