MUTAGENESIS BY THE ANTINEOPLASTIC MUSTARDS

抗肿瘤芥子的诱变作用

• 批准号: 3199709
• 负责人: WILLIAM D HENNER
• 金额: $ 16.51万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-15 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3199709
• 关键词: antineoplastics; chemical carcinogen; chlorambucil; cyclophosphamide; drug adverse effect; gene deletion mutation; gene rearrangement; genetic mapping; human subject; hypoxanthine phosphoribosyltransferase; lymphoblast; mechlorethamine; melphalan; molecular cloning; molecular oncology; mutagen testing; mutagens; neoplasm /cancer genetics; nitrogen mustard; nucleic acid sequence; polymerase chain reaction; tissue /cell culture

This project will determine the frequency and molecular structure of the mutations induced in human lymphocytes by the nitrogen mustard group of antineoplastic alkylating agents which are well-established as carcinogens in humans. Patients treated with these drugs have an increased risk of second malignancies particularly acute leukemia. Many of the induced leukemias are associated with characteristic chromosomal deletions and rearrangements. In spite of their wide use and established carcinogenicity, relatively less is known about the mechanism of mutagenicity for the nitrogen mustards than for many environmental carcinogens. Two complementary systems will be used to study mutations induced by these DNA-DNA cross-linking agents. In both systems the genetic locus to be studied is the human hypoxanthine-guanine- phosphoribosyltranferase (HPRT) locus. In the first system, the human lymphoblastoid cell line WIL-2 NS will be exposed in vitro to some of the clinically useful alkylating agents, primarily the nitrogen mustards, and the frequency of mutations induced at various levels of cytoxicity will be determined. The polymerase chain reaction, molecular cloning and DNA sequencing will be used to identify hprt mutations as base changes, deletions or rearrangements. Preliminary studies indicate that nitrogen mustard (mechlorethamine) induces a high frequency of large gene deletions at the HPRT locus. Moreover, within the HPRT gene, a "hotspot" for mechlorethamine-induced deletions has been identified in the region between exon4 and exon9, with 16/17 of such deletions comprising the 3' end of the gene. The nature of the sequence(s) that determines the increased frequency and unidirectionality for deletions in this region will be determined by cloning and sequencing of the mutant HPRT fragments containing the breakpoint. This sequence information will allow us to test various hypotheses as to the mechanism by which such deletions occur. In the second system, lymphocytes will be obtained from patients receiving therapy with cyclophosphamide for multiple sclerosis or chlorambucil for non-Hodgkin's lymphoma. In preliminary studies, we have identified an increased mutation frequency at the HPRT locus in lymphocytes of cyclophosphamide-treated patients. The molecular structure (base substitutions, deletion endpoints, etc.) of the mutations induced in vivo by cyclophosphamide, chlorambucil and melphalan will be analyzed. These complementary systems allow study of the frequency and structure of HPRT mutations under conditions of either in vitro or in vivo exposure. Patients, nurses, pharmacists and manufacturing workers are frequently exposed to the nitrogen mustards and to related compounds. This project should allow better characterization of the exact nature of the mutagenic events that occur in human cells exposed to the nitrogen mustards (or other environmental exposures) which may later lead to second malignancies, germ line mutations and birth defects in exposed individuals.

这个项目将确定频率和分子结构 氮芥子组诱发的人淋巴细胞突变 公认为致癌物的抗肿瘤烷基化药物 在人类身上。接受这些药物治疗的患者有更高的风险。 第二种恶性肿瘤，尤其是急性白血病。许多诱发的 白血病与特征性的染色体缺失和 重新安排。尽管它们被广泛使用和确立 致癌性，对其致癌机制知之甚少。 氮素芥菜的致突变性高于许多环境 致癌物质。将使用两个互补的系统来研究突变 由这些DNA-DNA交联剂诱导。在这两个系统中，基因 要研究的基因是人次黄嘌呤-鸟嘌呤- 磷酸核糖转移酶(HPRT)基因座。在第一个系统中，人类 淋巴母细胞系WIL-2 NS将在体外暴露于一些 临床上有用的烷化剂，主要是含氮芥末，以及 在不同水平的细胞毒性中诱导突变的频率将是 下定决心。聚合酶链式反应、分子克隆和DNA 测序将用于识别HPRT突变为碱基变化， 删除或重新排列。初步研究表明，氮 芥末(机械除草剂)会导致高频率的大基因缺失 在HPRT基因座上。此外，在HPRT基因中， 已在以下区域发现了由机氯乙胺引起的缺失 外显子4和外显子9，其中16/17的缺失包括 吉恩。序列的性质(S)决定了增加的 该区域删除的频率和单向性将为 由突变型HPRT片段的克隆和测序确定 包含断点的。这些序列信息将使我们能够测试 关于这种缺失发生的机制的各种假说。在……里面 第二个系统，淋巴细胞将从接受治疗的患者中获得 环磷酰胺治疗多发性硬化症或氯氨丁苯治疗 非霍奇金淋巴瘤。在初步研究中，我们发现了一种 大鼠淋巴细胞HPRT基因突变频率增加 接受环磷酰胺治疗的患者。分子结构(碱基 替换、删除终结点等)在体内诱导的突变 用环磷酰胺、百菌清和马法兰进行分析。这些 互补系统允许研究HPRT的频率和结构 在体外或体内暴露条件下的突变。 病人、护士、药剂师和制造业工人经常 暴露在氮素芥末和相关化合物中。这个项目 应该可以更好地描述诱变剂的确切性质 暴露在氮素芥菜(或其他 环境暴露)，这可能会导致第二种恶性肿瘤，细菌 暴露个体的基因突变和先天缺陷。