FLOW CYTOMETRIC STUDY OF ANTICANCER DRUG EFFECTS ON DNA

本研究的总体目标是发展一种预测性的 一种筛选肿瘤有效烷化剂的方法 治疗和检测耐药肿瘤细胞。 发展 将基于测定之间的线性关系。 单克隆抗体（MAb）F7-26与细胞DNA的结合， 氮芥衍生物和亚硝基脲的细胞毒性 1-6 log细胞杀伤的范围;以及MAb 区分敏感细胞和耐药细胞。 主要 在这项研究中要回答的问题是， MAb结合与药物敏感性或耐药性的相关性 可以在不同的鼠和人肿瘤细胞中观察到。 mAb 与单个细胞中DNA的结合将通过流式细胞仪测量。 细胞仪 单克隆抗体F7-26免疫反应性与 用烷化剂处理的细胞中的DNA和细胞杀伤 将在敏感和耐药的两种成对细胞系中进行研究 i）肺癌细胞系A549（Mer+ 表型）和A427（Mer-表型）耐药和敏感 ii）卵巢癌细胞系A2780 对L-PAM敏感和抗性。 在每一对线中， 细胞存活、初始DNA损伤和修复之间的关系 将研究药物去除后的DNA损伤。 鼠标固体 肿瘤刘易斯肺癌及其耐药亚系 亚硝基脲和环磷酰胺将与药物治疗， 体外和体内。 单克隆抗体与非整倍体肿瘤细胞的结合 将与肿瘤对治疗的反应进行比较。 的 低DNA细胞亚群的出现与 免疫反应性和耐药性的发展 将研究重复的药物治疗。 来描述类型的特征 单克隆抗体检测到的DNA变化的抗原决定簇 采用直接ELISA法和竞争ELISA法研究单克隆抗体F7-26的免疫原性 免疫测定法 单克隆抗体对不同抗体的相对亲和力 脱氧核糖均聚物和寡核苷酸中的碱基数 将确定结合MAb所需的量。 流式细胞术 研究药物对个别肿瘤细胞DNA的影响 在此，应能应用新的方法， 临床环境。

The general goal of this study is the development of a predictive assay which may select effective alkylating agents for tumor treatment and detect drug-resistant tumor cells. The development of the assay will be based on o linear relationship between the binding of monoclonal antibody (MAb) F7-26 to cellular DNA and the cytotoxicity of nitrogen mustard derivatives and nitrosoureas in the range of 1-6 log cell killing; and the ability of MAb to distinguish between sensitive and drug-resistant cells. The major question to be answered in this study is whether a quantitative correlation between MAb binding and drug sensitivity or resistance can be observed in different murine and human tumor cells. MAb binding to DNA in individual cells will be measured by flow cytometry. The correlation between MAb F7-26 immunoreactivity with DNA in the cells treated with alkylating agents and cell killing will be studied in two paired cell lines sensitive and resistant to alkylating agents: i) lung carcinoma lines A549 (Mer+ phenotype) and A427 (Mer- phenotype) resistant and sensitive to nitrosoureas, respectively; ii) ovarian carcinoma cell lines A2780 sensitive and resistant to L-PAM. In each pair of lines, relationship between cell survival, initial DNA damage and repair of DNA damage after drug removal will be studied. Mouse solid tumor Lewis lung carcinoma and its sublines resistant to nitrosoureas ans cyclophosphamide will be treated with drugs in vitro and in vivo. The binding of MAb to aneuploid tumor cells will be compared with tumor response to treatment. The relationship between the appearance of cell subsets with low DNA immunoreactivity and the development of drug resistance during repeated drug treatments will be studied. To characterize the type of change in DNA detected with the Mab the antigenic determinant of MAb F7-26 will be studied by direct and competitive ELISA immunoassay. The relative affinity of MAb to different deoxyribohomopolymers and the number of bases in oligonucleotides necessary to bind the MAb will be determined. Flow cytometric study of drug effects on DNA in individual tumor cells proposed herein should make it possible to apply the new methodology in clinical settings.