Development of a new PDT vaccine in a single molecule

开发新的单分子 PDT 疫苗

• 批准号: 04044184
• 负责人: SATO Hiroko
• 金额: $ 2.88万
• 依托单位: National Institute of Health
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for international Scientific Research
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-04044184/
• 关键词: PDT-Fusion vaccine; Fusion gene; Pertussis toxin; Tetanus toxin; Diphtheria toxin; Protective immunogenicity; PDT融合ワクチン

The introduction of the diphtheria-tetanus-pertussis (DTP) vaccine which consists of mainly detoxified diphtheria toxin (DT) , tetanus toxin (TT) , and pertussis toxin (PT) led to a dramatic decline in incidence and mortality of DTP in Japan. However, many children in the most of developing countries are still suffered with these infectious diseases and dying because of unavailability of the vaccine. Since the vaccination with DTP is the best way and must be peerless to prevent these diseases, development of a new DTP vaccine which include only immunoprotective antigens with no toxin activity should be next step towards obtaining an ideal vaccines.The purpose of this project is development of a simplified and well-defined DTP vaccine by combining the protective regions of each component in a single molecule. As a first step towards such a vaccine candidate, a recombinant PT S1 subunit-TT fragment C fusion protein was constructed. The recombinant hybrid protein was produced in Escherich … More ia coli. The 75-KDa fusion protein (p75) was over expressed as a soluble molecule and purified to near homogeneity by two consecutive chromatographic steps. Purified p75 retained its ability to be recognized by protective and neutralizing anti-pertussis toxin antibodies specific for conformational epitopes, such as anti-S1 mono-clonal antibody 1B7. When administered to mice, the hybrid protein was found to be nontoxic but immunogenic. It was capable of inducing strong protection against tetanus and some protection against pertussis, as well as eliciting a pertussis toxin-neutralizing antibody response. Although the levels of anti-pertussis toxin antibodies were rather low, their neutralizing titers correlated well with anti-pertussis toxin titers by ELISA,indicating that protective epitopes are conserved in the recombinant protein.Next, fusion proteins of S1-TTC and a part of DT,were constructed and characterized partially. Expression of these three antigens in E.coli was recognized by the SDS-PAGE and immunoblotting analysis. Furthermore, additional antigens such as HCV were attempted to fuse to the S1-TTC p75 antigen. Studies on the fusion proteins consisting of these triple antigens are in progress. Less

白喉-破伤风-百日咳（DTP）疫苗主要由白喉毒素（DT）、破伤风毒素（TT）和百日咳毒素（PT）组成，其引入导致日本DTP的发病率和死亡率急剧下降。然而，在大多数发展中国家，许多儿童仍然患有这些传染病，并因无法获得疫苗而死亡。由于百白破疫苗是预防这些疾病的最佳方法，而且必须是无与伦比的，因此开发一种只包含免疫保护性抗原而没有毒素活性的新型百白破疫苗应该是获得理想疫苗的下一步。作为这种疫苗候选物的第一步，构建重组PT S1亚基-TT片段C融合蛋白。重组杂合蛋白在土希土中表达， ...更多信息 大肠杆菌。75-KDa融合蛋白（p75）作为可溶性分子过表达，并通过两个连续的色谱步骤纯化至接近均一性。纯化的p75保留了其被对构象表位具有特异性的保护性和中和性抗百日咳毒素抗体（例如抗S1单克隆抗体1B 7）识别的能力。当给予小鼠时，发现杂交蛋白无毒但具有免疫原性。它能够诱导对破伤风的强烈保护和对百日咳的一些保护，以及引发百日咳毒素中和抗体反应。虽然百日咳毒素抗体水平较低，但其中和效价与ELISA检测的百日咳毒素抗体效价相关性良好，表明重组蛋白中存在保护性抗原表位。通过SDS-PAGE和免疫印迹分析识别这三种抗原在大肠杆菌中的表达。此外，还尝试将其他抗原如HCV与S1-TTC p75抗原融合。由这些三重抗原组成的融合蛋白的研究正在进行中。少

项目成果

Philip Boucher: "Neutralizing antibodies and immunoprotection against pertussis and tetanus obtained by use of a recombinant PT-TT fusion protein" Infection and Immunity. 62. 449-456 (1994)

Philip Boucher：“使用重组 PT-TT 融合蛋白获得的针对百日咳和破伤风的中和抗体和免疫保护”《感染与免疫》。

Sato,H.: "A role of pertussis toxin subunits in mouse encephalopathy." Microecology and Therapy. (in press). (1995)

Sato,H.：“百日咳毒素亚基在小鼠脑病中的作用。”

Sato,Y.: "Protective activity of specific antibodies to pertussis protective antigens in a passive aerosol challenge system." Biologicals. 21. 25-26 (1993)

Sato,Y.：“被动气溶胶攻击系统中百日咳保护性抗原的特异性抗体的保护活性。”

Sato, Y., Sato, H., Sasai, S.and Mates, S.: "Protective activity of specific antibodies to pertussis protective antigens in a passive aerosol challenge system." Biologicals. 21. 25-26 (1993)

Sato, Y.、Sato, H.、Sasai, S. 和 Mates, S.：“被动气溶胶攻击系统中百日咳保护性抗原的特异性抗体的保护活性。”

Bryant, A., Bergstrom, R., Zimmerman, G., Salyer, J., Hill, H., Tweten, R., Sato, H.and Stevens, L.: "Clostridium perfringens invasiveness is enhanced by effects of theta toxin upon PMNL structure and function : The roles of leukocytotoxicity and expressi

Bryant, A.、Bergstrom, R.、Zimmerman, G.、Salyer, J.、Hill, H.、Tweten, R.、Sato, H. 和 Stevens, L.：“产气荚膜梭菌的侵袭性因 θ 的影响而增强