Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen.
Identifieur interne : 000987 ( PubMed/Corpus ); précédent : 000986; suivant : 000988Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen.
Auteurs : Mun Peak Nyon ; Lanying Du ; Chien-Te Kent Tseng ; Christopher A. Seid ; Jeroen Pollet ; Kevin S. Naceanceno ; Anurodh Agrawal ; Abdullah Algaissi ; Bi-Hung Peng ; Wanbo Tai ; Shibo Jiang ; Maria Elena Bottazzi ; Ulrich Strych ; Peter J. HotezSource :
- Vaccine [ 1873-2518 ] ; 2018.
English descriptors
- KwdEn :
- Animals, Antigens, Viral (genetics), Antigens, Viral (immunology), CHO Cells, Coronavirus Infections (immunology), Coronavirus Infections (prevention & control), Cricetulus, Epitopes (chemistry), Epitopes (immunology), Gene Expression, Genetic Engineering, Genetic Vectors (genetics), Immunogenicity, Vaccine, Immunoglobulin Fc Fragments (genetics), Immunoglobulin Fc Fragments (immunology), Mice, Middle East Respiratory Syndrome Coronavirus (immunology), Protein Processing, Post-Translational, Recombinant Proteins (genetics), Recombinant Proteins (immunology), Viral Vaccines (immunology).
- MESH :
- chemical , chemistry : Epitopes.
- chemical , genetics : Antigens, Viral, Immunoglobulin Fc Fragments, Recombinant Proteins.
- chemical , immunology : Antigens, Viral, Epitopes, Immunoglobulin Fc Fragments, Recombinant Proteins, Viral Vaccines.
- genetics : Genetic Vectors.
- immunology : Coronavirus Infections, Middle East Respiratory Syndrome Coronavirus.
- prevention & control : Coronavirus Infections.
- Animals, CHO Cells, Cricetulus, Gene Expression, Genetic Engineering, Immunogenicity, Vaccine, Mice, Protein Processing, Post-Translational.
Abstract
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.
DOI: 10.1016/j.vaccine.2018.02.065
PubMed: 29496347
Links to Exploration step
pubmed:29496347Le document en format XML
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<author><name sortKey="Du, Lanying" sort="Du, Lanying" uniqKey="Du L" first="Lanying" last="Du">Lanying Du</name>
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<author><name sortKey="Tseng, Chien Te Kent" sort="Tseng, Chien Te Kent" uniqKey="Tseng C" first="Chien-Te Kent" last="Tseng">Chien-Te Kent Tseng</name>
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<author><name sortKey="Peng, Bi Hung" sort="Peng, Bi Hung" uniqKey="Peng B" first="Bi-Hung" last="Peng">Bi-Hung Peng</name>
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<author><name sortKey="Tai, Wanbo" sort="Tai, Wanbo" uniqKey="Tai W" first="Wanbo" last="Tai">Wanbo Tai</name>
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<author><name sortKey="Jiang, Shibo" sort="Jiang, Shibo" uniqKey="Jiang S" first="Shibo" last="Jiang">Shibo Jiang</name>
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</affiliation>
</author>
<author><name sortKey="Bottazzi, Maria Elena" sort="Bottazzi, Maria Elena" uniqKey="Bottazzi M" first="Maria Elena" last="Bottazzi">Maria Elena Bottazzi</name>
<affiliation><nlm:affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA. Electronic address: bottazzi@bcm.edu.</nlm:affiliation>
</affiliation>
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<author><name sortKey="Strych, Ulrich" sort="Strych, Ulrich" uniqKey="Strych U" first="Ulrich" last="Strych">Ulrich Strych</name>
<affiliation><nlm:affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</nlm:affiliation>
</affiliation>
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<author><name sortKey="Hotez, Peter J" sort="Hotez, Peter J" uniqKey="Hotez P" first="Peter J" last="Hotez">Peter J. Hotez</name>
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<author><name sortKey="Nyon, Mun Peak" sort="Nyon, Mun Peak" uniqKey="Nyon M" first="Mun Peak" last="Nyon">Mun Peak Nyon</name>
<affiliation><nlm:affiliation>Tropical Infectious Diseases Research and Education Centre (TIDREC), University of Malaya, Kuala Lumpur, Malaysia; Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</nlm:affiliation>
