INFERTILITY TESTING OF RECOMBINANT SP-10 VACCINE

重组 SP-10 疫苗的不孕症检测

• 批准号: 2198946
• 负责人: JOHN C HERR
• 金额: $ 9.06万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-02-01 至 1995-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2198946
• 关键词: Escherichia coli; acrosome; active immunization; anaphylaxis; baboons; cellular immunity; chimeric proteins; delayed hypersensitivity; drug adverse effect; drug screening /evaluation; female; female antifertility drug; fertility; genetic manipulation; human tissue; immediate hypersensitivity; immunocytochemistry; laboratory mouse; laboratory rabbit; messenger RNA; microorganism genetics; molecular cloning; monoclonal antibody; protein purification; protein structure function; sperm; surface antigens; testis; transfection; vaccines

The primary goal of this grant is to test the anti-fertility potential of a contraceptive vaccine in female baboons. The principal ingredient in the vaccine is an antigen, termed SP-10, existing on the human and baboon sperm head in association with the acrosomal membranes and recently cloned from the human testis. The vaccine will be a fusion protein construct, containing 248 amino acids of SP-10 linked to 26 kD of glutathione transferase. The vaccine will be injected intramuscularly. Toward this primary goal, an initial aim will be the genetic engineering of the SP-10 polypeptide into the pGex expression vector and the purification by affinity chromatography of pure fusion protein to serve as the immunogen and for various assays. To insure that the exact homology between human and baboon SP-10 is understood, baboon SP-10 will be cloned and sequenced from baboon testis utilizing cDNA libraries which we will construct and screen with a human SP-10 nucleotide probe. Messenger RNA from all major baboon organs will be isolated and probed by Northern blotting with the human SP-10 probe to determine if any baboon organ other than testis expresses mRNAs for SP-10. This dat will re-inforce immunohistochemical and ELISA data already obtained which indicate SP-10 is a tissue specific antigen unlikely to elicit autoimmune reactions. Fertility trials of the vaccine will be conducted in 15 female baboons with 15 females receiving the vaccine ingredients minus the SP-10 immunogen. Following immunization, each baboon will be monitored through 9 menstrual cycles for evidence of conception and for anti-SP-10 antibody responses in serum and cervial mucus. Histopathological examination of baboon organs will be conducted at the conclusion of the study. The baboon serum antibodies to SP-10 will be employed to determine the immunodominant portions of the SP-10 molecule. The research aims outlined in this proposal follow the direct path to development of a contraceptive vaccine with potential use in human females.

这笔赠款的主要目标是测试抗生育潜力 雌性狒狒的避孕疫苗。 主要成分 疫苗中存在一种称为 SP-10 的抗原，存在于人类和 狒狒精子头与顶体膜相连 最近从人类睾丸中克隆出来。 该疫苗将是一种融合疫苗 蛋白质构建体，含有与 26 kD 连接的 SP-10 的 248 个氨基酸 谷胱甘肽转移酶。 疫苗将被注射 肌肉注射。 为了实现这一主要目标，最初的目标将是基因工程 将 SP-10 多肽导入 pGex 表达载体中，并且 通过亲和层析纯化纯融合蛋白以供使用 作为免疫原和用于各种测定。 为了确保准确 人类和狒狒 SP-10 之间的同源性已被了解，狒狒 SP-10 将 利用 cDNA 文库从狒狒睾丸中克隆并测序 我们将使用人类 SP-10 核苷酸探针构建和筛选。 来自所有主要狒狒器官的信使RNA将被分离和探测 通过使用人类 SP-10 探针进行 Northern 印迹来确定是否存在 狒狒除睾丸外的器官表达 SP-10 的 mRNA。 这个数据将 强化已获得的免疫组织化学和 ELISA 数据 表明 SP-10 是一种组织特异性抗原，不太可能引起 自身免疫反应。 该疫苗的生育力试验将在 15 只雌性狒狒身上进行 其中 15 名女性接受了减去 SP-10 的疫苗成分 免疫原。 免疫后，每只狒狒都将受到监测 通过 9 个月经周期获取受孕证据和抗 SP-10 血清和宫颈粘液中的抗体反应。 组织病理学 狒狒器官的检查将在研究结束时进行 学习。 狒狒血清中的 SP-10 抗体将用于 确定 SP-10 分子的免疫显性部分。 本提案中概述的研究目标遵循以下直接路径： 开发具有人类潜在用途的避孕疫苗 女性。