Non-linear enhancement of mRNA delivery efficiencies by influenza A derived NS1 protein engendering host gene inhibition property

Nucleic acid induced immunogenicity remains a significant impediment in biomedical therapeutics because the innate immune system is a complex network overlaid with functional redundancies. Herein we report that non-structural protein 1 (NS1), an immune evasion protein derived from influenza A virus, when co-delivered in mRNA format is a potent mRNA transfection enhancer without toxicity. Transfection enhancement is mediated by NS1's effector domain through inhibition of IRF3 and PKR, activators of early anti-viral responses as well as CPSF30, a non immunostimulating protein. Importantly, host gene inhibition mediated via CPSF30 inhibition is a highly effective immune evasion mechanism because it blocks de novo gene expression non-specifically and inhibits global anti-viral responses during mRNA transfection. We show that only NS1 with CPSF30 inhibition property can enhance modified mRNA transfections. Furthermore, transfection efficiency of unmodified mRNA, if co-delivered with NS1-TX91 mRNA, can exceed that of modified mRNA in HepG2, RAW 264.7 and HeLa cells. The novel impact of NS1-TX91 lays the foundation of a virus inspired immune evasion genes co-delivery approach that can address problems arising from RNA immunogenicity for non-vaccine mRNA therapeutics in an affordable and scalable way. It is also transferable to applications that benefits from active inhibition of material-induced immunogenicity.

191