Isomeric HRT estrogen-DNA adducts: Structure and Repair

异构 HRT 雌激素-DNA 加合物：结构和修复

• 批准号: 7531045
• 负责人: Nicholas E Geacintov
• 金额: $ 26.84万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-01-01 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7531045
• 关键词: 4-hydroxy-equilenin; Adenine; Adverse effects; Affect; Base Sequence; Biological Markers; Biological Models; Breast Cancer Cell; Cancer Etiology; Cancer cell line; Catechols; Cell Line; Cells; Characteristics; Computational Technique; Computing Methodologies; Conjugated Equine Estrogens; Cultured Cells; Cytosine; DNA; DNA Adducts; DNA Repair; DNA Repair Enzymes; DNA Structure; DNA lesion; DNA-Directed DNA Polymerase; Data; Development; Drug Formulations; Equilenin; Equilin; Equus caballus; Estradiol; Estrogen Replacements; Estrogens; Estrone; Excision; Goals; Guanine; Health Benefit; Hormone replacement therapy; Hormones; Human; In Vitro; Kinetics; Lead; Lesion; Malignant Neoplasms; Mammary Gland Parenchyma; Mammary gland; Metabolic Activation; Methods; Molecular Conformation; Mutation; Nucleotide Excision Repair; Oligodeoxyribonucleotides; Oligonucleotides; Pattern; Pharmaceutical Preparations; Postmenopause; Predisposition; Process; Property; Rattus; Relative (related person); Resistance; Resolution; Risk; Site; Structure; System; Testing; Time; Tissue Model; Tissues; Woman; adduct; base; cancer risk; cell type; design; insight; malignant breast neoplasm; repair enzyme; repaired; tumor

DESCRIPTION (provided by applicant): A widely used hormone replacement therapy (HRT) formulation (~ 57 million prescriptions in 2003 alone) contains the conjugated equine estrogens equilenin (EN), equilin, and 8,9-dehydro-estrone (total ~ 54% composition), and the endogenous estrone and 17beta-estradiol. Although the long term risks of cancers of the breast and other hormone-sensitive tissues have been well publicized, women continue to use this formulation because of the significant health benefits associated with the relief of post-menopausal systems. The metabolic activation of equine and endogenous estrogens to genotoxic metabolites has been implicated in the etiology of cancers associated with HRT. It is known that the catechol 4-hydroxyequilenin (4-OHEN) is the most significant metabolite among all three equine estrogens, and that it forms unusual cyclic, stable DNA adducts in vitro, in rat mammary tissue model systems, and in human breast tissue. Adducts of 4-OHEN with cytosine, adenine, and guanine in DNA have been recognized, and each type of adduct is characterized by four stereoisomeric forms. If not removed by cellular DNA repair mechanisms, such adducts may be incorrectly replicated by DNA polymerases, thus causing mutations that may ultimately lead to the development of tumors. Nucleotide excision repair (NER) is the major repair mechanisms that removes bulky adducts from DNA in human cells with efficiencies that depend on the structural properties of the adducts and the distortions they cause in the DNA structure. However, there is no information on how the 12 different 4-OHEN-DNA adducts are processed by NER enzymes. The goal of this project is to determine how the conformations of the different stereoisomeric forms of these adducts influence the efficiencies of their removal by human NER enzymes. In Aim 1, site-specific oligonucleotides with single 4-OHEN-DNA lesions will be constructed. In Aim 2, their structural features will be analyzed by high resolution NMR and computational techniques, while in Aim 3, the resistance to DNA repair will be evaluated using cell free extracts in cells, and in several different types of cells in culture treated with 4-OHEN.The identification of the 4-OHEN-DNA adducts that are resistant to DNA repair in human cells is significant because this information could help to (1) develop biomarkers for identifying women at risk to the adverse effects of HR7, and (2) stimulate the design of new, modified estrogen replacement drugs that are less resistant to nucleotide excision repair enzymes at the DNA adduct level, and thus less active as potential cancer initiating agents.

描述（由申请人提供）：一种广泛使用的激素替代疗法（HRT）制剂（仅2003年就有约5700万张处方）含有结合马雌激素马烯雌酮（EN）、马烯雌酮和8，9-脱氢雌酮（总成分约54%），以及内源性雌酮和17 β-雌二醇。尽管乳腺癌和其他乳腺癌敏感组织的长期风险已经得到了广泛的宣传，但由于与缓解绝经后系统相关的显著健康益处，女性继续使用该制剂。马和内源性雌激素代谢活化为遗传毒性代谢物与HRT相关癌症的病因学有关。众所周知，邻苯二酚4-羟基马雌激素（4-OHEN）是所有三种马雌激素中最重要的代谢产物，并且它在体外、大鼠乳腺组织模型系统和人类乳腺组织中形成不寻常的环状稳定DNA加合物。4-OHEN与DNA中的胞嘧啶、腺嘌呤和鸟嘌呤的加合物已经被识别，并且每种类型的加合物的特征在于四种立体异构形式。如果不通过细胞DNA修复机制去除，这种加合物可能会被DNA聚合酶错误地复制，从而导致突变，最终可能导致肿瘤的发展。核苷酸切除修复（NER）是从人类细胞中的DNA去除大体积加合物的主要修复机制，其效率取决于加合物的结构特性及其在DNA结构中引起的扭曲。然而，没有关于12种不同的4-OHEN-DNA加合物如何被NER酶加工的信息。该项目的目标是确定这些加合物的不同立体异构形式的构象如何影响它们被人NER酶去除的效率。在目标1中，将构建具有单个4-OHEN-DNA损伤的位点特异性寡核苷酸。在目标2中，将通过高分辨率NMR和计算技术分析它们的结构特征，而在目标3中，将使用细胞中的无细胞提取物评估对DNA修复的抗性，在用4-OHEN处理的培养物中的几种不同类型的细胞中。鉴定对人类细胞中的DNA修复具有抗性的4-OHEN-DNA加合物是重要的，因为这些信息可以帮助（1）开发生物标志物，用于识别有HR 7不良反应风险的女性，以及（2）刺激新的、修饰的雌激素替代药物的设计，这些药物在DNA加合物水平上对核苷酸切除修复酶的抗性较低，因此作为潜在的癌症引发剂的活性较低。