Journal
BIOCHEMICAL ENGINEERING JOURNAL
Volume 67, Issue -, Pages 88-96Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bej.2012.05.007
Keywords
Virus-like particles; Self-assembly; Bioprocess design; Downstream processing; Process integration; Scale-up
Funding
- National Institutes of Health (National Center for Research Resources) [2P41RR001081]
- National Institutes of Health (National Institute of General Medical Sciences) [9P41GM103311]
- Universiti Sains Malaysia through the Ministry of Higher Education, Malaysia
- Queensland Government Smart Futures Fund
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Virus-like particles (VLPs) have been developed as safe and efficacious vaccines, and are increasingly used as carriers for foreign peptide epitopes in modular architectures. An emerging technology for low-cost and rapid-response VLP vaccine manufacture is based on controlled cell-free assembly of capsomeres, which are expressed in Escherichia call, into VLPs presenting pharmaceutically relevant antigenic modules. A key bioprocessing challenge in this technology is VLP self-assembly, which has until now been studied using qualitative laboratory methods and without sufficient quantitation. In this work, the yield and size distribution of VLPs assembled by dialysis or by ten-fold dilution were compared using quantitative metrics. Membrane-based steps used for dialysis and particle concentration were identified as the key inefficiencies in each method, resulting in 13-18% protein loss. Key inefficiencies were circumvented through process intensification that led to development of a two-fold dilution assembly method. The new process eliminated a unit operation, improved the final concentration of assembly products by a factor of five and reduced buffer consumption nine-fold. Using this process, modular capsomeres presenting a group A Streptococcus (GAS) antigenic module were assembled into high-quality VLPs with a final yield of 54%, which was 18-22 percentage points higher than obtained using conventional methods described in the literature. This study demonstrates the feasibility of manufacturing VLP vaccines in cell-free reactors at high yield, with high structural integrity, using a scalable and simple process. Ongoing development based on these results will be conducive to process-intensified VLP bioprocessing for low-cost vaccine delivery at global scale. (C) 2012 Elsevier B.V. All rights reserved.
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