title: Optimization of Lipid Nanoparticle Formulations for mRNA Delivery in Vivo with Fractional Factorial and Definitive Screening Designs.
abstract: Intracellular delivery of messenger RNA (mRNA) has the potential to induce protein production for many therapeutic applications. Although lipid nanoparticles have shown considerable promise for the delivery of small interfering RNAs (siRNA), their utility as agents for mRNA delivery has only recently been investigated. The most common siRNA formulations contain four components: an amine-containing lipid or lipid-like material, phospholipid, cholesterol, and lipid-anchored polyethylene glycol, the relative ratios of which can have profound effects on the formulation potency. Here, we develop a generalized strategy to optimize lipid nanoparticle formulations for mRNA delivery to the liver in vivo using Design of Experiment (DOE) methodologies including Definitive Screening and Fractional Factorial Designs. By simultaneously varying lipid ratios and structures, we developed an optimized formulation which increased the potency of erythropoietin-mRNA-loaded C12-200 lipid nanoparticles 7-fold relative to formulations previously used for siRNA delivery. Key features of this optimized formulation were the incorporation of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) and increased ionizable lipid:mRNA weight ratios. Interestingly, the optimized lipid nanoparticle formulation did not improve siRNA delivery, indicating differences in optimized formulation parameter design spaces for siRNA and mRNA. We believe the general method described here can accelerate in vivo screening and optimization of nanoparticle formulations with large multidimensional design spaces.
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Kauffman
authors.forename: J Robert
authors.initials: JR
authors.lastname: Dorkin
authors.forename: Jung H
authors.initials: JH
authors.lastname: Yang
authors.affiliation: Shire Pharmaceuticals, Lexington, Massachusetts 02421, United States.
authors.forename: Michael W
authors.initials: MW
authors.lastname: Heartlein
authors.affiliation: Shire Pharmaceuticals, Lexington, Massachusetts 02421, United States.
authors.forename: Frank
authors.initials: F
authors.lastname: DeRosa
authors.forename: Faryal F
authors.initials: FF
authors.lastname: Mir
authors.forename: Owen S
authors.initials: OS
authors.lastname: Fenton
authors.forename: Daniel G
authors.initials: DG
authors.lastname: Anderson
mesh_terms: D045744:Cell Line, Tumor; D018014:Gene Transfer Techniques; D006801:Humans; D008055:Lipids / Q000008:administration & dosage / Q000737:chemistry*; D008081:Liposomes / Q000008:administration & dosage / Q000737:chemistry; D008099:Liver / Q000187:drug effects; D053758:Nanoparticles / Q000008:administration & dosage / Q000737:chemistry*; D010714:Phosphatidylethanolamines / Q000008:administration & dosage / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry; D012333:RNA, Messenger / Q000008:administration & dosage* / Q000737:chemistry; D014162:Transfection
chemical_list: D008055:Lipids; D008081:Liposomes; D010714:Phosphatidylethanolamines; D012333:RNA, Messenger; C094877:dioleoyl phosphatidylethanolamine; D011092:Polyethylene Glycols
keywords: Lipid nanoparticle; design of experiment; in vivo; mRNA; nucleic acid