Exploring the impact of commonly used ionizable and pegylated lipids on mRNA-LNPs: A combined in vitro and preclinical perspective

Nov 9, 2024Journal of controlled release : official journal of the Controlled Release Society

How common charged and coated fats affect mRNA lipid nanoparticles in lab and animal studies

AI simplified

mRNA Technology on OpenScience ↗PubMed ↗DOI ↗

Abstract

All lipid nanoparticle (LNP) formulations exhibited similar critical quality attributes, including particle size <100 nm and high encapsulation efficiency (>90%).

The potency of LNPs varied significantly based on the type of ionizable lipid used, with SM-102 showing the highest in vitro mRNA expression.
In vivo studies indicated that both SM-102 and ALC-0315 LNPs resulted in significantly higher mRNA expression compared to DLin-MC3-DMA, DODAP, and DOTAP LNPs.
The choice of PEG lipid influenced mRNA expression levels, with DSPE-PEG2k associated with reduced expression, although it did not affect other quality attributes or clearance from the injection site.
Critical quality attributes (CQAs) serve as indicators of the quality of LNP production but do not correlate with LNP potency.
Standard in vitro studies may not adequately reflect in vivo potency, highlighting the complexities of formulation design.

AI simplified

Ionizable lipids are widely recognized as the crucial component of lipid nanoparticles (LNPs). They enable mRNA encapsulation, shield it from enzymatic degradation, facilitate cellular uptake, and foster its cytosolic release for subsequent translation into proteins. In addition, PEGylated lipids are added to stabilize the particles in storage and in vivo. In this study, we investigate the potency of LNPs prepared using commonly adopted ionizable and pegylated lipids in vitro (using HEK293 cells) and in vivo (mouse studies) to consider the impact of structure on potency. LNPs were prepared using a fixed molar ratio of DSPC: Cholesterol: ionizable/cationic lipid: PEG lipid (10:38.5:50:1.5 mol%). All LNP formulations exhibited similar critical quality attributes (CQAs), including particle size <100 nm, low PDI (<0.2), near-neutral zeta potential, and high encapsulation efficiency (>90%). However, the potency of these LNPs, as measured by in vitro mRNA expression and in vivo expression following intramuscular injection in mice varied significantly. LNPs formulated with SM-102 exhibited the highest expression in vitro, whilst in vivo SM-102 and ALC-0315 LNPs showed significantly higher mRNA expression than DLin-MC3-DMA (MC3), DODAP and DOTAP LNPs. We also investigated the effect of PEG lipid choice (ALC-0159, DMG-PEG2k, and DSPE-PEG2k), which did not impact LNP CQAs, nor their clearance from the injection site. However, PEG lipid choice significantly influenced mRNA expression with the incorporation of DSPE-PEG2k reducing expression. This work contributes valuable insights to the evolving landscape of mRNA research, emphasizing that CQAs are a marker of the quality of the LNP production process, but not discriminatory regarding LNP potency. Similarly, standard in vitro studies do not provide insights into in vivo potency. These results further emphasize the intricacies of formulation design and the importance of bridging gaps between experimental outcomes in different settings.

Full Text

Full text is available at the source.

View full text (XML) ↗View full text ↗

Exploring the impact of commonly used ionizable and pegylated lipids on mRNA-LNPs: A combined in vitro and preclinical perspective

Abstract

Full Text

You found one interesting study. We’ll send the next 7.

what lands in your inbox each week:

Recent issues from the mrna technology brief

Abstract

Full Text

Related papers

You found one interesting study. We’ll send the next 7.

what lands in your inbox each week:

Recent issues from the mrna technology brief