A stable two-component cationic liposome platform for mRNA delivery induces CD8⁺ T-cell responses and protection in a...

The mRNA-LNP-based SARS-CoV-2 vaccines were a breakthrough for the technology; however, their production requires specialized equipment, and the complex formulation involving four distinct lipid classes imposes stringent requirements on both characterization and quality control. We tested a cationic adjuvant formulation containing a simple two-component liposome formulation based on dimethyldioctadecylammonium bromide and monomycoloyl glycerol-1 (CAF^®04) for mRNA-delivery, which can be complexed with mRNA by simple admixing on-site. The physicochemical characteristics of the resulting vaccine particles depended on the CAF04:mRNA N/P ratio, where an N/P ratio ≤ 1.09 resulted in anionic particles with little aggregation and N/P ratios ≥ 2.18 resulted in increasingly cationic particles that aggregated. These attributes influenced the magnitude of the CD8⁺ T-cell responses induced against OVA-encoding mRNA and at optimal CAF04:mRNA N/P ratios (0.76-1.42), the antigen-specific CD8⁺ T-cell responses reached magnitudes comparable to the DOTMA:DOPE Lipoplex (RNA-LPX) platform. Therapeutic vaccination using CAF04:mRNA delayed tumor growth and enhanced survival in an E.G7-OVA lymphoma model. We present a highly stable mRNA delivery platform that can be freeze-dried for extended storage and admixed with mRNA on-site to induce highly functional CD8⁺ T-cell responses with potential for therapeutic cancer vaccines.