Engineered thermoswitch in the baculovirus expression vector system for production of virus-like particle vaccines with minimized baculovirus contaminants
The baculovirus expression vector system (BEVS) is commercially used for producing recombinant (glyco)proteins, gene therapy vectors, and virus-like particles (VLPs) in insect cells. A major challenge in VLP downstream processing (DSP) is the removal of similarly sized baculovirus particle contaminants. Herein we engineered a genetic ON/OFF thermoswitch in the baculovirus vector by mutating the essential gp41 gene. This temperature-sensitive (ts) baculovirus expression system (BacFree<sup>ts</sup>) produces high baculovirus titers at 27°C, while contaminating baculovirus production switches off at elevated temperatures. We demonstrate scalable VLP production for chikungunya virus (CHIKV), West Nile virus (WNV), coxsackievirus A6, and foot-and-mouth disease virus (FMDV) at 33°C, concomitant with baculovirus particle reduction up to 99.97%. However, elevated temperature moderately affected the yield of two out of four VLP designs. BacFree<sup>ts</sup> is compatible with commonly used insect cells and scalable infection strategies. BacFree<sup>ts</sup> effectively reduces baculovirus contaminants in the upstream VLP production phase, which will facilitate DSP of human and veterinary VLP-based vaccines.
Authors
- Louis Kuijpers (ORCID: https://orcid.org/0000-0001-7245-3031)
- René H. Wijffels (ORCID: https://orcid.org/0000-0001-7630-4295)
- Gwen Nowee
- Monique M. van Oers (ORCID: https://orcid.org/0000-0002-6664-4445)
- Ricardo Correia (ORCID: https://orcid.org/0000-0002-9434-7974)
- Vivian I. P. Tegelbeckers (ORCID: https://orcid.org/0000-0002-9669-9439)
- Gorben P. Pijlman (ORCID: https://orcid.org/0000-0001-9301-0408)
- Erwin van den Born (ORCID: https://orcid.org/0000-0002-0341-2874)
- Saskia Mentink (ORCID: https://orcid.org/0000-0002-4905-7257)
- Jort J. Altenburg (ORCID: https://orcid.org/0000-0002-9292-5339)
- António Roldão (ORCID: https://orcid.org/0000-0003-0414-4899)
- Christina Sophie Hausjell
- Linda van Oosten (ORCID: https://orcid.org/0000-0003-4346-7156)
- Dirk E. Martens (ORCID: https://orcid.org/0000-0002-5662-0466)
- Joris van der Heijden (ORCID: https://orcid.org/0000-0002-1730-3521)
- Leo van der Pol
- Wessel Willemsen
- Dennis Kenbeek
- Tessa Neef
- Thomas Rouw
Institutions
- Intravacc (Netherlands) (NL)
- Instituto de Biologia Experimental e Tecnológica (PT)
- MSD (Netherlands) (NL)
- Universidade Nova de Lisboa (PT)
- Wageningen University & Research (NL)
- BOKU University (AT)
Publication Details
- Journal
- Trends in biotechnology
- Published
- 2025-05-09
- DOI
- https://doi.org/10.1016/j.tibtech.2025.04.004
- Citations
- 3
- Primary Topic
- Viral Infectious Diseases and Gene Expression in Insects
- Type
- article
- Field-Weighted Citation Impact
- 2.42
Funders
- Chinese Academy of Sciences
- Nederlandse Organisatie voor Wetenschappelijk Onderzoek
- Wageningen University and Research