Thesis

End-to-end optimization of lipid nanoparticle manufacturing for mRNA delivery

Creator
Rights statement
Awarding institution
  • University of Strathclyde
Date of award
  • 2025
Thesis identifier
  • T17308
Person Identifier (Local)
  • 202291971
Qualification Level
Qualification Name
Department, School or Faculty
Abstract
  • With the emergence of SARS-CoV-2 in 2020, mRNA vaccines have gained global attention. Currently, lipid nanoparticles (LNPs) are the most clinically advanced drug delivery system for the delivery of nucleic acids. Despite the extensive literature on LNPs in recent years, challenges persist regarding their development and production. In fact, most research papers focus on the therapeutic targets of LNPs, while less attention is given to understanding the challenges associated with their manufacturing, especially on an industrial-production scale, including scalability, reproducibility, encapsulation efficiency and long-term storage. This thesis focused on the end-to-end workflow of LNPs manufacturing, covering production, purification, and freeze-drying, while also addressing storage conditions. Beginning with LNP production, the effects of microfluidic parameters on LNP manufacturing were investigated while the preclinical scalable production of LNPs using various microfluidic devices was also evaluated. Moving on to purification, the second step of LNP manufacturing, the typical bottlenecks associated with this stage were assessed, with a focus on tangential flow filtration (TFF) as this method is commonly used on an industrial level. The effect of TFF speed and diafiltration volumes on LNPs characteristics were evaluated, along with the challenges related to scaling up the purification process. mRNA LNPs storage also represents a challenge due to the fragile nature of mRNA. With the aim of exploring lyophilisation as a technique for preserving mRNA LNPs, a series of freeze-drying cycles were conducted to identify the optimal parameters for producing mRNA LNPs with acceptable critical quality attributes (CQAs) and the in vitro and in vivo activity of the lyophilised product was evaluated to determine the effectiveness of the method. This thesis also explored the role of lipid selection in shaping the quality, stability, and performance of the final product. In particular, the contribution of PEGylated lipids having different alkyl chain lengths (DMG-PEG 2000 versus DSG-PEG 2000) to the physicochemical characteristics and performance of mRNA LNPs was investigated, as well as the impact in vitro and in vivo of the ionisable lipid (ALC-0315, DLin-MC3, and SM-102). The results presented demonstrate that all steps of LNP manufacturing influence the CQAs of the particles, from the choice of lipids, which can either limit or enhance their efficiency, to the selection of microfluidic parameters, buffers, purification methods, and lyophilisation conditions, highlighting the importance of carefully considering each individual step.
Advisor / supervisor
  • Perrie, Yvonne
Resource Type
DOI
Date Created
  • 2025
Funder

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