Project 1: Synthesis, Characterization and Biomonitoring of DNA Containing Site-Specific Chemotherapy-Induced Complex Alkylation Products

项目1：含有位点特异性化疗诱导的复杂烷基化产物的DNA的合成、表征和生物监测

• 批准号: 9788283
• 负责人: Carmelo J Rizzo
• 金额: $ 42.07万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9788283
• 关键词: Abbreviations; Address; Adriamycin PFS; Alkylating Agents; Alkylation; Amination; Anthracyclines; Antineoplastic Agents; Attention; Aziridines; Benign; Biological; Biological Markers; Biological Monitoring; Biology; Breast; Breast Cancer Cell; Breast Cancer Patient; Breast Cancer cell line; Cations; Cells; Chemicals; Chemotherapy-Oncologic Procedure; Clinical Treatment; Complex; Cyclophosphamide; DNA; DNA Adducts; DNA Alkylation; DNA Damage; DNA biosynthesis; DNA crosslink; DNA lesion; Data; Deoxyguanosine; Depurination; Doxorubicin; Drug Design; Exposure to; FAPy; Fapy-dG; Future; Genetic Transcription; Guanine; Human; Imidazole; Imines; In Vitro; Ions; Lead; Leukocytes; Link; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Mechlorethamine; Mitoxantrone; Molecular Conformation; Mutagenesis; New Agents; Oligonucleotides; Oximes; Patients; Pharmaceutical Preparations; Phase; Protocols documentation; Reaction; Regimen; Resources; Rodent Model; Second Primary Cancers; Site; Solid; Structure; Testing; Thiotepa; Topoisomerase II; Treatment Efficacy; Work; adduct; alpha Dystroglycan; amino group; analog; analytical method; antitumor agent; base; calf thymus DNA; carcinogenicity; chemotherapy; crosslink; cytotoxic; cytotoxicity; improved; in vivo; insight; interest; methoxyamine; novel; prevent; repaired; synergism; targeted agent

Project Summary/Abstract The combination of doxorubicin (adriamycin) and cyclophosphamide, so-called AC chemotherapy, is commonly used for the clinical treatment of breast and other cancers. DNA apurinic/apyrimidinic (AP) sites are a common result of alkylating agents such as nitrogen mustards. The cytotoxicity of alkylating agents can be enhanced by methoxyamine, which forms a conjugate with AP sites and inhibits repair. We propose a new reactivity of doxorubicin and related agents in which they form a covalent conjugate with AP sites in DNA, and this adduct is highly cytotoxic. The following interconnected Aims are proposed to address this hypothesis. Project 1 will demonstrate that the anthracycline antitumor agents doxorubicin, mitoxantrone and pixantrone form covalent conjugates with AP sites in duplex DNA through an imine linkage (Specific Aim 1). Imines linkages form reversibily; we therefore propose to synthesize new analogues that form more stable hydrazine, semicarbazone, and oxime covalent linkages to AP sites. Nitrogen mustards cause complex forms of DNA damage. In order to evaluate the biological processing of nitrogen mustard DNA adduct, we will synthesize structurally defined oligonucleotides that contain DNA adducts of nor-nitrogen mustard (the active form of cyclophosphamide) and thioTEPA. This includes N5-substituted formamidopyrimidine-dG adducts derived from imidazole ring-opened of the cationic N7-alkylated dG (Specific Aim 2). We will also synthesize complex secondary products from nitrogen mustards cross-links in which one of the cross-linked dGs has undergone deglycosylation to an AP site. Based on deglycosylation rates, we predict thioTEPA and nor- nitrogen mustard will lead to a high burden of AP sites compared to other alkylating agents. The extent to which these alkylating agents form AP sites in DNA in vitro (calf thymus DNA and cultured human breast cancer cells) and in vivo (rodent models and white blood cells from breast cancer patients undergoing AC chemotherapy) as well as AP site conjugates of doxorubicin and related agents will be quantified by ion trap multistage mass spectrometry (Specific Aim 3). Project 1 will work closely with the DNA Synthesis Resource Core to provide site-specifically modified oligonucleotides containing nor-nitrogen mustard and thioTEPA adducts (Aim 1) as well as covalent drug conjugates of an AP site (Aim 2) to Project 2 for replication, mutagenesis and repair studies, and Project 3 for structural analysis. Project 2 will examine the cytotoxicity of the anthracyclines and new analogues in combination with nor-nitrogen mustard or thioTEPA in cultured human breast cancer cell lines.

项目摘要/摘要 阿霉素（阿霉素）和环磷酰胺（所谓的AC化学疗法）的组合是 通常用于乳腺癌和其他癌症的临床治疗。 DNA apurinic/apyrimidinic（AP）位点是 烷基化剂（例如氮芥末）的常见结果。烷基化剂的细胞毒性可以是 通过甲氧基因增强，甲氧基因与AP位点形成结合并抑制修复。我们提出了一个新的 阿霉素和相关剂的反应性，它们与DNA中的AP位点形成共价结合，并 该加合物具有高度的细胞毒性。提出了以下相互联系的目的来解决这一假设。 项目1将证明蒽环类抗肿瘤剂阿霉素，mitoxantrone和 Pixantrone通过亚胺连接形成双链DNA中的AP位点共轭（特定AIM 1）。 imines链接形成反向;因此，我们建议合成形成更稳定的新类似物 与AP位点的氢氮，半肺巴唑和Oxime共价连接。氮芥末引起复杂形式 DNA损伤。为了评估氮芥末DNA加合物的生物处理，我们将 合成的结构定义的寡核苷酸，该寡核苷酸含有北硝基的DNA加合物（活性 环磷酰胺的形式）和thiotepa。这包括N5取代的Formamidopyrimidine-DG加合物 源自阳离子N7烷基化DG的咪唑环开环（特定目标2）。我们还将合成 来自氮芥末交联的复杂二级产品，其中一种交联的DGS具有 经过直接糖基化为AP位点。根据脱糖基化速率，我们预测thiotepa和nor- 与其他烷基化剂相比，氮芥末将导致AP位点的高负担。范围 这些烷基化剂在体外形成DNA中的AP位点（小牛胸腺DNA和培养的人乳 癌细胞）和体内（乳腺癌患者的啮齿动物模型和白细胞 化学疗法）以及阿霉素和相关药物的AP位点缀合物将通过离子陷阱来定量 多阶段质谱法（特定目标3）。 项目1将与DNA合成资源核心紧密合作，以提供特定于现场的修改 含有非硝基芥末和硫代加合物的寡核苷酸（AIM 1）以及共价药物 AP站点的共轭（AIM 2）进行项目2进行复制，诱变和维修研究，以及项目3 结构分析。项目2将检查蒽环类药物和新类似物的细胞毒性 在培养的人类乳腺癌细胞系中与北硝基芥末或Thiotepa结合。