Pulmonary delivery of small circular RNA vaccines for influenza prevention

Lipid nanoparticles (LNPs) have played an instrumental role in the delivery of RNA therapeutics and vaccines, including the emerging class of synthetic circular RNA (circRNA). Pulmonary vaccines hold the potential to prevent various respiratory infectious diseases, such as influenza caused by influenza infection. Here, we report the pulmonary delivery of LNPs loaded with highly stable small circRNA vaccine for influenza prevention. Upon intratracheal (i.t.) instillation of circRNA LNPs into mice, circRNA was efficiently retained in the lung and was delivered to antigen-presenting cells (APCs). To elicit immune responses against influenza, we designed and synthesized a small circRNA-M2e encoding matrix protein 2 ectodomain (M2e), a highly conserved but poorly immunogenic antigen from influenza A. I.t. administration of circRNA-M2e LNPs elicited robust anti-M2e immunity in both young adult mice and 18-month-old aged mice. As a result, pulmonary delivery of LNPs loaded with circRNA-M2e vaccine protected mice from lethal influenza challenge, without significant adverse side effects. Collectively, these results demonstrate the potential of pulmonary circRNA vaccines for respiratory infectious diseases such as influenza.