HUMAN TESTIS CDNAS IDENTIFIED BY INFERTILITY SERA

这项研究计划的长期目标是开发一种安全和 妇女使用的有效的免疫避孕药。在以前的研究中，一个 基于精子序列的完全合成的嵌合肽- 特异性乳酸脱氢酶(LDH-C4表位HC-5-19-TT)可逆性 将雌性狒狒的生育力抑制了70%。在当前 项目期间，我们使用免疫不孕症患者的血清 确定三种新的精子特异性抗原，它们可以单独使用或 联合应用HC5-19-TT以加强对妊娠的保护 到90%以上。这个项目的近期目标是 测试其中一种B细胞表位的安全性和有效性 抗原，精子特异性的钙调蛋白亚型(t-cast)，单独和在 与LDH-C4表位结合。卡巴斯丁是多肽 钙蛋白酶的抑制剂，一种半胱氨酸蛋白酶，扮演着重要的 裂解前在囊泡胞吐和膜融合事件中的作用。 1例不孕症患者血清中抗t-Cast抗体(YM) 凝集人精子和抑制精子对宫颈的穿透 粘液。人工合成嵌合B细胞表位蛋白(HTC25-41-TT) 在兔和雌性猕猴中具有免疫原性，并且抗肽 人精子凝集和固定的血清。因此，t-cast是 存在于顶体表面的完整精子，是一种初级的 开发一种免疫避孕药的候选者。四个具体的 提出的目标是：1)确定T-CAST在 通过靶向基因破坏在小鼠中受精；2)建立 HTC25-41-TT的女性避孕效果和可逆性 食蟹猴；3)测试由两者组成的亚单位疫苗 雌性猕猴HTC25-41-TT和HC5-19-TT； Coli表达大量重组形式的亚基 在良好的实验室实践下用于大规模生产的疫苗 指导方针。这项技术将提供一种安全有效的 替代目前可用的避孕方法。

The long-term objective of this research plan is to develop a safe and effective immuocontraceptive for use by women. In previous studies, a completely synthetic chimeric peptide based on the sequence of sperm- specific lactate dehydrogenase (LDH-C4 epitope HC-5-19-TT) reversibly suppressed fertility in female baboons by 70 percent. In the current project period, we used sera from immunologically infertile patients to identify three new sperm-specific antigens which can be used alone or in combination with HC5-19-TT to increase the protection from pregnancy to greater than 90 percent. The immediate goal of this project is to test the safety and efficacy of a B-cell epitope from one of these antigens, the sperm-specific isoform of calpastatin (t-CAST), alone and in combination with the LDH-C4 epitope. Calpastatin is the peptide inhibitor of calpain, a cysteine protease that plays an important preteolytic role in vesicle exocytosis and membrane fusion events. Antibodies to t-CAST in the serum of an infertile patient (YM) agglutinated human sperm and inhibited sperm penetration of cervical mucus. A synthetic chimeric B-cell epitope of the protein (HTC25-41-TT) was immunogenic in rabbits and female macaques, and the anti-peptide sera agglutinated and immobilized human sperm. Therefore, t-CAST is present on the surface of acrosome intact sperm, and is a primary candidate for development of an immunocontraceptive. Four Specific Aims are proposed: 1) to establish the functional role of t-CAST during fertilization by targeted gene disruption in mice; 2) to establish contraceptive efficacy and reversibility of HTC25-41-TT in female cynomologus macaques; 3) to test a subunit vaccine composed of both HTC25-41-TT and HC5-19-TT in female macaques; and 4) to engineer E. coli to express large amounts of a recombinant form of the subunit vaccine for large-scale production under Good Laboratory Practices guidelines. This technology would provide a safe and effective alternative to presently available methods of contraception.