Shielding of Lipid Nanoparticles for siRNA Delivery: Impact on Physicochemical Properties, Cytokine Induction, and Efficacy.

Akin Akinc,Benjamin Brigham,Yongfeng Jiang,Shannon Fishman,Pia V Kasperkovitz,Scott A Barros,June Qin,Richard G Duncan,Varun Kumar,Jayaprakash K Nair

doi:10.1038/mtna.2014.61

Akin Akinc, Benjamin Brigham

Open Access

https://doi.org/10.1038/mtna.2014.61

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Abstract

Formulation of short interfering RNA (siRNA) into multicomponent lipid nanoparticles (LNP) is an effective strategy for hepatic delivery and therapeutic gene silencing. This study systematically evaluated the effect of polyethylene glycol (PEG) density on LNP physicochemical properties, innate immune response stimulation, and in vivo efficacy. Increased PEG density not only shielded LNP surface charge but also reduced hemolytic activity, suggesting the formation of a steric barrier. In addition, increasing the PEG density reduced LNP immunostimulatory potential as reflected in cytokine induction both in vivo and in vitro. Higher PEG density also hindered in vivo efficacy, presumably due to reduced association with apolipoprotein E (ApoE), a protein which serves as an endogenous targeting ligand to hepatocytes. This effect could be overcome by incorporating an exogenous targeting ligand into the highly shielded LNPs, thereby circumventing the requirement for ApoE association. Therefore, these studies provide useful information for the rational design of LNP-based siRNA delivery systems with an optimal safety and efficacy profile.

Highlights

Recent progress in clinical development of RNA interference (RNAi) therapeutics has demonstrated that formulation of short interfering RNA into multi-component lipid nanoparticles (LNPs) is an effective strategy for liver delivery and silencing of therapeutically relevant gene targets.[1,2] Careful design of various parameters is critical for optimizing pharmacokinetic and pharmacodynamic parameters while minimizing potential for adverse effects due to activation of the immune response
Another method that is useful for assessing the cationic charge on particles is the hemolytic assay, which is based on the electrostatic interaction and fusion of LNPs with anionic membranes of red blood cells (RBCs), resulting in lysis of the RBCs
When we performed a series of studies in apoE−/− mice, we found that preincubation of LNPs with recombinant apolipoprotein E (ApoE) before injection rescued the activity of LNP1.5, whereas no improvement in efficacy was observed for LNP5

Summary

Introduction

Recent progress in clinical development of RNA interference (RNAi) therapeutics has demonstrated that formulation of short interfering RNA (siRNA) into multi-component lipid nanoparticles (LNPs) is an effective strategy for liver delivery and silencing of therapeutically relevant gene targets.[1,2] Careful design of various parameters is critical for optimizing pharmacokinetic and pharmacodynamic parameters while minimizing potential for adverse effects due to activation of the immune response. More recent clinical applications of RNAi typically employ siRNAs that contain chemical modification patterns that minimize immunostimulatory potential without interfering with gene silencing.[1,2,21,22,23] even when using modified siRNA within LNPs, minor and transient elevations of serum cytokines can still sometimes be observed both preclinically and clinically,[24,25] suggesting that the lipid components may contribute to the immunostimulatory potential of LNP-formulated siRNAs. To date, the study of the impact of PEG shielding in the context of LNP-mediated siRNA delivery has been limited[26] and the effect on proinflammatory cytokine induction is still unclear. We performed a systematic characterization of the effects of LNP PEG density on physicochemical properties, cytokine induction and delivery efficacy

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Results

Conclusion