A Platform Approach to mRNA Purification by Tangential Flow Filtration: Application to a Pentavalent Norovirus Vaccine.
- 1Vaccines & Advanced Biotechnologies Process R&D, Merck & Co., Inc., Rahway, NJ 07065, USA.
Abstract
Background/objectives
Multivalent mRNA vaccines are being evaluated for prevention of norovirus infection caused by multiple circulating strains. Platform purification processes may accelerate development and reduce manufacturing complexity by enabling multiple mRNA constructs to be processed using a common downstream workflow. This study aimed to develop a high-yielding tangential flow filtration (TFF) process for a pentavalent norovirus vaccine comprising five VP1-encoding mRNA constructs from genotypes GI.1, GII.2, GII.3, GII.4 and GII.6.Methods
TFF studies evaluated membrane MWCOs, transmembrane pressures, shear rates, mRNA loadings, and scale-up. Process performance was assessed based on mRNA recovery, integrity, functionality, and apparent hydrodynamic size.Results
Although the VP1 constructs had similar molecular weights (590-610 kDa), initial studies using 100 kDa membranes revealed construct-dependent losses due to permeation of full-length mRNA. Apparent hydrodynamic size was associated with observed membrane sieving behavior, suggesting that transcript-specific structural properties may influence membrane transmission. Reducing the membrane MWCO from 100 to 50 kDa eliminated detectable mRNA permeation while maintaining membrane performance. A shear rate of 4000 s⁻¹ supported higher mRNA loading without compromising recovery or integrity. The resulting 50 kDa process was successfully scaled from 40 cm² to 1600 cm² and achieved product yields exceeding 95% across all five VP1 mRNA constructs.Conclusions
These findings support a framework for developing standardized TFF conditions for multivalent mRNA vaccines. The results indicate that transcript molecular weight and nominal membrane MWCO alone may not predict mRNA retention and demonstrate the applicability of a common TFF process across five VP1 mRNA constructs. The proposed influence of RNA conformation on membrane transmission requires confirmation using orthogonal structural methods.
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BibTeX
@article{dewar2026platform,
title = {{A Platform Approach to mRNA Purification by Tangential Flow Filtration: Application to a Pentavalent Norovirus Vaccine.}},
author = {Emily A Dewar and Andrew R Swartz and Keira Tuberty and Adam Gouveia},
journal = {Vaccines},
year = {2026},
month = oct,
doi = {10.3390/vaccines14100865},
url = {https://europepmc.org/article/MED/42931394},
note = {CC BY},
}
RIS
TY - JOUR
TI - A Platform Approach to mRNA Purification by Tangential Flow Filtration: Application to a Pentavalent Norovirus Vaccine.
AU - Emily A Dewar
AU - Andrew R Swartz
AU - Keira Tuberty
AU - Adam Gouveia
PY - 2026
DA - 2026/10/01
JO - Vaccines
DO - 10.3390/vaccines14100865
UR - https://europepmc.org/article/MED/42931394
AB - Background/objectives Multivalent mRNA vaccines are being evaluated for prevention of norovirus infection caused by multiple circulating strains. Platform purification processes may accelerate development and reduce manufacturing complexity by enabling multiple mRNA constructs to be processed using a common downstream workflow. This study aimed to develop a high-yielding tangential flow filtration (TFF) process for a pentavalent norovirus vaccine comprising five VP1-encoding mRNA constructs from genotypes GI.1, GII.2, GII.3, GII.4 and GII.6.Methods TFF studies evaluated membrane MWCOs, transmembrane pressures, shear rates, mRNA loadings, and scale-up. Process performance was assessed based on mRNA recovery, integrity, functionality, and apparent hydrodynamic size.Results Although the VP1 constructs had similar molecular weights (590-610 kDa), initial studies using 100 kDa membranes revealed construct-dependent losses due to permeation of full-length mRNA. Apparent hydrodynamic size was associated with observed membrane sieving behavior, suggesting that transcript-specific structural properties may influence membrane transmission. Reducing the membrane MWCO from 100 to 50 kDa eliminated detectable mRNA permeation while maintaining membrane performance. A shear rate of 4000 s⁻¹ supported higher mRNA loading without compromising recovery or integrity. The resulting 50 kDa process was successfully scaled from 40 cm² to 1600 cm² and achieved product yields exceeding 95% across all five VP1 mRNA constructs.Conclusions These findings support a framework for developing standardized TFF conditions for multivalent mRNA vaccines. The results indicate that transcript molecular weight and nominal membrane MWCO alone may not predict mRNA retention and demonstrate the applicability of a common TFF process across five VP1 mRNA constructs. The proposed influence of RNA conformation on membrane transmission requires confirmation using orthogonal structural methods.
SN - 2076-393X
ER -