Cell-free synthesis of influenza virus-like particles (VLPs) as prototyping platform for vaccine development and variant characterization

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SUMMARY The absence of prophylactic and low-cost influenza vaccines along with slow production rate makes us unprepared for the next pandemic outbreak worldwide. Recombinant virus-like particles (VLPs) vaccines provide us with unique opportunities to address these difficulties, but their recombinant synthesis strategies are limited to cell culture production, which has its challenges such as limited throughput and architectural heterogeneity of the obtained products. In this project, we aim to combine wheat germ cell-free protein expression platform with active microsomes of different origins (including human) to synthesize glycosylated influenza virus-like particles (VLPs). These particles will further undergo detailed morphological and compositional examination using electron microscopy and native mass spectrometry to understand the architecture of obtained products and understand the mechanisms of their assembly and composition variability. High- resolution tomography will be utilized to determine the best VLP candidates with maximal antigen occupancy and thus maximal vaccine efficiency. We hypothesize that this approach will be suitable for a stable and streamlined generation of Influenza VLPs and will offer much more flexibility and control over their architectural heterogeneity and eventually serve as a rapid prototyping platform for influenza vaccine development. The current project will also inform us about the utility of using such approaches for virology and for glycoprotein synthesis in general.