World Vaccine Congress 2025

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Speaker: Tristan Kovačič, Project Manager, Sartorius BIA Separations
Time and Date: Thursday, April 24, 2025 | 9:40 AM
Title: Monolithic Columns for a High Recovery Purification Process of RNA-LNP Vaccines and Therapeutics

Abstract:

Lipid nanoparticles (LNPs) provide the most advanced platform for in vivo drug delivery of nucleic acids. However, they have not yet been developed into well-characterized biopharmaceuticals. Among others, challenges are of biological, manufacturing, and characterization type. Manufacturing challenges include proper formation of the drug product, maintaining the integrity of the drug product, achieving sufficient recovery, and obtaining a well-separated and characterized product.
Tangential flow filtration (TFF) has been implemented for the scale-up of downstream processing for the Comirnaty and the Spikevax vaccines. It is known in the field that TFF produces low recovery of the encapsulated RNA, ranging around 50%. As an alternative to the TFF process, a new separation process has been developed using Convective Interaction Media (CIM) OH monolithic columns. CIM monolithic columns are uniquely suitable for LNP purification due to the low shear stress imparted due to the purely laminar flow, and a wide selection of chemically modified surfaces. LNPs are loaded onto the columns under kosmotropic conditions, directly following the encapsulation process and neutralization. Elution with reducing conductivity produces a high recovery collection of particles. A robust recovery (based on RNA quantification) of >90% is achieved. Alternative elution conditions with cryopreservants enable freezing and long-term stable particles.
This process, which is also much faster than other comparable processes, achieves the desired concentration, ethanol removal, and buffer exchange functions. At the same time, free, non-encapsulated RNA is removed by tuning the buffers and achieving chromatographic separation. Additionally, subpopulations of the sample are collected with the most active portion of the drug product retained. Such collected particles show up to three-times higher in vitro protein expression and comparable particle toxicity. This result is robust across different experiments and different cell lines tested (HEK-293 and Caco2).
Additionally, analytical methods developed using monolithic columns enable the characterization of such formed drug products. They enable determination of mRNA quantity and purity inside LNPs, mRNA-lipid adduct quantity, and lipid purity.

Location: Washington D.C., USA

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