Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigens
We have developed a single process for producing two key COVID-19 vaccine antigens: SARS-CoV-2 receptor binding domain (RBD) monomer and dimer. These antigens are featured in various COVID-19 vaccine formats, including SOBERANA 01 and the licensed SOBERANA 02, and SOBERANA Plus. Our approach involve...
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Format: | Article |
Language: | English |
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Frontiers Media S.A.
2023-11-01
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Series: | Frontiers in Bioengineering and Biotechnology |
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Online Access: | https://www.frontiersin.org/articles/10.3389/fbioe.2023.1287551/full |
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author | Tammy Boggiano-Ayo Julio Palacios-Oliva Sumlai Lozada-Chang Ernesto Relova-Hernandez Jose Gomez-Perez Gonzalo Oliva Lourdes Hernandez Alexi Bueno-Soler Daidee Montes de Oca Osvaldo Mora Roberto Machado-Santisteban Dayana Perez-Martinez Beatriz Perez-Masson Yanelys Cabrera Infante Lisandra Calzadilla-Rosado Yaima Ramirez Judey Aymed-Garcia Ingrid Ruiz-Ramirez Yamile Romero Tania Gomez Luis A. Espinosa Luis Javier Gonzalez Annia Cabrales Osmany Guirola Kathya Rashida de la Luz Franciscary Pi-Estopiñan Belinda Sanchez-Ramirez Dagmar Garcia-Rivera Yuri Valdes-Balbin Gertrudis Rojas Kalet Leon-Monzon Eduardo Ojito-Magaz Eugenio Hardy |
author_facet | Tammy Boggiano-Ayo Julio Palacios-Oliva Sumlai Lozada-Chang Ernesto Relova-Hernandez Jose Gomez-Perez Gonzalo Oliva Lourdes Hernandez Alexi Bueno-Soler Daidee Montes de Oca Osvaldo Mora Roberto Machado-Santisteban Dayana Perez-Martinez Beatriz Perez-Masson Yanelys Cabrera Infante Lisandra Calzadilla-Rosado Yaima Ramirez Judey Aymed-Garcia Ingrid Ruiz-Ramirez Yamile Romero Tania Gomez Luis A. Espinosa Luis Javier Gonzalez Annia Cabrales Osmany Guirola Kathya Rashida de la Luz Franciscary Pi-Estopiñan Belinda Sanchez-Ramirez Dagmar Garcia-Rivera Yuri Valdes-Balbin Gertrudis Rojas Kalet Leon-Monzon Eduardo Ojito-Magaz Eugenio Hardy |
author_sort | Tammy Boggiano-Ayo |
collection | DOAJ |
description | We have developed a single process for producing two key COVID-19 vaccine antigens: SARS-CoV-2 receptor binding domain (RBD) monomer and dimer. These antigens are featured in various COVID-19 vaccine formats, including SOBERANA 01 and the licensed SOBERANA 02, and SOBERANA Plus. Our approach involves expressing RBD (319-541)-His6 in Chinese hamster ovary (CHO)-K1 cells, generating and characterizing oligoclones, and selecting the best RBD-producing clones. Critical parameters such as copper supplementation in the culture medium and cell viability influenced the yield of RBD dimer. The purification of RBD involved standard immobilized metal ion affinity chromatography (IMAC), ion exchange chromatography, and size exclusion chromatography. Our findings suggest that copper can improve IMAC performance. Efficient RBD production was achieved using small-scale bioreactor cell culture (2 L). The two RBD forms - monomeric and dimeric RBD - were also produced on a large scale (500 L). This study represents the first large-scale application of perfusion culture for the production of RBD antigens. We conducted a thorough analysis of the purified RBD antigens, which encompassed primary structure, protein integrity, N-glycosylation, size, purity, secondary and tertiary structures, isoform composition, hydrophobicity, and long-term stability. Additionally, we investigated RBD-ACE2 interactions, in vitro ACE2 recognition of RBD, and the immunogenicity of RBD antigens in mice. We have determined that both the monomeric and dimeric RBD antigens possess the necessary quality attributes for vaccine production. By enabling the customizable production of both RBD forms, this unified manufacturing process provides the required flexibility to adapt rapidly to the ever-changing demands of emerging SARS-CoV-2 variants and different COVID-19 vaccine platforms. |
first_indexed | 2024-03-11T02:30:10Z |
format | Article |
id | doaj.art-e0706f0426fc462d821685ef6ce26f54 |
institution | Directory Open Access Journal |
issn | 2296-4185 |
language | English |
last_indexed | 2024-03-11T02:30:10Z |
publishDate | 2023-11-01 |
publisher | Frontiers Media S.A. |
record_format | Article |
series | Frontiers in Bioengineering and Biotechnology |
