Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen
Middle East respiratory syndrome coronavirus (MERS-CoV) has infected at least 2040 patients and caused 712 deaths since its first appearance in 2012, yet neither pathogen-specific therapeutics nor approved vaccines are available. To address this need, we are developing a subunit recombinant protein...
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Elsevier
2018
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| author | Nyon, Mun Peak Du, Lanying Tseng, Chien Te Kent Seid, Christopher A. Pollet, Jeroen Naceanceno, Kevin S. Agrawal, Anurodh Algaissi, Abdullah Peng, Bi Hung Tai, Wanbo Jiang, Shibo Bottazzi, Maria Elena Strych, Ulrich Hotez, Peter J. |
| author_facet | Nyon, Mun Peak Du, Lanying Tseng, Chien Te Kent Seid, Christopher A. Pollet, Jeroen Naceanceno, Kevin S. Agrawal, Anurodh Algaissi, Abdullah Peng, Bi Hung Tai, Wanbo Jiang, Shibo Bottazzi, Maria Elena Strych, Ulrich Hotez, Peter J. |
| author_sort | Nyon, Mun Peak |
| collection | UM |
| description | Middle East respiratory syndrome coronavirus (MERS-CoV) has infected at least 2040 patients and caused 712 deaths since its first appearance in 2012, yet neither pathogen-specific therapeutics nor approved vaccines are available. To address this need, we are developing a subunit recombinant protein vaccine comprising residues 377–588 of the MERS-CoV spike protein receptor-binding domain (RBD), which, when formulated with the AddaVax adjuvant, it induces a significant neutralizing antibody response and protection against MERS-CoV challenge in vaccinated animals. To prepare for the manufacture and first-in-human testing of the vaccine, we have developed a process to stably produce the recombinant MERS S377-588 protein in Chinese hamster ovary (CHO) cells. To accomplish this, we transfected an adherent dihydrofolate reductase-deficient CHO cell line (adCHO) with a plasmid encoding S377-588 fused with the human IgG Fc fragment (S377-588-Fc). We then demonstrated the interleukin-2 signal peptide-directed secretion of the recombinant protein into extracellular milieu. Using a gradually increasing methotrexate (MTX) concentration to 5 μM, we increased protein yield by a factor of 40. The adCHO-expressed S377-588-Fc recombinant protein demonstrated functionality and binding specificity identical to those of the protein from transiently transfected HEK293T cells. In addition, hCD26/dipeptidyl peptidase-4 (DPP4) transgenic mice vaccinated with AddaVax-adjuvanted S377-588-Fc could produce neutralizing antibodies against MERS-CoV and survived for at least 21 days after challenge with live MERS-CoV with no evidence of immunological toxicity or eosinophilic immune enhancement. To prepare for large scale-manufacture of the vaccine antigen, we have further developed a high-yield monoclonal suspension CHO cell line. |
| first_indexed | 2024-03-06T05:53:08Z |
| format | Article |
| id | um.eprints-21091 |
| institution | Universiti Malaya |
| last_indexed | 2024-03-06T05:53:08Z |
| publishDate | 2018 |
| publisher | Elsevier |
| record_format | dspace |
| spelling | um.eprints-210912019-04-26T08:50:15Z http://eprints.um.edu.my/21091/ Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen Nyon, Mun Peak Du, Lanying Tseng, Chien Te Kent Seid, Christopher A. Pollet, Jeroen Naceanceno, Kevin S. Agrawal, Anurodh Algaissi, Abdullah Peng, Bi Hung Tai, Wanbo Jiang, Shibo Bottazzi, Maria Elena Strych, Ulrich Hotez, Peter J. R Medicine Middle East respiratory syndrome coronavirus (MERS-CoV) has infected at least 2040 patients and caused 712 deaths since its first appearance in 2012, yet neither pathogen-specific therapeutics nor approved vaccines are available. To address this need, we are developing a subunit recombinant protein vaccine comprising residues 377–588 of the MERS-CoV spike protein receptor-binding domain (RBD), which, when formulated with the AddaVax adjuvant, it induces a significant neutralizing antibody response and protection against MERS-CoV challenge in vaccinated animals. To prepare for the manufacture and first-in-human testing of the vaccine, we have developed a process to stably produce the recombinant MERS S377-588 protein in Chinese hamster ovary (CHO) cells. To accomplish this, we transfected an adherent dihydrofolate reductase-deficient CHO cell line (adCHO) with a plasmid encoding S377-588 fused with the human IgG Fc fragment (S377-588-Fc). We then demonstrated the interleukin-2 signal peptide-directed secretion of the recombinant protein into extracellular milieu. Using a gradually increasing methotrexate (MTX) concentration to 5 μM, we increased protein yield by a factor of 40. The adCHO-expressed S377-588-Fc recombinant protein demonstrated functionality and binding specificity identical to those of the protein from transiently transfected HEK293T cells. In addition, hCD26/dipeptidyl peptidase-4 (DPP4) transgenic mice vaccinated with AddaVax-adjuvanted S377-588-Fc could produce neutralizing antibodies against MERS-CoV and survived for at least 21 days after challenge with live MERS-CoV with no evidence of immunological toxicity or eosinophilic immune enhancement. To prepare for large scale-manufacture of the vaccine antigen, we have further developed a high-yield monoclonal suspension CHO cell line. Elsevier 2018 Article PeerReviewed Nyon, Mun Peak and Du, Lanying and Tseng, Chien Te Kent and Seid, Christopher A. and Pollet, Jeroen and Naceanceno, Kevin S. and Agrawal, Anurodh and Algaissi, Abdullah and Peng, Bi Hung and Tai, Wanbo and Jiang, Shibo and Bottazzi, Maria Elena and Strych, Ulrich and Hotez, Peter J. (2018) Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen. Vaccine, 36 (14). pp. 1853-1862. ISSN 0264-410X, DOI https://doi.org/10.1016/j.vaccine.2018.02.065 <https://doi.org/10.1016/j.vaccine.2018.02.065>. https://doi.org/10.1016/j.vaccine.2018.02.065 doi:10.1016/j.vaccine.2018.02.065 |
| spellingShingle | R Medicine Nyon, Mun Peak Du, Lanying Tseng, Chien Te Kent Seid, Christopher A. Pollet, Jeroen Naceanceno, Kevin S. Agrawal, Anurodh Algaissi, Abdullah Peng, Bi Hung Tai, Wanbo Jiang, Shibo Bottazzi, Maria Elena Strych, Ulrich Hotez, Peter J. Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen |
| title | Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen |
| title_full | Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen |
| title_fullStr | Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen |
| title_full_unstemmed | Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen |
| title_short | Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen |
| title_sort | engineering a stable cho cell line for the expression of a mers coronavirus vaccine antigen |
| topic | R Medicine |
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