Molecular Biological Analysis for Augmented Brain Tumor Chemotherapy

恶性脑肿瘤化疗的一个主要问题是存在对化疗药物耐药的肿瘤细胞。氯乙基亚硝胺(Cenus)已被广泛用于单独或联合使用。这些试剂来自于DNA上的O_6-烷基鸟嘌呤，被O_6-甲基鸟嘌呤-DNA甲基转移酶(MGMT)修复。本研究的第一个目的是研究MGMT活性和MGMT mRNA在脑肿瘤中的分布和定量。利用pKW100探针和RT-PCR方法检测到高MGMT活性的脑瘤细胞有大量的MGMT mRNA，而低MGMT活性的脑瘤细胞MGMT mRMA的扩增与MGMT活性密切相关。因此，MGMT mRNA的检测将是CENU化疗应用的有用指标。这表明了EVALUA…的巨大优势研究了O_6-甲基鸟嘌呤、O_6-(对甲基苯基)鸟嘌呤、O_6-(对氯苯基)鸟嘌呤、O_6-(对甲氧基苯基)鸟嘌呤、O_6-甲基次黄嘌呤、O_6-(对甲氧基苯基)鸟嘌呤、O_6-甲基次黄嘌呤、O_6-苄基鸟嘌呤、O_6-(对甲基苯基)鸟嘌呤、O_6-(对甲基苯基)鸟嘌呤、O_6-(对甲基苯基)鸟嘌呤、O_6-甲基次黄嘌呤对10种肿瘤细胞系ACNU敏感性的影响。和O_6-(对氯苯基)鸟嘌呤，而不是O_6-甲基鸟嘌呤或O_6-甲基次黄嘌呤。O_6-(对甲氧基苄基)鸟嘌呤对ACNU细胞毒性有中度增敏作用。生物学效应提示O_6-苄基鸟嘌呤骨架上的外环2-氨基和O_6-苄基都是抑制MGMT活性所必需的。本研究结果为CENU与DNA修复酶MGMT相关的选择性化疗提供了分子生物学基础，有望提高恶性脑肿瘤的疗效和延长生存期。较少

A major problem of chemotherapy for malignant brain tumors is existence of tumor cells refractory to chemotherapeutic agents. Chloroetylnitrosouresa (CENUs) have widely been used singly or in combination. These agents from O_6-alkylguanines on DNA,which are repaired by the enzyme O_6-methylguanine-DNA methyltransferase (MGMT) . The first aim of the present research project was to study the distribution and quantification of MGMT activities and MGMT mRNA in brain tumors. The second was to facilitate a strategy aiming at overcoming MGMT-related resistance through inactivation of MGMT by exogenous deplators.Brain tumor cells showing high MGMT activities had a large number of MGMT mRNA using a probe pKW100 and RT-PCR method, whereas those showing low MGMT activities had less MGMT mRNA.Amplification of MGMT mRMA is closely related with MGMT activities. Measurements of MGMT mRNA will be therefore a useful index of application of CENU chemotherapy. This indicates great advantage in the evalua … More tion of MGMT activity of brain tumors.resected samples of which are too limited.We tested effect of O_6-methylguanine, O_6-benzylguanine, O_6- (p-methylbenzyl) guanine, O_6- (p-chlorobenzy) guanine, O_6- (p-methoxybenzyl) guanine, O_6-methylhypoxantine, and O_6-benzylhypoxanthine on the sensitivity of ten tumor cell lines to ACNU.The sensitivity of MGMT-proficient tumor cells was greatly enhanced by pretreatment O_6-benzylguanine, O_6- (p-methylbenzl) guanine, and O_6- (p-chlorobenzyl) guanine, but not by O_6-methylguanine or O_6-methylhypoxanthine. O_6- (p-Methoxybenzyl) guanine moderately sensitized two cell lines to ACNU cytotoxicity. Biological effects O_6-alkylguanine derivatives suggest that the exocyclic 2-amino and O_6-benzyl groups in O_6-benzylguanine skeleton are both essential for the inhibition of MGMT activity.The results of the present study offer molecular biological basis for selective chemotherapy of CENU in relation to DNA repair enzyme MGMT,and promise to potentiate therapeutic effects and to increase long survivors in malignant brain tumors. Less