Delivery and Expression of mRNA in the Secondary Lymphoid Organs Drive Immune Responses to Lipid Nanoparticle-mRNA Vaccines after Intramuscular Injection

Lipid nanoparticles (LNPs) are the prime delivery vehiclefor mRNAvaccines. Previous hypotheses suggested that LNPs contribute to innatereactogenicity and lead to the establishment of a vaccine adaptiveresponse. It has not been clear whether LNP adjuvancy in the muscleis the prime driver of adaptive immune responses or whether deliveryto secondary lymphatic organs is necessary to induce strong adaptiveresponses. To address this, we formulated reporter gene (NLuc) orOVA mRNA into LNP or coadministered the mRNA with empty LNP. AfterIM injection, we correlated the delivery with adaptive immune responses.Additionally, we investigated humoral responses to modified mRNA encodingthe SARS-CoV-2 spike protein. Compared to unformulated mRNA encodingnanoluciferase, with or without co-administered empty LNPs, LNP-formulatedmRNA resulted in high levels of nanoluciferase in the secondary lymphoidorgans. Similarly, LNP-mRNA encoding ovalbumin led to a cellular immuneresponse against OVA while free mRNA, with or without empty adjuvantedLNPs, caused little or no immune response. Finally, only mice injectedwith LNP-formulated mRNA encoding SARS-CoV-2 spike protein elicitedrobust cellular and humoral immune responses. Our results suggestthat the mRNA delivery and transfection of secondary lymphatic organs,not LNP adjuvancy or RNA expression in muscle, are the main driversfor adaptive immune response in mice. This work informs the designof next-generation mRNA delivery systems where better delivery tosecondary lymphatic organs should lead to a better vaccine response.