人膀胱癌非经典型多药耐受性研究

Study on Non－classical Multidrug Resistance in Human Bladder Cancer

目的：为了建立人膀胱癌耐阿霉素细胞系并研究它们的生物学特性及药物耐受性的机理。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1年，建立一株耐药亚株EJ／DOR，对其生物学特性及耐药机理进行了研究，并应用反转录PCR检测了MDR1、MRP和DNA TopoⅡ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 TopoⅡ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DNA TopoⅡ基因表达低于亲本细胞。结论：细胞内DNA TopoⅡ基因表达下降及MRP基因过表达是EJ／DOR表现为多药耐受性亚型的主要原因，这种非P－gp介导的非经典型多药耐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 its biological characteristics and mechanism of drug resistrance were studied． Methods A human bladder cancer cell line resistant to doxorubicin DOR EJ／DOR has been established in vitro by exposing EJ parent cells to increasing DOR concentration during one year． The cell line was characterized in terms of growth kinetics morphology cross－resistance to other anti－cancer agents and pharmacokinetics of daunorubicin and the MDR1 MRP multidrug resistance associated and DNA topoisomerase Ⅱ TopoⅡ gene expression were assayed using the reverse transcription polymerase chain reaction RT／PCR． Results The EJ／DOR cells were found to be 14．3 times higher resistant to DOR than that of the EJ parent and exhibited cross－resistasnce to DOR derivatives vinca alkaloids and a DNA Topo II－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 that in drug－sensitive EJ cells． DNA Topo Ⅱ gene expression in the EJ／DOR cells was apparently lower than that in EJ parent． Conclusion These results suggest that a decreased cellular level of DNA Topo Ⅱ and an overexpression of MRP gene may be responsible for the expression of a MDR phenotype in the EJ／DOR cells and that such non－Pgp－mediated atypical MDR may develop in bladder cancer treated with chemotherapy including DOR．