Lipid nanoparticle chemistry determines how nucleoside base modifications alter mRNA delivery

Area of Science:

• Biotechnology
• Molecular Biology
• Drug Delivery

Background:

• Therapeutic mRNA offers revolutionary disease treatment potential, accelerated by rapid vaccine development.
• Base modifications in mRNA can reduce immunogenicity and increase protein translation.
• However, base modifications do not consistently improve protein expression.

Purpose of the Study:

• To investigate how lipid nanoparticle (LNP) chemistry, formulation, and organ accumulation affect mRNA base modification efficacy.
• To compare the impact of five distinct base modifications and four ionizable lipids on protein expression in vivo.
• To determine the influence of delivery vehicle and target cell type on mRNA modification benefits.

Main Methods:

• Compared IV-injected LNPs with reporter mRNA containing five base modifications (ψ, m1ψ, m5U, m5C/ψ, m5C/s2U) and four ionizable lipids.
• Assessed protein expression levels in various organs, focusing on spleen and monocytic lineage splenocytes.
• Utilized firefly luciferase and erythropoietin mRNA constructs to evaluate modification effects.

Main Results:

• The m1ψ base modification generally yielded the greatest translation enhancement (up to 15-fold increase).
• Spleen exhibited the most dramatic expression improvements (up to 50-fold), linked to monocytic splenocytes.
• Enhancements were delivery vehicle-dependent and correlated with innate immunogenicity; benefits were limited outside the spleen.

Conclusions:

• mRNA base modification effectiveness is complex and highly dependent on LNP formulation, delivery vehicle, target cells, and organ-specific expression.
• The m1ψ modification shows promise for enhancing protein expression, particularly in the spleen.
• Optimizing LNP design is crucial for maximizing the therapeutic potential of modified mRNA.