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Holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in Pichia pastoris by means of an acid pH pulse during fed‐batch fermentation
To meet the challenges of global health, vaccine design and development must be reconsidered to achieve cost of goods as low as 15¢ per dose. A new recombinant protein‐based rotavirus vaccine candidate derived from non‐replicative viral subunits fused to a P2 tetanus toxoid CD4(+) T cell epitope is...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
John Wiley & Sons, Inc.
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7317458/ https://www.ncbi.nlm.nih.gov/pubmed/31960616 http://dx.doi.org/10.1002/btpr.2966 |
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author | Velez‐Suberbie, M. Lourdes Morris, Stephen A. Kaur, Kawaljit Hickey, John M. Joshi, Sangeeta B. Volkin, David B. Bracewell, Daniel G. Mukhopadhyay, Tarit K. |
author_facet | Velez‐Suberbie, M. Lourdes Morris, Stephen A. Kaur, Kawaljit Hickey, John M. Joshi, Sangeeta B. Volkin, David B. Bracewell, Daniel G. Mukhopadhyay, Tarit K. |
author_sort | Velez‐Suberbie, M. Lourdes |
collection | PubMed |
description | To meet the challenges of global health, vaccine design and development must be reconsidered to achieve cost of goods as low as 15¢ per dose. A new recombinant protein‐based rotavirus vaccine candidate derived from non‐replicative viral subunits fused to a P2 tetanus toxoid CD4(+) T cell epitope is currently under clinical development. We have sought to simplify the existing manufacturing process to meet these aims. To this end, we have taken a holistic process development approach to reduce process complexity and costs while producing a product with the required characteristics. We have changed expression system from Escherichia coli to Pichia pastoris, to produce a secreted product, thereby reducing the number of purification steps. However, the presence of proteases poses challenges to product quality. To understand the effect of fermentation parameters on product quality small‐scale fermentations were carried out. Media pH and fermentation duration had the greatest impact on the proportion of full‐length product. A novel acidic pH pulse strategy was used to minimize proteolysis, and this combined with an early harvest time significantly increased the proportion of full‐length material (60–75%). An improved downstream process using a combination of CIEX and AIEX to further reduce proteases, resulted in maintaining product quality (95% yield). |
format | Online Article Text |
id | pubmed-7317458 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | John Wiley & Sons, Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-73174582020-06-30 Holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in Pichia pastoris by means of an acid pH pulse during fed‐batch fermentation Velez‐Suberbie, M. Lourdes Morris, Stephen A. Kaur, Kawaljit Hickey, John M. Joshi, Sangeeta B. Volkin, David B. Bracewell, Daniel G. Mukhopadhyay, Tarit K. Biotechnol Prog RESEARCH ARTICLES To meet the challenges of global health, vaccine design and development must be reconsidered to achieve cost of goods as low as 15¢ per dose. A new recombinant protein‐based rotavirus vaccine candidate derived from non‐replicative viral subunits fused to a P2 tetanus toxoid CD4(+) T cell epitope is currently under clinical development. We have sought to simplify the existing manufacturing process to meet these aims. To this end, we have taken a holistic process development approach to reduce process complexity and costs while producing a product with the required characteristics. We have changed expression system from Escherichia coli to Pichia pastoris, to produce a secreted product, thereby reducing the number of purification steps. However, the presence of proteases poses challenges to product quality. To understand the effect of fermentation parameters on product quality small‐scale fermentations were carried out. Media pH and fermentation duration had the greatest impact on the proportion of full‐length product. A novel acidic pH pulse strategy was used to minimize proteolysis, and this combined with an early harvest time significantly increased the proportion of full‐length material (60–75%). An improved downstream process using a combination of CIEX and AIEX to further reduce proteases, resulted in maintaining product quality (95% yield). John Wiley & Sons, Inc. 2020-02-03 2020 /pmc/articles/PMC7317458/ /pubmed/31960616 http://dx.doi.org/10.1002/btpr.2966 Text en © 2020 The Authors. Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers. This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | RESEARCH ARTICLES Velez‐Suberbie, M. Lourdes Morris, Stephen A. Kaur, Kawaljit Hickey, John M. Joshi, Sangeeta B. Volkin, David B. Bracewell, Daniel G. Mukhopadhyay, Tarit K. Holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in Pichia pastoris by means of an acid pH pulse during fed‐batch fermentation |
title | Holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in Pichia pastoris by means of an acid pH pulse during fed‐batch fermentation |
title_full | Holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in Pichia pastoris by means of an acid pH pulse during fed‐batch fermentation |
title_fullStr | Holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in Pichia pastoris by means of an acid pH pulse during fed‐batch fermentation |
title_full_unstemmed | Holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in Pichia pastoris by means of an acid pH pulse during fed‐batch fermentation |
title_short | Holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in Pichia pastoris by means of an acid pH pulse during fed‐batch fermentation |
title_sort | holistic process development to mitigate proteolysis of a subunit rotavirus vaccine candidate produced in pichia pastoris by means of an acid ph pulse during fed‐batch fermentation |
topic | RESEARCH ARTICLES |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7317458/ https://www.ncbi.nlm.nih.gov/pubmed/31960616 http://dx.doi.org/10.1002/btpr.2966 |
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