CHARACTERIZATION OF PSORALEN DNA PHOTOADDUCTS

补骨脂素 DNA 照片加合物的表征

• 批准号: 3456693
• 负责人: FRANCIS P GASPARRO
• 金额: $ 12万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-01-01 至 1990-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3456693
• 关键词: DNA; DNA footprinting; DNA repair; adduct; enzyme linked immunosorbent assay; flow cytometry; gel electrophoresis; high performance liquid chromatography; laboratory mouse; leukapheresis; monoclonal antibody; mycosis fungoides lymphoma; neoplasm /cancer photoradiation therapy; nucleic acid sequence; photobiology; photochemistry; phototherapy; psoralens; psoriasis; skin disorder chemotherapy; tissue /cell culture; ultraviolet radiation

8-Methoxypsoralen and ultra-violet A (PUVA) therapies are used in the treatment of psoriasis and cutaneous T cell lymphoma (CTCL). Recently, a new phototherapy for CTCL, photopheresis, has been developed which may serve as a prototype of a new form of photopharmacology. The efficacy of PUVA in these diseases has been attributed to the formation of 8-MOP photoadducts in cellular DNA. The nature and quantity of these adducts in mammalian systems has not been determined, nor has their repair been fully characterized. AlphaDNA, a 92 base pair fragment widely distributed throughout the human genome, will be used to elucidate the effects of specific base sequences on photoadduct formation. Three psoralens (8-methoxypsoralen, 4'-aminomethyl-4,5',8-trimethylpsoralen, and 4',5-dimethylangelicin) will be used in comparative studies to discern the effects of base sequences on monoadduct and crosslink formation. The overall objective of the proposed studies is to determine the nature, quantity and location of the UVA-induced psoralen photoadducts in AlphaDNA. Specifically the following aspects will be examined. 1. the quantitation of psoralen photoadducts in AlphaDNA by HPLC analysis of enzymatically hydrolyzed AlphaDNA samples. AlphaDNA from PUVA treated lymphoid cells will also be analyzed in the same way. 2. the determination of base sequences in which photoadducts have formed, as well as the identity of adducts in specific DNA sites using footprinting techniques in combination with psoralen-DNA monoclonal antibodies. 3. cytofluorometric determination of the effects of PUVA on cultured lymphoid cells using both propidium iodide staining and specific psoralen-DNA monoclonal antibodies. The development of the techniques described in this proposal will eventually lead to the detailed characterization of the repair of psoralen photoadducts. Knowledge of the sites of unrepaired photoadducts will lead to a better understanding of phototherapies based on psoralen plus UVA and culminate in the rational design of new, potentially more effective, psoralens.

8-甲氧补骨脂素和紫外线A(PUVA)疗法用于 治疗银屑病和皮肤T细胞淋巴瘤(CTCL)。最近，一位 已经开发出治疗CTCL的新的光疗法，即光分离，它可能 作为一种新形式的光药学的原型。的功效。 在这些疾病中，PUVA被归因于8-MOP的形成 细胞DNA中的光加合物。这些加合物的性质和数量 哺乳动物的系统还没有确定，它们的修复也没有完全完成 特色化的。 AlphaDNA，一个广泛分布于人类体内的92bpDNA片段 基因组，将被用来阐明特定碱基序列对 光加合物形成。三种补骨脂素(8-甲氧基补骨脂素， 4‘-氨甲基-4，5’，8-三甲基补骨脂素和4‘，5-二甲基当归 在比较研究中用来辨别碱基序列对 单加合物和交联剂的形成。建议的总体目标是 研究的目的是确定这些物质的性质、数量和位置 UVA诱导补骨脂素在αDNA中的光加合物。特别是以下几点 我们将对这些方面进行研究。 1.补骨脂素光加合物在alphadna中的含量 酶解后的AlphaDNA样本。经治疗的PUVA中的AlphaDNA 淋巴样细胞也将以同样的方式进行分析。 2.光加合物形成的碱基序列的测定， 以及使用足迹技术确定特定DNA位置中加合物的身份 补骨脂素-DNA单抗结合技术。 3.细胞荧光法测定PUVA对培养细胞的影响 淋巴样细胞碘化丙啶染色和特异性 补骨脂素-DNA单抗。 本建议书中描述的技术的发展将 最终得到补骨脂素修复的详细表征 光电子加合物。对未修复的光加合物的位置的了解将导致 为了更好地了解基于补骨脂素加UVA和 最终在合理设计新的，可能更有效的， 补骨脂。