(T1330-08-49) Continuous Processing of Liposomal Nanoparticles as Potential Reference Materials for Drug Product Development

Purpose: With the recent success of lipid nanoparticle delivery systems for nucleic acid therapeutics, such as mRNA COVID-19 vaccines, there is an anticipated increase in demand for liposomal/lipid-based therapeutics and vaccines. Liposomal reference materials would provide significant benefits to regulatory agencies and industry for further development, quality control, and technology transfer. The objectives of this project were to investigate long-term stability and characterization of candidate liposomal reference materials at different storage and processing conditions.

Methods: Candidate liposomal reference formulations were prepared with a coaxial turbulent jet in co-flow that uses an ethanol injection method¹. By changing fluid flow parameters, monodispersed liposomes at 50, 100, 150, 200 diameter nanometer (d.nm) with 5% charged lipid and 100 d.nm particles with 0.0%, 2.5%, 5.0% PEG-lipid were produced. All formulations were fully characterized using techniques such as dynamic light scattering for particle size and UPLC for lipid component and degradation product analysis. In addition, pH and Zeta-potential were monitored at selected time points. These characteristics were evaluated over 18 months at 25 °C and 4 °C storage conditions, with sampling each formulation monthly in triplicate.

Results: All samples stored for 18 months remained stable and homogeneous as the particle size remained below 2-5% error and the polydispersity index values remained below 0.13. While 5% charged lipid formulations were the most consistent, both the 2.5% and 5% PEG also remained below 5% change. The pH was stable for all samples, although the 2.5% and 5% PEG samples had a consistently lower pH when stored at 25°C than when stored at 4°C. Zeta potential remained as expected with the 5% charged lipid formulations around -40 mV and 2.5%, 5% PEG-lipid around -15 mV. Beyond 18 months, UPLC results showed cholesterol had no degradation while PG, DSPE-mPEG2000, and HSPC produced degradation products, which accounted for 5.5% of the total lipid (average mole percentage), mainly in the form of Lyso-18 and to a lesser extent Lyso-16 and Stearic Acid.

Conclusion: To develop liposomal reference materials, it is critical to ensure long-term stability consistent product characteristics, particularly particle size distribution. This report describes a stable liposomal particle system that has potential to be used in developing future liposomal reference materials. This system yields consistent quality attributes such as particle size distribution, while maintaining biological relevance.

References: 1. Costa., et al. Liposome Formation Using a Coaxial Turbulent Jet in Co-Flow. Pharm. Res, 2016. 404-416.

Acknowledgements: Funding for this project was made possible by a U.S. Food and Drug Administration grant (75F40120C002-01). This abstract reflects the views of the authors and should not be construed to represent FDA’s views or policies. Drs. Diane J. Burgess and Antonio P. Costa may have a potential conflict of interest with DIANT Pharma Inc., as some of the technology used in this work is licensed to DIANT.

Schematic of the injection port for continuous liposome formulation.

Long term particle size stability of seven liposmal formulations stored at 4°C.

Long term polydispersity data of seven liposomal formulations stored at 4°C.