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）：该项目的目标是测试 改进乳腺癌诊断技术的假设 使用识别 GRSA 蛋白的试剂生成。此类试剂包括 单克隆抗体和工程单链可变区 (scFv) 形式 这些抗体。 GRSA（糖肽相关细胞表面抗原）蛋白 是由加压素表达产生的特定肿瘤产物 （VP）基因。它们似乎是由所有乳腺癌和所有导管癌产生的 原位癌（DCIS）。或者，VP 基因表达和 GRSA 蛋白 正常乳腺组织和所有乳腺纤维囊性病变中均不存在 包括非典型导管增生（ADH）。对这种区别的确认 应提供一种早期检测初始和复发的方法 疾病。初步研究表明 GRSA 蛋白可以代表一个靶标 用于免疫诊断和未来的治疗干预。因此一个 资助期间研究的主要重点是生成和评估 来自用于这些目的的可用抗体的 ScFv GRSA 定向分子。 具体目标是： (i) 评估新的免疫组织化学和 RT-PCR 乳腺癌和 DCIS 的单克隆诊断程序 针对 GRSA 蛋白的抗体或 VPmRNA 的引物； (二) 制造业 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 GRSA 蛋白的抗体； (三) 评估 这些 scFv 分子因其能力而成为有用的体外临床药物 提供档案中所有乳腺癌的阳性免疫染色 文库，并与培养物中的癌细胞结合； (iv)评价这些scFv 分子作为体内有用的临床药物，基于它们的能力 选择性靶向小鼠肿瘤异种移植物上的 GRSA 蛋白。研究将雇用 RT-PCR、连接和克隆； ABC免疫组化、Western分析；流动 细胞计数术；放射免疫分析；组织的辐射分析；和伽马相机闪烁扫描。 这项研究的预期成果是开发简单、灵敏、 区分导管原位癌 (DCIS) 的可靠且普遍可用的诊断方法 组织活检中的非典型增生和乳腺癌，这可以 通过放射免疫成像检测和定位患者的复发性疾病。 这些方法可能适用于所有乳腺癌患者或 疑似乳腺癌。