Nagane M, Asai A, Shibui S, Nomura K, Kuchino Y
Department of Neurosurgery, National Cancer Center Hospital, National Cancer Center Research Institute, Tokyo, Japan.
Neurosurgery. 1997 Aug;41(2):434-40; discussion 440-1. doi: 10.1097/00006123-199708000-00021.
A derivative of chloroethylnitrosoureas, 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosourea (ACNU), is a drug of choice for the chemotherapy of human malignant brain tumors. However, the cytocidal effect of ACNU is effectively repressed through repair of ACNU-mediated deoxyribonucleic acid lesions by O6-methylguanine-deoxyribonucleic acid methyltransferase (MGMT). Because a variety of human tumors, including brain tumors, contain high levels of MGMT activity, we investigated the effect of antisense ribonucleic acid (RNA) complementary to MGMT messenger RNA on ACNU resistance in tumor cells.
We established a stable ACNU-resistant clone, C6AR, from the rat glioma cell line C6 exposed to a stepwise increasing concentration of ACNU. We transfected a plasmid deoxyribonucleic acid-encoding antisense MGMT RNA under the control of the human metallothionein promoter into C6AR cells and determined the effect of the antisense RNA on ACNU resistance of tumor cells by a colony-forming efficiency assay.
C6AR cells expressed abundant MGMT messenger RNA, although the transcription level of the MGMT gene in parental C6 cells was below the lower limits of detection under the same assay conditions. ACNU resistance of C6AR cells was significantly repressed by transfected gene-dependent antisense MGMT RNA expression that resulted in decreased survival of the tumor cells.
ACNU resistance resulting from the expression of MGMT in rat glioma cells is significantly overcome by the expression of antisense MGMT RNA. This result suggests that the antisense MGMT RNA system might be a useful strategy for overcoming ACNU resistance in the treatment of intractable malignant gliomas.
氯乙基亚硝脲的衍生物1-(4-氨基-2-甲基-5-嘧啶基)甲基-3-(2-氯乙基)-3-亚硝基脲（ACNU）是治疗人类恶性脑肿瘤的化疗首选药物。然而，O6-甲基鸟嘌呤-脱氧核糖核酸甲基转移酶（MGMT）对ACNU介导的脱氧核糖核酸损伤的修复可有效抑制ACNU的杀细胞作用。由于包括脑肿瘤在内的多种人类肿瘤都含有高水平的MGMT活性，我们研究了与MGMT信使核糖核酸互补的反义核糖核酸（RNA）对肿瘤细胞中ACNU耐药性的影响。
我们从暴露于逐步增加浓度ACNU的大鼠胶质瘤细胞系C6中建立了一个稳定的ACNU耐药克隆C6AR。我们将在人金属硫蛋白启动子控制下编码反义MGMT RNA的质粒脱氧核糖核酸转染到C6AR细胞中，并通过集落形成效率测定法确定反义RNA对肿瘤细胞ACNU耐药性的影响。
C6AR细胞表达丰富的MGMT信使核糖核酸，尽管在相同检测条件下，亲代C6细胞中MGMT基因的转录水平低于检测下限。转染基因依赖性反义MGMT RNA表达可显著抑制C6AR细胞对ACNU的耐药性，导致肿瘤细胞存活率降低。
反义MGMT RNA的表达可显著克服大鼠胶质瘤细胞中因MGMT表达而产生的ACNU耐药性。这一结果表明，反义MGMT RNA系统可能是克服难治性恶性胶质瘤治疗中ACNU耐药性的一种有用策略。
