RNA干扰抑制FANCD2基因对多药耐药骨髓瘤细胞株药物敏感性的影响

Effects of inhibition of FANCD2 expression by RNA interference on chemotherapy sensitivity of multidrug resistant human multiple myeloma cell line

目的:探讨FA/BRCA(fanconi anemia/BRCA)中siRNA干扰对多药耐药骨髓瘤细胞株的影响。方法:体外培养多发性骨髓瘤细胞RPMI-8226细胞系,通过相应渐增浓度的马法兰药物处理,选择性获得多药耐药的子代肿瘤细胞株RPMI-8226/R细胞。MTT法检测RPMI-8226和RPMI-8226/R细胞马法兰、ADM、VCR、CTX、Ara-C、VP-16及DDP的敏感性。siRNA干扰抑制FA/BRCA途径FANCD2蛋白表达,观察多发性骨髓瘤多药耐药细胞株对烷化剂药物敏感性的变化。结果:成功构建针对FANCD2基因的siRNA,将其转染多药耐药骨髓瘤细胞株,可以抑制细胞中FANCD2基因表达,转染细胞较转染前细胞FANCD2基因表达降低,对烷化剂的敏感性升高。结论:人多发性骨髓瘤多药耐药细胞株RPMI-8226/R其多药耐药性的产生可能与FA/BRCA途径中FANCD2表达增强有关,可通过抑制FANCD2表达而逆转细胞的耐药性达到增强烷化剂对耐药肿瘤细胞的增殖抑制及诱导凋亡作用。沉默FANCD2表达可以提高多药耐药MM细胞对烷化剂的敏感性,而RNA干扰是一种沉默FANCD2表达的理想方法,因此它可能是一种有潜力的耐药MM辅助治疗新方法。

Objective：To explore effects of siRNA interference of FANCD2 on muhidrug resistant human multiple myeloma cell line. Methods ： Multiple myeloma cell line RPMI - 8226 was cultured in vitro, and the progency multi- drug resistant cell line RPMI - 8226/R was selected by stepwise exposure of parental RPMI - 8226 cells to increasing concentrations of melphalan. The MTI＂ assay was used to detect the sensitivity to chemotherapeutic agents, such as melphalan,ADM,VCR,CTX,Ara- C, VP- 16 and DDP in the RPMI- 8226 and RPMI- 8226/R sublines. Using siRNA interference suppression the FANCD2 protein expression of the FA/BRCA pathway, and observed changes of the drug sensitivity in MM multidrug resistant cell line. Results：We successfully constructed the siRNA directed at the FANCD2 gene, and transfected the muhidrug resistant myeloma cell lines, which can inhibit the FANCI）2 gene ex- pression in cells. The expression levels of the FANCD2 gene of transfected cells was decreased compared with the be- fore - transfection cells, and the sensitivity to the alkylating agent （CTX） was also elevated in transfected cells. Con- clusion： The RNA interference to FANCD2 gene is a good method to silence expression of FANCD2 in human multiple myeloma cells. Inhibiting the expression of FANCD2 can enhance sensitivity to alkylating agents in multidrug resistant human multiple myeloma cells. It may be an available therapeutic method in the treatment of muhidrug resistant multi- ple myeloma.