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Published on: February 1, 2019
Incorporation of mRNA in Lamellar Lipid Matrices for Parenteral Administration
Antje Ziller1, Sara S Nogueira1,2, Eva Hühn1
1Department of Pharmaceutics and Biopharmaceutics, Johannes Gutenberg University Mainz , 55099 Mainz, Germany.
Abstract:
Insertion of high molecular weight messenger RNA (mRNA) into lyotropic lipid phases as model systems for controlled release formulations for the mRNA was investigated. Low fractions of 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) were used as an anchor to load the mRNA into a lamellar lipid matrix. Dispersions of zwitterionic lipid in the aqueous phase in the presence of increasing fractions of mRNA and cationic lipid were prepared, and the molecular organization was investigated as a function of mRNA and cationic lipid fraction. Insertion of both cationic lipid and mRNA was clearly proven from the physicochemical characteristics. The d-spacing of the lipid bilayers, as determined by small-angle X-ray scattering (SAXS) measurements, responded sensitively to the amount of inserted DOTAP and mRNA. A concise model of the insertion of the mRNA in the lipid matrices was derived, indicating that the mRNA was accommodated in the aqueous slab between lipid bilayers. Depending on the DOTAP and mRNA fraction, a different excess of water was present in this slab. Results from further physicochemical characterization, including determination of free and bound mRNA, zeta potential, and calorimetry data, were in line with this assumption. The structure of these concentrated lipid/mRNA preparations was maintained upon dilution. The functionality of the inserted mRNA was proven by cell culture experiments using C2C12 murine myoblast cells with the luciferase-encoding mRNA. The described lipid phases as carriers for the mRNA may be applicable for different routes of local administration, where control of the release kinetics and the form of the released mRNA (bound or free) is required.
Insights
Messenger RNA (mRNA) was successfully loaded into lyotropic lipid phases using DOTAP as an anchor. These lipid-mRNA formulations show potential for controlled local drug delivery applications.
Area of Science:
- Biomaterials Science
- Drug Delivery Systems
- Nanotechnology
Background:
- Controlled release formulations are crucial for effective mRNA therapeutics.
- Lyotropic lipid phases offer a promising matrix for encapsulating biomolecules.
- Understanding the interaction between mRNA and lipid matrices is key for developing stable delivery systems.
Purpose of the Study:
- To investigate the insertion of high molecular weight messenger RNA (mRNA) into lyotropic lipid phases.
- To explore the use of 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) as an anchor for mRNA loading.
- To characterize the molecular organization and physicochemical properties of mRNA-lipid dispersions.
Main Methods:
- Preparation of lipid dispersions with varying fractions of mRNA and DOTAP.
- Small-angle X-ray scattering (SAXS) for structural analysis.
- Physicochemical characterization including zeta potential and calorimetry.
- Cell culture experiments with luciferase-encoding mRNA.
Main Results:
- Successful insertion of both DOTAP and mRNA into the lamellar lipid matrix was confirmed.
- SAXS measurements showed d-spacing sensitivity to DOTAP and mRNA fractions.
- A model indicated mRNA accommodation in the aqueous slab between lipid bilayers.
- Lipid-mRNA structures remained stable upon dilution.
- Functional luciferase expression was observed in C2C12 cells.
Conclusions:
- Lyotropic lipid phases effectively accommodate mRNA, with insertion occurring in the aqueous layer.
- DOTAP acts as an anchor, influencing the lipid-mRNA organization.
- These lipid-mRNA formulations are stable and maintain mRNA functionality.
- The system holds potential for controlled local administration of mRNA therapeutics.