spelling | doaj.art-e0706f0426fc462d821685ef6ce26f542023-11-18T10:20:33ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852023-11-011110.3389/fbioe.2023.12875511287551Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigensTammy Boggiano-Ayo0Julio Palacios-Oliva1Sumlai Lozada-Chang2Ernesto Relova-Hernandez3Jose Gomez-Perez4Gonzalo Oliva5Lourdes Hernandez6Alexi Bueno-Soler7Daidee Montes de Oca8Osvaldo Mora9Roberto Machado-Santisteban10Dayana Perez-Martinez11Beatriz Perez-Masson12Yanelys Cabrera Infante13Lisandra Calzadilla-Rosado14Yaima Ramirez15Judey Aymed-Garcia16Ingrid Ruiz-Ramirez17Yamile Romero18Tania Gomez19Luis A. Espinosa20Luis Javier Gonzalez21Annia Cabrales22Osmany Guirola23Kathya Rashida de la Luz24Franciscary Pi-Estopiñan25Belinda Sanchez-Ramirez26Dagmar Garcia-Rivera27Yuri Valdes-Balbin28Gertrudis Rojas29Kalet Leon-Monzon30Eduardo Ojito-Magaz31Eugenio Hardy32Process Development Direction, Center of Molecular Immunology, Havana, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaQuality Direction, Center of Molecular Immunology, Havana, CubaProcess Direction, Center of Molecular Immunology, Havana, CubaProcess Direction, Center of Molecular Immunology, Havana, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaProcess Direction, Center of Molecular Immunology, Havana, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaQuality Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaQuality Direction, Center of Molecular Immunology, Havana, CubaCenter for Genetic Engineering and Biotechnology, Playa, CubaCenter for Genetic Engineering and Biotechnology, Playa, CubaCenter for Genetic Engineering and Biotechnology, Playa, CubaCenter for Genetic Engineering and Biotechnology, Playa, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaFinlay Vaccine Institute, Havana, CubaFinlay Vaccine Institute, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaImmunology and Immunobiology Direction, Center of Molecular Immunology, Havana, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaProcess Development Direction, Center of Molecular Immunology, Havana, CubaWe have developed a single process for producing two key COVID-19 vaccine antigens: SARS-CoV-2 receptor binding domain (RBD) monomer and dimer. These antigens are featured in various COVID-19 vaccine formats, including SOBERANA 01 and the licensed SOBERANA 02, and SOBERANA Plus. Our approach involves expressing RBD (319-541)-His6 in Chinese hamster ovary (CHO)-K1 cells, generating and characterizing oligoclones, and selecting the best RBD-producing clones. Critical parameters such as copper supplementation in the culture medium and cell viability influenced the yield of RBD dimer. The purification of RBD involved standard immobilized metal ion affinity chromatography (IMAC), ion exchange chromatography, and size exclusion chromatography. Our findings suggest that copper can improve IMAC performance. Efficient RBD production was achieved using small-scale bioreactor cell culture (2 L). The two RBD forms - monomeric and dimeric RBD - were also produced on a large scale (500 L). This study represents the first large-scale application of perfusion culture for the production of RBD antigens. We conducted a thorough analysis of the purified RBD antigens, which encompassed primary structure, protein integrity, N-glycosylation, size, purity, secondary and tertiary structures, isoform composition, hydrophobicity, and long-term stability. Additionally, we investigated RBD-ACE2 interactions, in vitro ACE2 recognition of RBD, and the immunogenicity of RBD antigens in mice. We have determined that both the monomeric and dimeric RBD antigens possess the necessary quality attributes for vaccine production. By enabling the customizable production of both RBD forms, this unified manufacturing process provides the required flexibility to adapt rapidly to the ever-changing demands of emerging SARS-CoV-2 variants and different COVID-19 vaccine platforms.https://www.frontiersin.org/articles/10.3389/fbioe.2023.1287551/fullSARS-CoV-2RBDCHO cellsperfusion cultureCOVID-19vaccine |
spellingShingle | Tammy Boggiano-Ayo Julio Palacios-Oliva Sumlai Lozada-Chang Ernesto Relova-Hernandez Jose Gomez-Perez Gonzalo Oliva Lourdes Hernandez Alexi Bueno-Soler Daidee Montes de Oca Osvaldo Mora Roberto Machado-Santisteban Dayana Perez-Martinez Beatriz Perez-Masson Yanelys Cabrera Infante Lisandra Calzadilla-Rosado Yaima Ramirez Judey Aymed-Garcia Ingrid Ruiz-Ramirez Yamile Romero Tania Gomez Luis A. Espinosa Luis Javier Gonzalez Annia Cabrales Osmany Guirola Kathya Rashida de la Luz Franciscary Pi-Estopiñan Belinda Sanchez-Ramirez Dagmar Garcia-Rivera Yuri Valdes-Balbin Gertrudis Rojas Kalet Leon-Monzon Eduardo Ojito-Magaz Eugenio Hardy Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigens Frontiers in Bioengineering and Biotechnology SARS-CoV-2 RBD CHO cells perfusion culture COVID-19 vaccine |
title | Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigens |
title_full | Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigens |
title_fullStr | Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigens |
title_full_unstemmed | Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigens |
title_short | Development of a scalable single process for producing SARS-CoV-2 RBD monomer and dimer vaccine antigens |
title_sort | development of a scalable single process for producing sars cov 2 rbd monomer and dimer vaccine antigens |
topic | SARS-CoV-2 RBD CHO cells perfusion culture COVID-19 vaccine |
url | https://www.frontiersin.org/articles/10.3389/fbioe.2023.1287551/full |
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