Generation and characterization of adduct-specific anti cisplatin DNA antibodies

加合物特异性抗顺铂 DNA 抗体的生成和表征

• 批准号: 8951743
• 负责人: Orlando D. Scharer
• 金额: $ 20.41万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8951743
• 关键词: Address; Adverse effects; Affect; Antibodies; Antibody Specificity; Antigens; BRCA2 gene; Basic Science; Binding; Biological Assay; Biopsy; Breast; Cancer cell line; Carboplatin; Carrier Proteins; Cell Line; Cells; Chemotherapy-Oncologic Procedure; Cisplatin; Clinic; Clinical; Collection; Communities; Cytotoxic Chemotherapy; Cytotoxic agent; DNA; DNA Adducts; DNA Damage; DNA Repair; DNA Repair Pathway; Defect; Down-Regulation; Drug Combinations; Drug Targeting; Drug resistance; Drug toxicity; Drug usage; ERCC1 gene; Excision; Future; Generations; Genome; Goals; Hybridomas; Individual; Keyhole Limpet Hemocyanin; Link; Lung; Malignant Neoplasms; Malignant neoplasm of cervix uteri; Malignant neoplasm of lung; Malignant neoplasm of ovary; Malignant neoplasm of testis; Malignant neoplasm of urinary bladder; Measurement; Measures; Medical Research; Metabolism; Metals; Methods; Monitor; Monoclonal Antibodies; Mus; Neck Cancer; Nucleotide Excision Repair; Oligonucleotides; Outcome; Pathway interactions; Patients; Pharmaceutical Preparations; Platinum; Platinum adduct; Point Mutation; Process; Property; Purines; Reaction; Reagent; Resistance; Sampling; Site; Specificity; Testing; Therapeutic; Therapeutic Effect; Toxic effect; Treatment Efficacy; Tumor Cell Line; Up-Regulation; Validation; Work; XRCC3 gene; Xeroderma Pigmentosum Complementation Group A; Xeroderma Pigmentosum Complementation Group G; adduct; antitumor agent; antitumor drug; base; cisplatin-DNA adduct; crosslink; cytotoxic; drug metabolism; drug sensitivity; endonuclease; inhibitor/antagonist; insight; interest; malignant breast neoplasm; oncology; public health relevance; purine; repaired; resistance mechanism; response; targeted treatment; tumor; tumor DNA; ultraviolet irradiation

 DESCRIPTION (provided by applicant): Cisplatin and carboplatin (platinum drugs) are among the most successful antitumor drugs and are used to treat testicular, breast, ovarian, bladder, neck and lung cancer. As with many antitumor agents, the efficacy of treatment can vary greatly from patient to patient and the occurrence of resistance is a significant problem. The therapeutic effect of the platinums is based on the formation of different types of DNA adducts, primarily intra- and interstrand crosslinks, by the reaction with two purine bases on one or two complementary strands of DNA. The resistance to platinum treatment occurs by a the up- or down-regulation of several processes, including metal transport in and out of the cell, intracellular metabolism and the removal of the cisplatin DNA adducts by DNA repair pathways. The primary hypothesis of the work proposed in this application is that by developing a monoclonal antibodies with specificity for the individual platinum DNA adducts in the genomes of cells we will be able to 1) determine which platinum adduct is the most therapeutically relevant one, 2) understand the repair mechanisms responsible for resistance, 3) test the sensitivity or resistance to platinum in tumor biopsies and 4) to select patients that are most likely to benefit from therapy and sparing those that are likely to be resistant the severe side effect associated with this type of treatment. In this R21 application, we propose to synthesize oligonucleotides containing the 1,2-GG-, 1,2-AG-, 1,3-GNG-intrastrand and 1,2-GC-interstrand platinum crosslinks, couple them to the KLH carrier protein and use them to immunize mice to generate monoclonal antibodies that recognize the individual adducts with high specificity. These monoclonal antibodies will be then further validated in cell lines with specific DNA repair defects. We predict that these reagents will enable us to show that platinum intrastrand crosslinks persist in cells with defects in the nucleotide excision repair (NER) pathway, while interstrand crosslinks will persist in cells with defects interstrand crosslink (ICL) repair pathwa. We will then measure platinum levels in breast and ovarian cancer cell lines of the NCI-60 collection, which have been characterized for drug sensitivity or resistance. We will determine whether the levels of a specific adduct or all adduct are elevated in platinum-responsive cell lines. Conversely, we will test whether a specific adduct is absent in resistant cell lines. We expect that upon completion of the studies proposed here we will have developed a set of unique reagents that will be of tremendous use for the research and medical community to determine which platinum adducts are clinically most relevant and to work toward developing a robust method to predict clinical outcomes of platinum treatments.

 描述（由申请人提供）：顺铂和卡铂（铂类药物）是最成功的抗肿瘤药物之一，用于治疗睾丸癌、乳腺癌、卵巢癌、膀胱癌、颈癌和肺癌。与许多抗肿瘤药物一样，治疗效果因患者而异，并且耐药性的出现是一个重大问题。铂的治疗效果基于不同类型的 DNA 加合物的形成，主要是通过与一条或两条互补 DNA 链上的两个嘌呤碱基反应而形成的链内和链间交联。对铂治疗的耐药性是由几个过程的上调或下调引起的，包括金属进出细胞的转运、细胞内代谢以及通过 DNA 修复途径去除顺铂 DNA 加合物。本申请中提出的工作的主要假设是，通过开发对细胞基因组中各个铂 DNA 加合物具有特异性的单克隆抗体，我们将能够 1) 确定哪种铂加合物在治疗上最相关，2) 了解导致耐药性的修复机制，3) 测试肿瘤活检中对铂的敏感性或耐药性，以及 4) 选择最有可能受益的患者 避免那些可能对此类治疗产生耐药性的严重副作用。在此 R21 应用中，我们建议合成含有 1,2-GG-、1,2-AG-、1,3-GNG-链内和 1,2-GC-链间铂交联的寡核苷酸，将它们与 KLH 载体蛋白偶联，并用它们免疫小鼠以生成高特异性识别单个加合物的单克隆抗体。这些单克隆抗体将在具有特定 DNA 修复缺陷的细胞系中得到进一步验证。我们预测这些试剂将使我们能够证明铂链内交联在核苷酸切除修复（NER）途径缺陷的细胞中持续存在，而链间交联将在链间交联（ICL）修复途径缺陷的细胞中持续存在。然后，我们将测量 NCI-60 收集的乳腺癌和卵巢癌细胞系中的铂水平，这些细胞系已被表征为药物敏感性或耐药性。我们将确定铂敏感细胞系中特定加合物或所有加合物的水平是否升高。相反，我们将测试耐药细胞系中是否不存在特定的加合物。我们预计，在完成此处提出的研究后，我们将开发出一套独特的试剂，这些试剂将对研究和医学界发挥巨大作用，以确定哪些铂加合物在临床上最相关，并致力于开发一种稳健的方法来预测铂治疗的临床结果。