SRP493249 PRJNA1083572 GSE260817 Time-dependent enhancement of mRNA vaccines by 4-1BB costimulatory antibodies mRNA vaccines have demonstrated efficacy against COVID-19. However, concerns regarding waning immunity and breakthrough infections have motivated the development of next-generation vaccines with enhanced efficacy. In this study, we investigated the impact of 4-1BB costimulation on immune responses elicited by mRNA vaccines in mice. We first vaccinated mice with an mRNA vaccine encoding the SARS-CoV-2 spike antigen like the Moderna and Pfizer-BioNTech vaccines, followed by administration of 4-1BB costimulatory antibodies at various times post-vaccination. Administering 4-1BB costimulatory antibodies during the priming phase did not enhance immune responses. However, administering 4-1BB costimulatory antibodies after 96 hours elicited a significant improvement in CD8 T cell responses, leading to enhanced protection against breakthrough infections. A similar improvement in immune responses was observed with multiple mRNA vaccines, including vaccines against common cold coronavirus, human immunodeficiency virus (HIV), and arenavirus. These findings demonstrate a time-dependent effect by 4-1BB costimulation and provide insights for developing improved mRNA vaccines. Overall design: Bulk RNA-Sequencing of SARS-CoV-2 specific CD8 T cells taken 7 days after vaccination of mice with an mRNA vaccine followed with treatment with a control antibody (n=4) or 4-1BB agonistic antibody (n=4). GSE260817 pubmed 39432667