GRSA Proteins in the Diagnosis of Breast Cancer

GRSA 蛋白在乳腺癌诊断中的应用

• 批准号: 6515212
• 负责人: WILLIAM G NORTH
• 金额: $ 15.9万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6515212
• 关键词: MCF7 cell; athymic mouse; breast neoplasm /cancer diagnosis; breast neoplasms; diagnosis design /evaluation; early diagnosis; glycopeptides; human tissue; immunocytochemistry; molecular cloning; monoclonal antibody; neoplasm /cancer immunodiagnosis; neoplasm /cancer relapse /recurrence; polymerase chain reaction; surface antigens; tumor antigens; vasopressins; western blottings

DESCRIPTION (provided by applicant): The objective of this project is to test the hypothesis that improved diagnostic techniques for breast cancer can be generated with agents recognizing GRSA proteins. Such agents comprise monoclonal antibodies and engineered single-chain variable region (scFv) forms of these antibodies. GRSA (Glycopeptide-Related Cell Surface Antigen) proteins are specific tumor products that result from the expression of the vasopressin (VP) gene. They are produced by seemingly all breast cancer and all ductal carcinoma in Situ (DCIS). Alternatively, VP gene expression and GRSA proteins are absent from normal breast tissue and from all breast fibrocystic conditions including atypical ductal hyperplasia (ADH). A confirmation of this distinction should provide a method for the early detection of initial and recurrent disease. Preliminary studies indicate GRSA proteins could represent a target for immunodiagnosis and for future therapeutic interventions. Therefore a principal focus of research over the funding period is to generate and evaluate ScFv GRSA-directed molecules from available antibodies for these purposes. Specific Aims are directed at: (i) assessing new immunohistochemical and RT-PCR diagnostic procedures for breast cancer and DCIS that feature monoclonal antibodies against GRSA proteins or primers for VPmRNA; (ii) manufacturing single-chain scFv diabodies and triabodies representing our monoclonal antibodies that recognize GRSA proteins on breast cancer; (iii) appraising these scFv molecules as useful clinical agents in vitro based on their ability to provide positive immunostaining of all breast carcinoma in an archival library, and to bind to cancer cells in culture, and; (iv)appraising these scFv molecules as useful clinical agents in vivo based on their ability to selectively target GRSA proteins on mouse tumor xenografts. Studies will employ RT-PCR, ligation, and cloning; ABC immunohistochemistry, Western analysis; flow cytometry; RIA; radiometric analysis of tissues; and gamma-camera scintigraphy. Expected outcomes of this research are the development of simple, sensitive, reliable, and universally available diagnostic methods that distinguish DCIS and breast cancer from atypical hyperplasia in tissue biopsies, and that can detect and localize recurrent disease in patients through radioimmunoimaging. These methods could be applicable to all patients with breast cancer or suspected breast cancer.

描述（由申请人提供）：本项目的目的是测试 乳腺癌诊断技术的改进， 用识别GRSA蛋白的试剂产生。这样的试剂包括 单克隆抗体和工程化单链可变区（scFv）形式 这些抗体。GRSA（糖肽相关细胞表面抗原）蛋白 是由血管加压素表达引起的特异性肿瘤产物 (VP)基因它们是由似乎所有的乳腺癌和所有的导管癌产生的。 原位癌（DCIS）。或者，VP基因表达和GRSA蛋白 不存在于正常乳腺组织和所有乳腺纤维囊性疾病中 包括非典型导管增生（ADH）。对这种区别的确认 应该提供一种早期发现初始和复发的方法， 疾病初步研究表明，GRSA蛋白可能是一个靶点， 用于免疫诊断和未来的治疗干预。因此 在资助期间，研究的主要重点是产生和评估 ScFv GRSA定向分子来自用于这些目的的可用抗体。 具体目标是：（i）评估新的免疫组化和RT-PCR 乳腺癌和DCIS的诊断程序， 针对GRSA蛋白的抗体或针对VPmRNA的引物;（ii）制造 代表我们的单克隆抗体的单链scFv双抗体和三抗体 识别乳腺癌上的GRSA蛋白的抗体;（iii）评估 这些scFv分子基于它们的能力在体外作为有用的临床试剂 提供存档中所有乳腺癌的阳性免疫染色 （iv）鉴定这些scFv， 分子作为体内有用的临床试剂的能力， 选择性靶向小鼠肿瘤异种移植物上的GRSA蛋白。研究将采用 RT-PCR、连接和克隆; ABC免疫组织化学、Western分析;流式细胞术 细胞术;放射免疫分析;组织放射性分析;和γ照相机放射照相术。 这项研究的预期成果是开发简单，灵敏， 可靠且普遍可用的诊断方法， 和乳腺癌组织活检中的非典型增生， 通过放射免疫成像检测和定位患者的复发性疾病。 这些方法可以适用于所有乳腺癌患者， 疑似乳腺癌