Abstract LB-304: Customizable, in vitro protein glycosylation for antibacterial and anti-cancer vaccines

Abstract

Abstract Changes in protein glycosylation are important hallmarks of cancer biology, serving as biomarkers for cancer progression and predictors of cancer prognosis. Moreover, tumor-associated carbohydrate antigens (TACAs) provide potential targets for therapeutic and prophylactic interventions. However, development of anti-cancer immunotherapies and vaccines directed against TACAs has been limited due to the significant challenges associated with synthesizing proteins bearing defined glycan structures in vivo. These challenges arise due to the complex, non-templated nature of the glycosylation process and the inability to control glycosylation components at precise ratios in living cells. To address these challenges, we describe a novel cell-free glycoprotein synthesis (CFGpS) technology that seamlessly integrates protein biosynthesis with asparagine-linked (N-linked) protein glycosylation. The platform is flexible and modular, allowing the decoration of proteins with structurally diverse, yet homogeneous glycans, including the eukaryotic trimannosyl N-glycan (mannose3-N-acetylglucosamine2, Man3GlcNAc2), which forms the core of all tumor-associated N-glycans. We demonstrate the utility of this platform through the production of antibacterial glycoconjugate vaccines directed against the highly infectious pathogenic bacterium Franciscella tularensis. Importantly, we are able to synthesize and glycosylate a variant of E. coli maltose binding protein (MBP) that has been shown to elicit humoral and cell-based immunity to polysaccharide antigens in mice. Our work demonstrates a novel approach for the customizable glycosylation of immunogenic proteins with TACAs in a cell-free system. The ability of the CFGpS platform to synthesize defined glycan structures uniquely enables the production of novel antibacterial and anti-cancer vaccines. Citation Format: Jessica C. Stark, Thapakorn Jaroentomeechai, Rachel S. Dubner, Karen J. Hsu, Cameron Glasscock, Matthew P. DeLisa, Michael C. Jewett. Customizable, in vitro protein glycosylation for antibacterial and anti-cancer vaccines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-304.