Design and in vitro evaluation of an Fc-fusion epitope-based vaccine candidate against Helicobacter pylori.

Helicobacter pylori, a Group I carcinogen that infects over 50% of the global population, is a major contributor to gastric cancer and the growing crisis of antibiotic resistance. However, vaccine development has remained challenging due to the poor immunogenicity of candidate antigens and limitations in delivery strategies. To address this, we engineered a novel Fc-fusion vaccine targeting antigen-presenting cells (APCs) by fusing H. pylori UreB317-329 and UreB409-421 epitopes with Lpp20 lipoprotein to the murine IgG2a Fc domain (UreB: Lpp20: mFcγ2a). This study presents a preliminary, proof-of-concept evaluation of an Fc-fusion epitope-based vaccine candidate, focusing on in vitro characterization and APC-targeting properties. The construct was codon-optimized, expressed in Pichia pastoris, and purified via affinity chromatography. Expression levels in the mg/mL range, as determined by biochemical analysis, with >95% purity confirmed by SDS-PAGE and Western blot. Co-localization assays demonstrated 4.2-fold higher APC uptake of the Fc-fusion protein versus a His-tagged control (p < 0.001, unpaired t-test), mediated by FcγRI (CD64) binding. The protein maintained its structural stability for 30 days at 4 °C (<5% batch variability), underscoring manufacturing feasibility. While in vitro results validate APC targeting, limitations include preclinical scope and murine Fc compatibility gaps for human translation. This study established P. pastoris as a scalable platform for Fc-fusion vaccines, bypassing adjuvants and enhancing immunogenicity through conserved epitopes. Future work must evaluate in vivo efficacy in infection models and human Fc adaptations. These findings offer a blueprint for next-generation vaccines against H. pylori and other intracellular pathogens, urgently needed in antimicrobial resistance containment strategies.