Targeted Delivery of RNAi Therapeutics With Endogenous and Exogenous Ligand-Based Mechanisms

Abstract

Lipid nanoparticles (LNPs) have proven to be highly efficient carriers of short-interfering RNAs (siRNAs) to hepatocytes in vivo; however, the precise mechanism by which this efficient delivery occurs has yet to be elucidated. We found that apolipoprotein E (apoE), which plays a major role in the clearance and hepatocellular uptake of physiological lipoproteins, also acts as an endogenous targeting ligand for ionizable LNPs (iLNPs), but not cationic LNPs (cLNPs). The role of apoE was investigated using both in vitro studies employing recombinant apoE and in vivo studies in wild-type and apoE(-/-) mice. Receptor dependence was explored in vitro and in vivo using low-density lipoprotein receptor (LDLR(-/-))-deficient mice. As an alternative to endogenous apoE-based targeting, we developed a targeting approach using an exogenous ligand containing a multivalent N-acetylgalactosamine (GalNAc)-cluster, which binds with high affinity to the asialoglycoprotein receptor (ASGPR) expressed on hepatocytes. Both apoE-based endogenous and GalNAc-based exogenous targeting appear to be highly effective strategies for the delivery of iLNPs to liver.

Multiparametric approach for the evaluation of lipid nanoparticles for siRNA delivery

“…The FVII model was primarily chosen for this evaluation because previous work from our laboratory and others have demonstrated that ionizable LNPs, similar to those prepared in this work, target liver hepatocytes presumably due to interactions with endogenous lipoproteins (25). All 32 members of the lipid library were formulated into LNPs with siRNA against FVII and administered i.v.…”

“…LNPs were used in this study because they have been shown to facilitate efficient delivery to hepatic and immune targets (23,(25)(26)(27). In the past few years, LNPs formulated with siRNAs against transthyretin (TTR) and proprotein convertase subtilisin/kexin type 9 (PCSK9) in rodents and nonhuman primates have produced promising preclinical results and several clinical trials involving LNPs are currently underway (28)(29)(30).…”

RNAi-mediated silencing of hepatic Alas1 effectively prevents and treats the induced acute attacks in acute intermittent porphyria mice

“…To identify a potent Alas1-siRNA, a panel of siRNAs targeting Alas1 was initially screened for their ability to inhibit Alas1 mRNA expression in cultured hepatic cells. The most active compounds were formulated into lipid nanoparticles (LNPs) for efficient hepatic delivery (21)(22)(23) and evaluated for their ability to down-regulate hepatic Alas1 mRNA expression in wild-type mice. The selected Alas1-siRNA was then evaluated in the mouse model for AIP for its effectiveness to prevent or treat an induced acute attack.…”

The Niemann-Pick C1 Inhibitor NP3.47 Enhances Gene Silencing Potency of Lipid Nanoparticles Containing siRNA

“…The endocytosis of LNP-siRNA systems containing ionizable cationic lipids is dependent on association with apolipoprotein E (ApoE) and proceeds by the classic clathrin-mediated endocytosis mechanism used for uptake of cholesterol-containing LDL. 20 However, it has been noted that low NPC1 levels result in reduced internalization and lower transfection efficiencies for a polymer-based gene delivery system which enters cells primarily through caveolaemediated endocytosis. 21 Thus, the benefits of NP3.47 on LNPsiRNA intracellular accumulation and gene silencing potencies may depend on the particular endocytotic mechanism employed.…”