In vivo conjugated multivalent toxoid-polysaccharide vaccine for S. aureus

描述(申请人提供)：金黄色葡萄球菌是一种革兰氏阳性人类病原体，可引起多种感染，从皮肤和软组织感染(SSTI)到危及生命的败血症和肺炎。金黄色葡萄球菌的致病性依赖于许多毒力因子，包括细胞表面蛋白和多糖，以及分泌的毒素。其中一类重要的毒素包括溶血素和杀白素等毛孔形成毒素，以及对红细胞和白细胞具有裂解活性的小肽毒素，特别是代表金黄色葡萄球菌主要防御线的中性粒细胞。这些毒素会造成组织损伤，促进细菌在远处器官的传播和生长，并使病原体能够逃避宿主的先天免疫反应。此外，荚膜多糖(CP)具有保护细菌免受多形核白细胞吞噬作用的作用，已被证实是重要的疫苗靶标。综合生物治疗公司(IBT)正在与CPS结合开发基于毒素的疫苗，以对抗金黄色葡萄球菌感染。致孔溶血素(Hla)，又称β-毒素(AT)，几乎所有菌株都能产生，与多种金黄色葡萄球菌侵袭性疾病有关。最近对金黄色葡萄球菌菌血症的微生物学和血清学研究也表明毒素和苯酚可溶性调节素(PSM)在金黄色葡萄球菌致病中的重要性。根据我们最近的研究，预先存在的针对人类白细胞抗原、毒素和PSM？3的抗体显著降低了患有金黄色葡萄球菌菌血症的成年人的败血症风险。这一阶段SBIR的目标是评估一种新型糖结合葡萄球菌毒素疫苗的可行性。已经设计的候选疫苗代表了融合毒素和PSM？3的人类白细胞抗原A(AT62)N末端的一个关键结构域。我们以前已经证明AT62疫苗对菌血症和肺炎具有保护作用。这项建议利用GlycoVaxyn公司开发的独特和新颖的体内生物结合技术，将这种杂交类毒素与胶囊多糖5型(CP5)结合，作为生物结合物有效性的概念证明。这项生物结合技术涉及到工程设计的大肠杆菌细胞，以表达弯曲杆菌酶PglB以及金黄色葡萄球菌CP8生物合成所需的基因。PglB将N-连接的CP8链转移到引入这些细胞的疫苗构建物上。这项新技术是糖结合疫苗领域的一项重大发展，与传统的化学结合相比具有多种优势。在具体目标1中，将产生三种毒素的融合，并识别减弱突变并将其并入融合构建体中。候选疫苗将进行免疫原性测试，然后在目标2中产生生物结合物。在目标3中，将在代表菌血症/败血症和皮肤和软组织感染的两种主要疾病模型中评估候选疫苗的保护效果。在这项拟议的研究成功完成后，我们设想了第二阶段的SBIR，其中将为另一种主要的金黄色葡萄球菌壳多糖CP8生产类似的生物结合物，并对多价疫苗的效力(包括我们小组目前正在开发的其他毒素疫苗的组合)进行广泛的测试。最终目标是与大型制药公司建立战略合作伙伴关系，将候选疫苗转化为临床开发。

DESCRIPTION (provided by applicant): Staphylococcus aureus is a Gram-positive human pathogen that causes a wide range of infections from skin and soft tissue infections (SSTI) to life threatening sepsis and pneumonia. The pathogenicity of S. aureus is dependent on numerous virulence factors, including cell surface proteins and polysaccharides, as well as secreted toxins. An important group of these toxins includes pore forming toxins such as hemolysins and leukocidins, as well as small peptidic toxins, with lytic activity towards erythrocytes and leukocytes particularly the neutrophils that represent the major line of defense against S. aureus. These toxins cause tissue damage, promote bacterial dissemination and growth in distant organs, and enable the pathogen to evade the host innate immune response. Furthermore, capsular polysaccharide (CP) are known to protect bacteria from phagocytic activity of polymorphonuclear leukocytes and have been validated as important vaccine targets. Integrated BioTherapeutics (IBT) is pursuing development of toxin- based vaccines in combination with CPs against S. aureus infections. The pore-forming ?-hemolysin (Hla), also known as ?-toxin (AT), is produced by nearly all strains and is implicated in several S. aureus invasive diseases. Recent microbiological and serological studies in humans with S. aureus bacteremia also show the importance of ¿-toxin and phenol-soluble modulins (PSM) for S. aureus pathogenicity. Based on our recent study, pre-existing antibodies against Hla, ¿-toxin and PSM?3 significantly reduce the risk of sepsis in adults with S. aureus bacteremia. The goal of this Phase I SBIR is to evaluate the feasibility of a novel glycoconjugate staphylococcal toxin based vaccine. Vaccine candidates have been designed that represent a critical structural domain at the N terminus of Hla (AT62) fused to ¿-toxin and PSM?3. We have shown previously that AT62 vaccine provides protection against bacteremia and pneumonia. This proposal utilizes a unique and novel in vivo bioconjugation technology developed by Glycovaxyn Inc. to conjugate this hybrid toxoid with capsular polysaccharide Type 5 (CP5) as proof of concept of the efficacy of the bioconjugate. The bioconjugation technology involves E. coli cells engineered to express the Campylobacter enzyme PglB as well as the genes required for S. aureus CP8 biosynthesis. PglB transfers N-linked CP8 chains to the vaccine construct introduced into these cells. This novel technology is a major development in glycoconjugate vaccine field with multiple advantages over the conventional chemical conjugation. In the Specific Aim 1 fusions of the three toxins will be generated and attenuating mutations identified and incorporated into the fusion construct. The candidate vaccine will be tested for immunogenicity and then bioconjugates generated in Aim 2. In Aim 3 the protective efficacy of the vaccine candidate will be evaluated in two major disease models representing bacteremia/sepsis and skin and soft tissue infections. Upon successful completion of this proposed research we envision a Phase II SBIR in which similar bioconjugate will be produced for the other major S. aureus capsular polysaccharide CP8 and the efficacy of the multivalent vaccine (including combination other toxoid vaccines currently under development in our group) extensively tested. The ultimate goal is to build a strategic partnership with large pharma to transition the vaccine candidate into clinical development.