Replacing cholesterol and PEGylated lipids with zwitterionic ionizable lipids in LNPs for spleen-specific mRNA translation

Oct 8, 2025Science advances

Using special ionizable lipids instead of cholesterol and PEGylated lipids in lipid nanoparticles to target mRNA delivery to the spleen

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mRNA Technology on OpenScience ↗PubMed ↗DOI ↗

Abstract

Replacing cholesterol and PEGylated lipids in LNPs with zwitterionic pyridine carboxybetaine results in ~70% lower liver accumulation.

The new three-component lipid nanoparticles (ThrCo LNPs) lead to a 4.5-fold increase in spleen-specific mRNA translation.
Zwitterionic pyridine carboxybetaine ionizable lipids enhance the hydrophilicity of LNPs, stabilizing their outer membrane.
Strong protonation at endosomal pH by PyCB groups aids in mRNA translation.
The zwitterionic surface reduces protein adsorption, potentially avoiding accelerated clearance from the bloodstream during repeated doses.
ThrCo LNPs may improve the delivery of mRNA to splenic antigen-presenting cells, enhancing immune responses.

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The spleen is emerging as a key vaccination target. However, existing lipid nanoparticles (LNPs) primarily accumulate in the liver, limiting their efficacy in vaccine therapy. The cholesterol in current LNP formulations promotes their uptake by hepatocytes, while the polyethylene glycol-modified (PEGylated) lipids induce PEG immunogenicity, further reducing the efficacy in the setting of repeated administrations. We develop a three-component (ThrCo) LNP by replacing cholesterol and PEGylated lipids in Pfizer-BioNTech LNPs with zwitterionic pyridine carboxybetaine (PyCB) ionizable lipids (ILs), achieving ~70% lower liver accumulation and a 4.5-fold increase in spleen-specific mRNA translation. PyCB ILs enhance LNP hydrophilicity, stabilizing the outer membrane to compensate for cholesterol removal. PyCB groups also exhibit strong protonation at endosomal pH, facilitating mRNA translation. The zwitterionic surface of ThrCo LNP reduces protein adsorption, thereby preventing the accelerated blood clearance effect caused by PEGylated lipids following repeated administrations. Thus, ThrCo LNP-based vaccines efficiently deliver mRNA to splenic antigen-presenting cells, boosting immune responses and improving therapeutic outcomes.