Optimized LNP-encapsulated IVT mRNA as a robust platform for the development of next-generation cancer immunotherapies 2309761

Introduction Chimeric antigen receptor T cell therapy (CAR-T) and therapeutic cancer vaccines show great promise as next-generation cancer immunotherapies. However, these approaches traditionally rely on plasmid DNA or viral-vector-based gene delivery systems which are limited by potential safety risks and suboptimal antigen expression. Lipid nanoparticle (LNP)-encapsulated in-vitro-transcribed (IVT) mRNA offers a safer alternative while facilitating enhanced targeting, antigen expression, and overall therapeutic efficacy. Methods IVT mRNA vectors were rationally designed and vector components optimized for enhanced expression. Modified nucleotides were incorporated into the RNA during in vitro transcription prior to encapsulation with optimized LNP formulations. To assess efficacy versus other approaches, we compared antigen expression of our optimized LNP-mRNA to a protein-based vaccine in mice. Finally, we tested anti-CD19 CAR-T cells generated using LNP-mRNA in a cytotoxicity assay, and evaluated the anti-tumor effects of an LNP-mRNA anti-TRP2 vaccine in vivo. Results Through considered vector design and optimization, we successfully engineered IVT mRNAs with enhanced stability and translation efficiency, achieving robust transgene expression both in vitro and in vivo. By incorporating modified nucleotides and optimizing LNP formulations, we reduced unwanted host immune responses that might otherwise hamper therapeutic efficacy. Compared with a recombinant protein-based vaccine, optimized LNP-mRNA was more effective, eliciting markedly higher antibody titers against a complex transmembrane antigen. Functionally, anti-CD19 CAR-T cells engineered using LNP-mRNA exhibited potent cytotoxicity against target cells ex vivo, and mRNA-based anti-TRP2 vaccines inhibited melanoma growth in vivo. Conclusion Overall, these findings highlight the potential of optimized LNP-encapsulated IVT mRNA as a platform for engineering safer and more effective cancer immunotherapies.