Nucleoside modifications of in vitro transcribed mRNA to reduce immunogenicity and improve translation of prophylactic and therapeutic antigens

RNA molecules are prone to nuclease degradation and recognition by the innate immune system. Chemical modifications of the phosphate backbone, sugar, and/or nucleobase have helped increase resistance to degradation, while reducing recognition by immune sensors, and have proven crucial for the bench-to-bedside translation of several small RNA-based therapeutics (i.e., Onpattro and Givlaari). RNA molecules produced using in vitro transcription (IVT) demonstrated superior performance compared to other vaccine platforms gaining prominence in the fight against the COVID-19 pandemic (i.e., Comirnaty and Spikevax). This chapter discusses the elements of RNA recognition by innate immune sensors, the origins of the immunogenicity of in vitro transcribed RNA, as well as strategies to mitigate immunogenicity and improve translation of IVT-produced mRNA. We further discuss different nucleoside modifications and their influence on the capacity of RNA to activate the innate immune system and improve the therapeutic potential of mRNA.