Towards a New Generation of Glycoengineered Pneumococcal Bioconjugate Vaccines

项目总结 肺炎球菌结合疫苗(PCV)，由肺炎球菌多糖共价连接到 载体蛋白，是用于预防肺炎球菌疾病的救命避孕药。重要的是，PCV提供了 所有年龄段的人，包括婴儿和两岁以下的儿童都享有豁免权，这不是纯粹的 多糖类疫苗。像所有的结合疫苗一样，PCV是用化学结合法生产的， 这是出了名的复杂，劳动密集型，最终阻碍了更好的版本的开发， 提供对更多致病血清型的免疫力。例如，PCV Prevnar 13在#年获得许可 2010年，只对13种肺炎球菌血清型有保护作用；而纯粹的多糖疫苗， 该病毒于1983年获得许可，可抵抗23种血清型。Pappovax 23被批准用于 对老年人的保护；然而，它不为婴儿和儿童提供保护。因此，三十多年来， 婴儿和儿童还没有预防20多种引起肺炎球菌疾病的疫苗选择 血清型。为了提供20多价的PCV供所有年龄段使用，VaxNewMo一直在开发一种 制造肺炎球菌和其他结合疫苗的方法，其绕过对 化学结合，在称为生物结合的过程中利用原核糖基化系统。 VaxNewMo的专利生物结合平台依赖于结合酶来转移细菌 多糖，就像肺炎链球菌的荚膜多糖一样，在实验室安全的情况下变成载体蛋白 细菌E.Coli.由于生物偶联是一种酶驱动的技术，因此产生的偶联物是 均一且易于提纯。重要的是，生物结合可以用来迅速产生过多的 只需通过引入编码不同血清型的遗传信息即可对抗多种血清型 肺炎球菌的血清型转化为具有生物结合能力的大肠杆菌菌株。因此，生物偶联可用于 一种涵盖20多个血清型的PCV的简化开发。在这个项目的第一阶段，我们 成功建立了VaxNewMo生物偶联平台可以产生PCV的原理证明 含有常规疫苗携带者。此外，VaxNewMo的PCV既具有免疫原性，又具有保护性 对抗肺炎链球菌疾病。第二阶段SBIR应用的拟议研究将集中在(目标1) 建立大规模生产VNM8的生物处理能力 生物结合物。建立单一血清型生物结合物的生物处理程序是重要的第一步 迈向商业规模生产，并将有助于简化未来其他 肺炎球菌生物结合物。随后，(目标2)我们将确认VNM8是大容量生产的 使用体外和体内模型，生物过程是保护性的。此外，(目标3)我们将生成一个大肠杆菌文库 能够产生24价PCV的菌株。对于第二阶段B，将寻求配制一种24价PCV(24v气动) 用于药代动力学和毒性研究，并为IND前与FDA的会议做准备。

PROJECT SUMMARY Pneumococcal conjugate vaccines (PCVs), composed of a pneumococcal polysaccharide covalently linked to a carrier protein, are life-saving prophylactics used to prevent pneumococcal disease. Importantly, PCVs provide immunity for all age groups, including, infants and children under the age of two, which is not the case for purely polysaccharide vaccines. Like all conjugate vaccines, PCVs are manufactured using chemical conjugation, which is notoriously complex, labor intensive, and ultimately hinders the development of better versions that provide immunity to more disease-causing serotypes. As an example, the PCV, Prevnar 13, was licensed in 2010 and only protects against 13 pneumococcal serotypes; whereas, the purely polysaccharide vaccine, Pneumovax 23, was licensed in 1983 and protects against 23 serotypes. Pneumovax 23 is approved for use in the elderly; however, it does not provide protection to infants and children. Thus, for more than three decades, infants and children have not had a vaccine option that protects against 20+ disease causing pneumococcal serotypes. In order to provide a 20+ valent PCV for use in all age groups, VaxNewMo has been developing a method for manufacturing pneumococcal and other conjugate vaccines that bypasses the dependency of chemical conjugation and instead exploits prokaryotic glycosylation systems in process termed bioconjugation. VaxNewMo’s proprietary bioconjugation platform relies on a conjugating enzyme to transfer a bacterial polysaccharide, like a pneumococcal capsular polysaccharide, to a carrier protein all within the lab safe bacterium E. coli. Since bioconjugation is an enzyme driven technology, the conjugates produced are homogenous and readily purified. Importantly, bioconjugation can be used to rapidly produce a plethora of conjugates against many serotypes simply by introducing the genetic information encoding for a different pneumococcal serotype into a bioconjugation competent strain of E. coli. Thus, bioconjugation can be used for the streamlined development of a PCV covering more than 20 serotypes. In Phase I of this project, we successfully established proof of principle that VaxNewMo’s bioconjugation platform could generate PCVs containing conventional vaccine carriers. Moreover, VaxNewMo’s PCVs were both immunogenic and protective against pneumococcal disease. The proposed research in this Phase II SBIR application will focus on (Aim 1) establishing bioprocessing capabilities for large volumetric production of VNM8, a serotype 8 pneumococcal bioconjugate. Establishing bioprocessing procedures for a single serotype bioconjugate is an important first step towards commercial scale production and will help streamline future upstream processing for other pneumococcal bioconjugates. Subsequently, (Aim 2) we will confirm that VNM8 produced in a large volumetric bioprocess is protective using in vitro and in vivo models. In addition, (Aim 3) we will generate a library of E. coli strains capable of producing a 24 valent PCV. For Phase IIB, will seek to formulate a 24 valent PCV (24vPneumo) for pharmacokinetic and toxicity studies and prepare for pre-IND meetings with the FDA.