(F-230) Engineering multiply adjuvanted mRNA vaccines through lipid and antigen design

Introduction:: The successful development of mRNA-based vaccines against SARS-CoV-2 has provoked broad interest in RNA-based technologies and demonstrated the promising potential of mRNA as a new vaccine class for combating infectious diseases. The widespread clinical translation of this technology requires overcoming certain limitations including eliciting a robust, protective immune response to the expressed antigen while ideally minimizing the side effects associated with vaccination. To further improve the safety and efficacy of mRNA vaccines, we here develop a dually self-adjuvanted mRNA vaccine system whereby both the encapsulating lipid nanoparticle as well as the mRNA-encoded antigen are designed to potentiate the immune response to mRNA-LNP vaccination.

Materials and Methods:: A novel, biodegradable library of ionizable lipids was tested both in vitro and in vivo for their transfection efficiency and ability to stimulate the immune system. Concurrently, an mRNA transcript was developed to encode an antigen combined with a bio-inspired adjuvanting peptide (AP). The efficacy of this adjuvanted construct in safely enhancing antigen-specific immune responses was evaluated in mice. Mice were then vaccinated both intramuscularly and intranasally using a combination of the adjuvanted mRNA transcript and immunostimulatory lipids, and the resulting immune responses were assessed.

Results, Conclusions, and Discussions:: Through screening a combinatorial library of 480 biodegradable ionizable lipids as part of LNP formulations for their potential as vaccines, a top-performing ionizable lipid was identified that exhibited enhanced immune stimulation resulting in potentiated immune response compared to FDA-approved ionizable lipids MC3 and ALC-0315. Next, vaccination with an adjuvanted mRNA encoding a fusion protein consisting of a SARS-CoV-2 antigen and a bio-inspired adjuvanting peptide (AP), within a single transcript, increased the magnitude of antigen-specific antibody titers by at least ten-fold in mouse sera for both wild-type SARS-CoV-2 and Delta variant antigens compared to mRNA encoding viral antigens alone without elevations in systemic inflammation. Vaccination with dually adjuvanted LNPs combining the immunostimulatory ionizable lipid and fusion mRNA transcript resulted in synergistic immune responses to the Delta variant of SARS-CoV-2 for both intramuscular and intranasal administrations. These results demonstrate that a multiply-adjuvanted system, whereby adjuvant properties are engineered into both the lipid carriers and the mRNA-encoded antigen, holds great potential in improving the efficacy, safety, and ease of administration of mRNA vaccines.The development of a multiply adjuvanted mRNA vaccine system, as presented in this study, holds many implications for future vaccine development. Our approach, which combines immunostimulatory lipids and an adjuvanted mRNA transcript encoding an antigen-adjuvanting peptide fusion, broadens the combinatorial design space available for engineering mRNA vaccines which carefully balance immunogenicity with reactogenicity. By achieving an enhanced immune response against SARS-CoV-2 and demonstrating promising safety profiles, this research could pave the way for improved protection against existing and emerging infectious diseases including influenza and other SARS-CoV-2 variants. Furthermore, the versatility of the mRNA platform and the possibility of intranasal administration suggest a more adaptable and user-friendly vaccination strategy, increasing the potential impact of this novel system on global public health.

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