Development of an immunocontraceptive DNA vaccine using sperm

利用精子开发免疫避孕 DNA 疫苗

• 批准号: 12557137
• 负责人: MAEGAWA Masahiko
• 金额: $ 8.32万
• 依托单位: THE UNIVERSITY OF TOKUSHIMA
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12557137/
• 关键词: immunocontraceptive vaccine; DNA vaccine; sperm antigen; peptide; 抗精子抗体; 粘膜ワクチン

We previously reported that active immunization of female rats with rSMP-230, residue 1-28 of rSMP-B protein, and 80 kDa human sperm antigen (80 kDa HSA), prevented conception. The purpose of this project was to develop an immunocontraceptive vaccine using an expression vector encoding rSMP-B or 80 kDa HSA. Results are as follows : (l) Compared with the nucleic acid sequence of RSD-1 encoding rSMP-B reported in GenBank"808109" (position ?70-773 : ATAT), 4 nucleotides were different (position 770-777 : CACTCTG), resulting in a difference in three amino acids (His-Met to Pro-Leu-Try). (2) When the p3xFLAG-CMV-14 vector containing RSD-1 was transfected into HEK293 cells, cell extracts of HEK293 cells showed a 20-kD band by Western blotting using an anti-FLAG antibody, but no immunoreaction was found in the lane of culture medium. (3) Active immunization of rabbits with synthetic peptides of 80 kDa HSA elicited significant antibody titers. These antibodies specifically recognized the naive protein in an ELISA and induced in-vitro agglutination of human, rat and monkey sperm. Western blot analysis showed that these antibodies bind to the 80 kDa band of sperm extractThese findings demonstrated that the sperm protein rSMP-B and 80 kDa HSA were pomising candidates for the development of an immunocontraceptive vaccine

我们以前报道过，主动免疫雌性大鼠rSMP-230，rSMP-B蛋白的残基1-28，和80 kDa的人精子抗原（80 kDa的HSA），防止受孕。本研究的目的是利用编码rSMP-B或80 kDa HSA的表达载体开发免疫避孕疫苗。结果如下：（1）与GenBank“808109”中报道的RSD-1编码rSMP-B的核酸序列（位置？70-773：ATAT），4个核苷酸不同（位置770-777：CACTCTG），导致3个氨基酸的差异（His-Met至Pro-Leu-Try）。(2)当将含有RSD-1的p3 xFLAG-CMV-14载体转染到HEK 293细胞中时，HEK 293细胞的细胞提取物通过使用抗FLAG抗体的Western印迹显示20-kD条带，但在培养基泳道中未发现免疫反应。(3)用80 kDa HSA合成肽主动免疫家兔，引起显著的抗体滴度。这些抗体在ELISA中特异性识别幼稚蛋白，并诱导人、大鼠和猴精子的体外凝集。这些结果表明，精子蛋白rSMP-B和80 kDa HSA是开发免疫避孕疫苗的候选蛋白。

项目成果

Maegawa M et al.: "Involvement of carbohydrate molecules on zona pellucida in human fertilization"J Reprod Immunol. 553. 79-89 (2002)

前川 M 等人：“人类受精中透明带上碳水化合物分子的参与”J ReprodImmunol。

Vernekar VJ et al.: "Studies with synthetic peptide of 80 kDa human sperm antigen (80 kDa HAS)"Am J Reprod Immunol. 51. 106-111 (2004)

Vernekar VJ 等人：“80 kDa 人类精子抗原 (80 kDa HAS) 的合成肽的研究”Am J Reprod Nutrition。

Takikawa,M.: "Evaluation of two sperm antigens, rSMP^B and YWK-II, as targets for immunocontraception."Zygote. (印刷中). (2001)

Takikawa, M.：“评估两种精子抗原，rSMP^B 和 YWK-II，作为免疫避孕的靶标。”Zygote（出版中）。

Kamada M et al.: "Specific cytotoxicity of antibody to YAL-198, a sperm antigen peptide, to murine zygote"Arch.Androl.. 47. 89-96 (2001)

Kamada M 等人：“YAL-198（一种精子抗原肽）抗体对鼠受精卵的特异性细胞毒性”Arch.Androl.. 47. 89-96 (2001)

Maegawa,M.: "A repertoire of cytokines in human seminal plasma."J.Reprod.Immunol.. (印刷中). (2001)

Maekawa, M.：“人精浆中的细胞因子库”。J.Reprod.Immunol..（出版中）。