An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures

Virus-like particles (VLPs) have gained interest over the last years as recombinant vaccine formats, as they generate a strong immune response and present storage and distribution advantages compared to conventional vaccines. Therefore, VLPs are being regarded as potential vaccine candidates for sev...

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Autores: Lavado-García, Jesús|||0000-0001-9993-6332, Cervera Gracia, Laura|||0000-0002-3639-2793, Gòdia, Francesc|||0000-0002-4060-9887
Tipo de recurso: artículo
Fecha de publicación:2020
País:España
Institución:Universitat Autònoma de Barcelona
Repositorio:Dipòsit Digital de Documents de la UAB
Idioma:inglés
OAI Identifier:oai:ddd.uab.cat:227845
Acceso en línea:https://ddd.uab.cat/record/227845
https://dx.doi.org/urn:doi:10.3389/fbioe.2020.00617
Access Level:acceso abierto
Palabra clave:Bioreactor
Perfusion
ATF
Design of experiments
VLP
HFM
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spelling An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 CulturesLavado-García, Jesús|||0000-0001-9993-6332Cervera Gracia, Laura|||0000-0002-3639-2793Gòdia, Francesc|||0000-0002-4060-9887BioreactorPerfusionATFDesign of experimentsVLPHFMVirus-like particles (VLPs) have gained interest over the last years as recombinant vaccine formats, as they generate a strong immune response and present storage and distribution advantages compared to conventional vaccines. Therefore, VLPs are being regarded as potential vaccine candidates for several diseases. One requirement for their further clinical testing is the development of scalable processes and production platforms for cell-based viral particles. In this work, the extended gene expression (EGE) method, which consists in consecutive media replacements combined with cell retransfections, was successfully optimized and transferred to a bioreactor operating in perfusion. A process optimization using design of experiments (DoE) was carried out to obtain optimal values for the time of retransfection, the cell specific perfusion rate (CSPR) and transfected DNA concentration, improving 86.7% the previously reported EGE protocol in HEK293. Moreover, it was successfully implemented at 1.5L bioreactor using an ATF as cell retention system achieving concentrations of 6.8·10 10 VLP/mL. VLP interaction with the ATF hollow fibers was studied via confocal microscopy, field emission scanning electron microscopy, and nanoparticle tracking analysis to design a bioprocess capable of separating unassembled Gag monomers and concentrate VLPs in one step. 22020-01-0120202020-01-01Articlehttp://purl.org/coar/resource_type/c_6501VoRhttp://purl.org/coar/version/c_970fb48d4fbd8a85info:eu-repo/semantics/articleapplication/pdfhttps://ddd.uab.cat/record/227845https://dx.doi.org/urn:doi:10.3389/fbioe.2020.00617reponame:Dipòsit Digital de Documents de la UABinstname:Universitat Autònoma de BarcelonaInglésengAgència de Gestió d'Ajuts Universitaris i de Recerca https://doi.org/10.13039/501100003030 2017/SGR-898open accesshttp://purl.org/coar/access_right/c_abf2Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.https://creativecommons.org/licenses/by/4.0/info:eu-repo/semantics/openAccessoai:ddd.uab.cat:2278452026-06-06T12:50:31Z
dc.title.none.fl_str_mv An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures
title An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures
spellingShingle An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures
Lavado-García, Jesús|||0000-0001-9993-6332
Bioreactor
Perfusion
ATF
Design of experiments
VLP
HFM
title_short An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures
title_full An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures
title_fullStr An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures
title_full_unstemmed An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures
title_sort An Alternative Perfusion Approach for the Intensification of Virus-Like Particle Production in HEK293 Cultures
dc.creator.none.fl_str_mv Lavado-García, Jesús|||0000-0001-9993-6332
Cervera Gracia, Laura|||0000-0002-3639-2793
Gòdia, Francesc|||0000-0002-4060-9887
author Lavado-García, Jesús|||0000-0001-9993-6332
author_facet Lavado-García, Jesús|||0000-0001-9993-6332
Cervera Gracia, Laura|||0000-0002-3639-2793
Gòdia, Francesc|||0000-0002-4060-9887
author_role author
author2 Cervera Gracia, Laura|||0000-0002-3639-2793
Gòdia, Francesc|||0000-0002-4060-9887
author2_role author
author
dc.subject.none.fl_str_mv Bioreactor
Perfusion
ATF
Design of experiments
VLP
HFM
topic Bioreactor
Perfusion
ATF
Design of experiments
VLP
HFM
description Virus-like particles (VLPs) have gained interest over the last years as recombinant vaccine formats, as they generate a strong immune response and present storage and distribution advantages compared to conventional vaccines. Therefore, VLPs are being regarded as potential vaccine candidates for several diseases. One requirement for their further clinical testing is the development of scalable processes and production platforms for cell-based viral particles. In this work, the extended gene expression (EGE) method, which consists in consecutive media replacements combined with cell retransfections, was successfully optimized and transferred to a bioreactor operating in perfusion. A process optimization using design of experiments (DoE) was carried out to obtain optimal values for the time of retransfection, the cell specific perfusion rate (CSPR) and transfected DNA concentration, improving 86.7% the previously reported EGE protocol in HEK293. Moreover, it was successfully implemented at 1.5L bioreactor using an ATF as cell retention system achieving concentrations of 6.8·10 10 VLP/mL. VLP interaction with the ATF hollow fibers was studied via confocal microscopy, field emission scanning electron microscopy, and nanoparticle tracking analysis to design a bioprocess capable of separating unassembled Gag monomers and concentrate VLPs in one step.
publishDate 2020
dc.date.none.fl_str_mv 2
2020-01-01
2020
2020-01-01
dc.type.none.fl_str_mv Article
http://purl.org/coar/resource_type/c_6501
VoR
http://purl.org/coar/version/c_970fb48d4fbd8a85
dc.type.openaire.fl_str_mv info:eu-repo/semantics/article
format article
dc.identifier.none.fl_str_mv https://ddd.uab.cat/record/227845
https://dx.doi.org/urn:doi:10.3389/fbioe.2020.00617
url https://ddd.uab.cat/record/227845
https://dx.doi.org/urn:doi:10.3389/fbioe.2020.00617
dc.language.none.fl_str_mv Inglés
eng
language_invalid_str_mv Inglés
language eng
dc.relation.none.fl_str_mv Agència de Gestió d'Ajuts Universitaris i de Recerca https://doi.org/10.13039/501100003030 2017/SGR-898
dc.rights.none.fl_str_mv open access
http://purl.org/coar/access_right/c_abf2
https://creativecommons.org/licenses/by/4.0/
dc.rights.openaire.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv open access
http://purl.org/coar/access_right/c_abf2
https://creativecommons.org/licenses/by/4.0/
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.source.none.fl_str_mv reponame:Dipòsit Digital de Documents de la UAB
instname:Universitat Autònoma de Barcelona
instname_str Universitat Autònoma de Barcelona
reponame_str Dipòsit Digital de Documents de la UAB
collection Dipòsit Digital de Documents de la UAB
repository.name.fl_str_mv
repository.mail.fl_str_mv
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