MUTATIONAL STUDIES OF AIDS THERAPEUTIC AGENTS

这项建议的目的是调查 抗艾滋病毒治疗药物对人体细胞的慢性治疗。一个 当前医学研究的主要重点是发展 通过人体免疫缺陷预防或逆转感染的抗病毒药物 病毒(HIV)，获得性免疫缺陷的病原体 综合症(艾滋病)。对艾滋病毒生命周期的研究针对的是相反的情况 将病毒RNA转录成DNA是病毒最可行的阶段 特定的治疗干预。在过去的三年里，有几个 脱氧核苷类似物，用作HIV逆转的抑制剂 转录酶，已经成功地应用于患者 艾滋病与艾滋病相关情结。公众要求加快步伐的压力 然而，这些组件的应用引发了对以下问题的担忧 它们对人类宿主DNA复制和修复过程的致突变作用 在慢性治疗期间。这些担忧促使我们提出一项研究 这些抗病毒药物对人体DNA合成的长期影响 细胞。提出了四个具体的实验：1.人的发展 适合突变研究的细胞系。在经历了长期增长之后 含有抗病毒化合物的培养液，细胞克隆将被筛选 活性核苷激酶活性并扩增到细胞系中用于 随后的突变研究。2.基因突变的比较分析 选定的人类细胞生长在对照培养液和含 DNA合成正向突变检测抗病毒化合物。3. 突变的分子特征。对于那些抗病毒药物来说 引起高突变频率，DNA合成的保真度 从细胞中纯化的主要复制和修复DNA聚合酶 对长期服用这些药物的患者进行分析。4.评估 通过比较确定和新开发的抗病毒药物 HIV逆转录酶和纯化的人DNA的抑制作用研究 聚合酶α、β和β，并分析细胞的DNA产物 聚合酶和这些抑制剂。抑制数据与 正向突变分析将为艾滋病提供宝贵的参考 未来的治疗策略和生化原理 抗病毒治疗药物设计。

The objective of this proposal is to investigate the mutagenic effect of chronic treatment of human cells by anti-HIV therapeutic agents. A major emphasis of current medical research is the development of antiviral agents to prevent or reverse infection by human immunodeficiency virus (HIV), the etiologic agent of acquired immunodeficiency syndrome (AIDS). Studies of the HIV life cycle have targeted the reverse transcription of viral RNA to DNA as the most feasible stage for virus- specific therapeutic intervention. In the past three years, several deoxynucleoside analogues, used as inhibitors to the HIV reverse transcriptase, have been applied with some success to patients with AIDS and AIDS-related complex. The public pressure to expedite application of these components has, however, engendered concerns about their mutagenic effect on human host DNA replication and repair process during chronic treatment. These concerns prompt us to propose a study of the long term effects of these antiviral agents on DNA synthesis in human cells. Four specific experiments are proposed: 1. Development of human cell lines suitable for mutation studies. Following prolonged growth in media with antiviral compounds, cell clones will be screened for active nucleoside kinase activity and expanded into cell lines for the subsequent mutation studies. 2. Comparative analysis of mutations of selected human cells grown in control medium and in medium containing antiviral compounds by DNA synthesis forward mutation assays. 3. Molecular characterization of mutations. For those antiviral agents that cause high mutational frequencies, DNA synthesis fidelity of the principal replicative and repair DNA polymerase purified from cells chronically treated by these drugs will be analyzed. 4. Evaluation of the established and newly developed antiviral drugs by comparative inhibition studies of HIV reverse transcriptase and purified human DNA polymerase alpha, beta, and delta and analyze the DNA products of cellular polymerase with these inhibitors. The inhibition data correlated with the forward mutation analyses will provide invaluable reference for AIDS therapeutic strategy and biochemical rationale for the future antiviral therapeutic drug design.