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Optimizing the Production of gp145, an HIV-1 Envelope Glycoprotein Vaccine Candidate and Its Encapsulation in Guanosine Microparticles
We have developed a pipeline to express, purify, and characterize HIV envelope protein (Env) gp145 from Chinese hamster ovary cells, to accelerate the production of a promising vaccine candidate. First in shake flasks, then in bioreactors, we optimized the growth conditions. By adjusting the pH to 6...
Autores principales: | , , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10221277/ https://www.ncbi.nlm.nih.gov/pubmed/37243079 http://dx.doi.org/10.3390/vaccines11050975 |
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author | Akamine, Pearl González-Feliciano, José A. Almodóvar, Ruth Morell, Gloriner Rivera, Javier Capó-Vélez, Coral M. Delgado-Vélez, Manuel Prieto-Costas, Luis Madera, Bismark Eichinger, Daniel Pino, Ignacio Rivera, José H. Ortiz-Ubarri, José Rivera, José M. Baerga-Ortiz, Abel Lasalde-Dominicci, José A. |
author_facet | Akamine, Pearl González-Feliciano, José A. Almodóvar, Ruth Morell, Gloriner Rivera, Javier Capó-Vélez, Coral M. Delgado-Vélez, Manuel Prieto-Costas, Luis Madera, Bismark Eichinger, Daniel Pino, Ignacio Rivera, José H. Ortiz-Ubarri, José Rivera, José M. Baerga-Ortiz, Abel Lasalde-Dominicci, José A. |
author_sort | Akamine, Pearl |
collection | PubMed |
description | We have developed a pipeline to express, purify, and characterize HIV envelope protein (Env) gp145 from Chinese hamster ovary cells, to accelerate the production of a promising vaccine candidate. First in shake flasks, then in bioreactors, we optimized the growth conditions. By adjusting the pH to 6.8, we increased expression levels to 101 mg/L in a 50 L bioreactor, nearly twice the previously reported titer value. A battery of analytical methods was developed in accordance with current good manufacturing practices to ensure a quality biopharmaceutical. Imaged capillary isoelectric focusing verified proper glycosylation of gp145; dynamic light scattering confirmed the trimeric arrangement; and bio-layer interferometry and circular dichroism analysis demonstrated native-like properties (i.e., antibody binding and secondary structure). MALDI-TOF mass spectrometry was used as a multi-attribute platform for accurate mass determination, glycans analysis, and protein identification. Our robust analysis demonstrates that our gp145 product is very similar to a reference standard and emphasizes the importance of accurate characterization of a highly heterogeneous immunogen for the development of an effective vaccine. Finally, we present a novel guanosine microparticle with gp145 encapsulated and displayed on its surface. The unique properties of our gp145 microparticle make it amenable to use in future preclinical and clinical trials. |
format | Online Article Text |
id | pubmed-10221277 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-102212772023-05-28 Optimizing the Production of gp145, an HIV-1 Envelope Glycoprotein Vaccine Candidate and Its Encapsulation in Guanosine Microparticles Akamine, Pearl González-Feliciano, José A. Almodóvar, Ruth Morell, Gloriner Rivera, Javier Capó-Vélez, Coral M. Delgado-Vélez, Manuel Prieto-Costas, Luis Madera, Bismark Eichinger, Daniel Pino, Ignacio Rivera, José H. Ortiz-Ubarri, José Rivera, José M. Baerga-Ortiz, Abel Lasalde-Dominicci, José A. Vaccines (Basel) Article We have developed a pipeline to express, purify, and characterize HIV envelope protein (Env) gp145 from Chinese hamster ovary cells, to accelerate the production of a promising vaccine candidate. First in shake flasks, then in bioreactors, we optimized the growth conditions. By adjusting the pH to 6.8, we increased expression levels to 101 mg/L in a 50 L bioreactor, nearly twice the previously reported titer value. A battery of analytical methods was developed in accordance with current good manufacturing practices to ensure a quality biopharmaceutical. Imaged capillary isoelectric focusing verified proper glycosylation of gp145; dynamic light scattering confirmed the trimeric arrangement; and bio-layer interferometry and circular dichroism analysis demonstrated native-like properties (i.e., antibody binding and secondary structure). MALDI-TOF mass spectrometry was used as a multi-attribute platform for accurate mass determination, glycans analysis, and protein identification. Our robust analysis demonstrates that our gp145 product is very similar to a reference standard and emphasizes the importance of accurate characterization of a highly heterogeneous immunogen for the development of an effective vaccine. Finally, we present a novel guanosine microparticle with gp145 encapsulated and displayed on its surface. The unique properties of our gp145 microparticle make it amenable to use in future preclinical and clinical trials. MDPI 2023-05-12 /pmc/articles/PMC10221277/ /pubmed/37243079 http://dx.doi.org/10.3390/vaccines11050975 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Akamine, Pearl González-Feliciano, José A. Almodóvar, Ruth Morell, Gloriner Rivera, Javier Capó-Vélez, Coral M. Delgado-Vélez, Manuel Prieto-Costas, Luis Madera, Bismark Eichinger, Daniel Pino, Ignacio Rivera, José H. Ortiz-Ubarri, José Rivera, José M. Baerga-Ortiz, Abel Lasalde-Dominicci, José A. Optimizing the Production of gp145, an HIV-1 Envelope Glycoprotein Vaccine Candidate and Its Encapsulation in Guanosine Microparticles |
title | Optimizing the Production of gp145, an HIV-1 Envelope Glycoprotein Vaccine Candidate and Its Encapsulation in Guanosine Microparticles |
title_full | Optimizing the Production of gp145, an HIV-1 Envelope Glycoprotein Vaccine Candidate and Its Encapsulation in Guanosine Microparticles |
title_fullStr | Optimizing the Production of gp145, an HIV-1 Envelope Glycoprotein Vaccine Candidate and Its Encapsulation in Guanosine Microparticles |
title_full_unstemmed | Optimizing the Production of gp145, an HIV-1 Envelope Glycoprotein Vaccine Candidate and Its Encapsulation in Guanosine Microparticles |
title_short | Optimizing the Production of gp145, an HIV-1 Envelope Glycoprotein Vaccine Candidate and Its Encapsulation in Guanosine Microparticles |
title_sort | optimizing the production of gp145, an hiv-1 envelope glycoprotein vaccine candidate and its encapsulation in guanosine microparticles |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10221277/ https://www.ncbi.nlm.nih.gov/pubmed/37243079 http://dx.doi.org/10.3390/vaccines11050975 |
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