The valency of fatty acid conjugates impacts siRNA pharmacokinetics, distribution, and efficacy in vivo

Abstract

Lipid-conjugated small-interfering RNAs (siRNAs) exhibit accumulation and gene silencing in extrahepatic tissues, providing an opportunity to expand therapeutic siRNA utility beyond the liver. Chemically engineering lipids may further improve siRNA delivery and efficacy, but the relationship between lipid structure/configuration and siRNA pharmacodynamics is unclear. Here, we synthesized a panel of mono-, di-, and tri-meric fatty acid-conjugated siRNAs to systematically evaluate the impact of fatty acid structure and valency on siRNA clearance, distribution, and efficacy. Fatty acid valency significantly altered the physicochemical properties of conjugated siRNAs, including hydrophobicity and micelle formation, which affected distribution. Trivalent lipid-conjugated siRNAs were predominantly retained at the site of injection with minimal systemic exposure, whereas monovalent lipid-conjugated siRNAs were quickly released into the circulation and accumulated primarily in kidney. Divalent lipid-conjugated siRNAs showed intermediate behavior, and preferentially accumulated in liver with functional distribution to lung, heart, and fat. The chemical structure of the conjugate, rather than overall physicochemical properties (i.e. hydrophobicity), predicted the degree of extrahepatic tissue accumulation necessary for productive gene silencing. Our findings will inform chemical engineering strategies for enhancing the extrahepatic delivery of lipophilic siRNAs.