(M1230-09-56) Enhancing Lipid Nanoparticles Shelf Life with a Vitamin A Derivative

Purpose: The objective of this study was to improve the stability and efficacy of lipid nanoparticles (LNPs) encapsulating messenger RNA (mRNA) by optimizing their formulation.

Methods: Lipid nanoparticles (LNPs) were formulated using the NanoAssemblr™ Ignite™ system (Cytiva, Marlborough, MA, USA). Each formulation included an ionizable lipid, PEGylated lipid, helper lipid, and cholesterol, with additional incorporation of various vitamins and vitamin derivatives. A modified luciferase mRNA was encapsulated to evaluate the stability and functional efficacy of the LNPs in human cells after 4 weeks of storage at 4°C. Only LNPs demonstrating acceptable physiochemical properties were tested and compared to a commercial mRNA vaccine formulation. Characterization was performed using a combination of light scattering techniques, including Dynamic Light Scattering (DLS) and Electrophoretic Light Scattering (ELS). The Zetasizer Nano ZS instrument (Malvern Panalytical, Worcestershire, UK) was used to measure mean hydrodynamic diameter, polydispersity index (PDI), and zeta potential. The mRNA concentration in the LNP formulations was quantified using a NanoDrop™ Spectrophotometer (Thermo Fisher Scientific, Basel, Switzerland). mRNA encapsulation efficiency (EE%) was assessed using the Quant-iT™ RiboGreen™ RNA assay (Thermo Fisher Scientific, Basel, Switzerland). Functional efficacy was measured with the Bright-Glo Luciferase Assay kit (Promega, Madison, WI).

Results: A range of lipophilic vitamins and derivatives were screened for their ability to enhance LNP-mRNA stability. Most compounds showed no significant ability to improve shelf-life after four weeks of storage at 4 °C. However, LNPs incorporating a vitamin A derivative demonstrated a marked increase in functional efficacy. Specifically, luciferase mRNA-loaded LNPs with this excipient exhibited up to six-fold increase in enzyme activity after four weeks of storage at 4°C compared to control LNPs lacking the vitamin A derivative.

Conclusion: Incorporation of a vitamin A derivate in a commercial mRNA vaccine LNP formulation significantly improved the stability and functional efficacy of luciferase mRNA after extended storage. These findings suggest that this vitamin A derivative could be considered as a stabilizing excipient to enhance shelf-life and performance of mRNA-based therapeutics.