新型肺炎链球菌全合成糖脂疫苗的设计、合成与免疫生物活性评价

Streptococcus pneumoniae (SP) infections cause severe threat to human health. The pneumococcal vaccines based on capsular polysaccharide and/or glycoprotein conjugates have been played the important roles on prevention and treatment of pneumococcal infection diseases. However, with the insurgence of high drug-resistant pneumococci and the lower efficiency of immune responds for some SP bacteria, the development of safe and efficient pneumococcal vaccines still remain new challenges and difficulties to be solved. The fully synthetic, self-adjuvanting glycoconjugate vaccines with monophosphoryl lipid A (MPLA ) as carrier molecule have been recently received more concerns. This research project aims at developing of novel fully synthetic anti-pneumococcal glycolipid vaccines based on the artificial oligosaccharide antigens of pneumococci type 3 and 23F and MPLA carrier. For this purpose, the structurally well-defined pneumococcal oligosaccharide antigens with different chain lengths will be first chemically synthesized using preactivation-based iterative one-pot and/or highly convergent glycosylation strategies, followed by incorporation with MPLA carrier via a bifunctional activated ester as linker, to furnish the fully synthetic MPLA-pneumococcal oligosaccharide conjugates. Moreover, the immunobiological studies on the these fully synthetic MPLA conjugates will be detailedly investigated by mice immunization, by antiserum titration and binding, by antibody-mediated complement-dependent cytotoxicity, and by the protection of mice against pneumococci infections, so as to find the effective anti-pneumococcal glycolipid vaccines. This research will not only yield novel fully synthetic MPLA conjugates or promising lead compounds worthy further investigations but also provide with the structure-activity relationships (SARs) of pneumococcal oligosaccharides as target antigens to guide further optimization of the anti-pneumococcal vaccine design. The result will have a major impact on antibacterial research.

肺炎链球菌（Streptococcus pneumoniae，SP）感染严重危害着人类的健康。基于SP细胞表面荚膜多糖抗原研制的多糖疫苗和糖蛋白疫苗在预防和治疗SP感染疾病方面发挥了重要作用。然而，随着耐药或多重耐药等高抗药性SP菌的出现，以及某些SP菌存在有效免疫应答率较低等问题，使得SP疫苗研制工作依然面临许多新的挑战和困难亟待解决。本项目结合最新发展的单磷酰类脂A（MPLA）为载体和免疫佐剂的全合成糖脂疫苗策略，以肺炎链球菌血清3和23F型为研究对象，采用MPLA为载体、人工合成的SP荚膜多糖重复片段为抗原，研发新型、高效和具有明确结构的全合成抗SP糖脂疫苗。项目研究方法采用化学法合成系列具有明确结构的各种长度不一的SP寡糖半抗原，通过双活化酯连接臂与MPLA进行缀合，获取制备系列具有确定结构的新型SP寡糖脂复合物，并进行相应免疫生物学和抗菌活性的研究，从而发现新型抗SP糖脂疫苗。

基于肺炎链球菌（Streptococcus pneumoniae，SP）细胞表面荚膜多糖抗原研制的多糖类疫苗和糖蛋白结合疫苗在预防和治疗SP感染疾病方面发挥了重要作用。 本项目选用肺炎链球菌包括3型（有效免疫应答率低）和23F型（高抗药性）的荚膜多糖为研究对象，通过化学法合成了一系列具有明确结构特征的各种长度不一的SP寡糖半抗原，包括SP3相关寡聚体五糖、六糖、七糖和八糖半抗原以及SP23F相关三糖和四糖半抗原；通过双活化酯连接臂将上述寡糖半抗原与具有免疫增强活性的载体蛋白TT进行缀合，制备了系列具有确定结构特征的半合成寡糖蛋白缀合物；通过点击化学将上述寡糖半抗原与新型单磷酰类脂A（MPLA）缀合，制备了系列具有确定结构的新型SP寡糖脂复合物；对上述制备的SP3寡糖-TT蛋白缀合物进行了一系列包括抗血清抗体滴度检测、抗血清与SP3菌体外结合和吞噬实验以及SP3菌体内冲击保护实验等免疫活性评价实验，揭示了SP3寡糖-TT蛋白缀合物能够在小鼠体内诱导具有保护作用的T细胞依赖性免疫反应。重要的是，与其他蛋白缀合物相比，SP3六糖-TT蛋白缀合物表现出了显著免疫活性，对SP3菌体内冲击起到显著的保护作用，因此推断SP3六糖是潜在的优选靶标抗原来发展新型抗SP3糖类疫苗。上述研究成果为进一步设计和优化下一代的新型半合成和全合成SP糖缀合物疫苗奠定实验基础。