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

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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.