Collaborative Research: Cell-free glycoprotein synthesis technology for point-of-care vaccine biomanufacturing

耐药细菌对人类健康的威胁越来越大。到2050年，每年可能有多达1 000万人的生命受到威胁。需要采取新的战略来应对这一威胁。已经开发出疫苗来安全有效地预防危险的细菌感染。该项目旨在解决目前疫苗生产的局限性。将开发易于扩大规模的无细胞疫苗生产技术。这可能导致便携式，按需疫苗的开发和生产。该项目将推进结合疫苗的生物制造。这将使资源贫乏的社区更容易获得这些服务。与此同时，将开发实践学习模块，并提供给代表性不足的高中学生，本科生和研究生。细菌细胞表面装饰有复杂的糖结构。这些包括荚膜多糖（CPS）和0-多糖（0-PS）。这些结构是重要的毒力因子、粘附介质和免疫调节剂。CPS和O-PS结构通常不存在于宿主细胞的表面。因此，这些多糖可以配制成疫苗亚单位，用于保护人类免受危及生命的细菌感染。偶联疫苗是一种亚单位疫苗，其中多糖抗原连接到蛋白质载体。它们是诱导对病原菌免疫的最安全和最有效的方法之一。目前的生产技术在技术上很复杂，并且依赖于活细胞。冷藏在分销链的沿着每一步都是必要的。因此，制造业是集中的，资本密集型的，需要高技能的劳动力。该项目将创建一种可扩展的无细胞生物合成技术，用于生成偶联疫苗候选物。实验和计算方法将结合起来，开发和优化一锅无细胞糖蛋白合成系统。这些糖缀合物可以以冷冻干燥的形式储存，并通过添加水简单地重构。一种补充技术，鸟枪扫描糖突变，将使糖基化效率作为结合位点和抗原负载密度的函数，这将与疫苗免疫原性的综合评价。这一奖项反映了NSF的法定使命，并已被认为是值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估的支持。

Drug-resistant bacteria are a growing threat to human health. By the year 2050, up to 10 million lives per year could be at risk. New strategies will be needed to counter this threat. Vaccines have been developed to safely and effectively prevent dangerous bacterial infections. This project seeks to address current limitations in vaccine production. Cell-free technology for vaccine production that can be easily scaled up will be developed. This could lead to portable, on-demand vaccine development and production. The project will advance the biomanufacturing of conjugate vaccines. This would improve their availability to resource-poor communities. In parallel, hands-on learning modules will be developed and delivered to underrepresented high school students, and to undergraduate and graduate students. The bacterial cell surface is decorated with complex sugar structures. These include capsular polysaccharides (CPS) and O-polysaccharides (O-PS). These structures are important virulence factors, adhesion mediators, and immunomodulators. CPS and O-PS structures are typically absent from the surfaces of host cells. These polysaccharides can, therefore, be formulated as vaccine subunits and used to protect humans against life-threatening bacterial infections. Conjugate vaccines are a type of subunit vaccine whereby polysaccharide antigens are linked to a protein carrier. They are among the safest and most effective methods for inducing immunity against pathogenic bacteria. Current production technology is technically complex and relies on living cells. Refrigeration is necessary at every step along the distribution chain. As a result, manufacturing is centralized, capital intensive, and requires highly skilled labor. This project will create a scalable, cell-free biosynthesis technology for generating conjugate vaccine candidates. Experimental and computational approaches will be combined to develop and optimize a one-pot cell-free glycoprotein synthesis system. These glycoconjugates can be stored in freeze-dried formats and reconstituted simply by adding water. A complementary technology, shotgun scanning glycomutagenesis, will enable comprehensive evaluation of glycosylation efficiency as a function of conjugation site and antigen loading density, which will be correlated with vaccine immunogenicity.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.