Immunogenicity and Innate Immunity to High-Dose and Repeated Vaccination of Modified mRNA versus Unmodified mRNA

mRNA vaccines represent a new era with several novel constructs underway. We compared the responses of high doses and multiple repetitive immunizations of a nucleoside-modified mRNA construct to a sequence-codon optimized unmodified mRNA construct encoding the identical model antigen (HIV-1 gag). Rhesus macaques were immunized five times at two-week intervals, with a final boost 20 weeks later. At 24 hours post-vaccination both unmodified (160 µg) and modified (400 µg and 800 µg) mRNA constructs elicited clear but transient increase of plasmacytoid dendritic cells, intermediate CD14+ CD16+ monocytes and neutrophils along with secretion of type I IFN-related and inflammatory cytokines. Unmodified mRNA induced higher IL-7 and IFN-a levels, while modified mRNA induced higher IL-6 levels. Transcriptomic profiling showed significant upregulation of genes related to type I IFN signaling, antigen presentation, and innate immune activation induced by both mRNA constructs. The high dose modified mRNA induced a higher number of differentially expressed genes at prime which further increased after the fifth immunization. These differences in innate immune activation nonetheless led to similar levels and kinetics of gag-specific antibody and T cell responses. These findings offer insights into the immunogenic and reactogenic potential of different mRNA vaccine modalities, guiding future vaccine and therapy development.