人膀胱癌非P-糖蛋白介导的多重抗药性

Study on non-Pgp-mediated pleiotropic resistance in a human bladder carcinoma cell line

目的 :建立人膀胱癌耐阿霉素细胞系及研究它们的生物学特性及药物耐受性的机制。 方法 :采用人膀胱癌细胞系 EJ,经递增阿霉素剂量的方法 ,历时一年 ,建立一株耐药亚株 EJ/ DOR,对其生物学特性及耐药机制进行了研究 ,并应用反转录 PCR检测了 MDR1、MRP和 DNA Topo 基因的表达。 结果 :EJ/ DOR对阿霉素的相对耐受度较亲本细胞提高了 14.3倍 ;对蒽环类、长春花属生物碱及 DNA Topo 靶制剂足叶乙甙有明显的交叉耐药性 ,但对顺铂、丝裂霉素无明显的交叉耐受性 ;耐药细胞对柔红霉素的细胞内聚集量显著减少 ;EJ/ DOR并不表达 MDR1基因 ,而 MRP基因过表达 ,但细胞内 DNATopo 基因表达低于亲本细胞。 结论 :细胞内 DNA Topo 基因表达下降及 MRP基因过表达是 EJ/ DOR表现为多耐药受性亚型的主要原因 ,这种非 P- gp介导的非经典型多耐药受性细胞为寻求包括阿霉素在内的化疗方案提供了良好的实验模型。

Objective To establish a doxorubicin resistant human bladder cancer cell line,and to study its biological characteristics and mechanism of drug resistance. Methods A human bladder carcinoma cell line resistant to doxorubicin(DOR),EJ/DOR has been established in vitro by exposing EJ parent cells to stepwise increase DOR concentrations over a period of one year.The cell line was characterized in terms of growth kinetics,morphology,cross resistance to other anticancerous agents and pharmacokinetics of daunorubicin and analyzed the MDR1, MRP(multidrug resistance associated), and DNA topoisomeraseⅡ(TopoⅡ) gene expression using the reverse transcription polymerase chain reaction assay(RT/PCR). Results The EJ/DOR cells were found to be 14.3 times more resistant to DOR than the EJ parent,and exhibited cross resistance to DOR derivatives,vinca alkaloids and a DNA TopoⅡ targeting agent,etoposide. The intracellular accumulation of daunorubicin was markedly decreased in the EJ/DOR cells in comparison with the parent cells. EJ/DOR cells did not express the MDR1 gene, but the expression levels of MRP were markedly higher than in drug sensitive EJ cells. DNA TopoⅡ gene expression in the EJ/DOR cells was apparently lower than in the EJ parent. Conclusions These results suggest that a decreased cellular level of DNA TopoⅡ and an overexpression of MRP gene may be responsible for the expression of an MDR phenotype in the EJ/DOR cells and that such non Pgp mediated, atypical MDR may develop in bladder carcinoma treated with chemotherapy including DOR.