</affiliation>
</author>
<author><name sortKey="Du, Lanying" sort="Du, Lanying" uniqKey="Du L" first="Lanying" last="Du">Lanying Du</name>
<affiliation><nlm:affiliation>Lindsley F. Kimball Research Institute, New York Blood Center, New York, NY, USA.</nlm:affiliation>
</affiliation>
</author>
<author><name sortKey="Tseng, Chien Te Kent" sort="Tseng, Chien Te Kent" uniqKey="Tseng C" first="Chien-Te Kent" last="Tseng">Chien-Te Kent Tseng</name>
<affiliation><nlm:affiliation>Department of Microbiology and Immunology & Center of Biodefense and Emerging Diseases, University of Texas Medical Branch, Galveston, TX, USA.</nlm:affiliation>
</affiliation>
</author>
<author><name sortKey="Seid, Christopher A" sort="Seid, Christopher A" uniqKey="Seid C" first="Christopher A" last="Seid">Christopher A. Seid</name>
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</affiliation>
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<author><name sortKey="Pollet, Jeroen" sort="Pollet, Jeroen" uniqKey="Pollet J" first="Jeroen" last="Pollet">Jeroen Pollet</name>
<affiliation><nlm:affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</nlm:affiliation>
</affiliation>
</author>
<author><name sortKey="Naceanceno, Kevin S" sort="Naceanceno, Kevin S" uniqKey="Naceanceno K" first="Kevin S" last="Naceanceno">Kevin S. Naceanceno</name>
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</affiliation>
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<author><name sortKey="Agrawal, Anurodh" sort="Agrawal, Anurodh" uniqKey="Agrawal A" first="Anurodh" last="Agrawal">Anurodh Agrawal</name>
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</affiliation>
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<author><name sortKey="Algaissi, Abdullah" sort="Algaissi, Abdullah" uniqKey="Algaissi A" first="Abdullah" last="Algaissi">Abdullah Algaissi</name>
<affiliation><nlm:affiliation>Department of Microbiology and Immunology & Center of Biodefense and Emerging Diseases, University of Texas Medical Branch, Galveston, TX, USA.</nlm:affiliation>
</affiliation>
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<author><name sortKey="Peng, Bi Hung" sort="Peng, Bi Hung" uniqKey="Peng B" first="Bi-Hung" last="Peng">Bi-Hung Peng</name>
<affiliation><nlm:affiliation>Department of Microbiology and Immunology & Center of Biodefense and Emerging Diseases, University of Texas Medical Branch, Galveston, TX, USA.</nlm:affiliation>
</affiliation>
</author>
<author><name sortKey="Tai, Wanbo" sort="Tai, Wanbo" uniqKey="Tai W" first="Wanbo" last="Tai">Wanbo Tai</name>
<affiliation><nlm:affiliation>Lindsley F. Kimball Research Institute, New York Blood Center, New York, NY, USA.</nlm:affiliation>
</affiliation>
</author>
<author><name sortKey="Jiang, Shibo" sort="Jiang, Shibo" uniqKey="Jiang S" first="Shibo" last="Jiang">Shibo Jiang</name>
<affiliation><nlm:affiliation>Lindsley F. Kimball Research Institute, New York Blood Center, New York, NY, USA; Key Laboratory of Medical Molecular Virology of MOE/MOH, School of Basic Medical Sciences, Fudan University, Shanghai, China.</nlm:affiliation>
</affiliation>
</author>
<author><name sortKey="Bottazzi, Maria Elena" sort="Bottazzi, Maria Elena" uniqKey="Bottazzi M" first="Maria Elena" last="Bottazzi">Maria Elena Bottazzi</name>
<affiliation><nlm:affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA. Electronic address: bottazzi@bcm.edu.</nlm:affiliation>
</affiliation>
</author>
<author><name sortKey="Strych, Ulrich" sort="Strych, Ulrich" uniqKey="Strych U" first="Ulrich" last="Strych">Ulrich Strych</name>
<affiliation><nlm:affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</nlm:affiliation>
</affiliation>
</author>
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<affiliation><nlm:affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</nlm:affiliation>
</affiliation>
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<series><title level="j">Vaccine</title>
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<imprint><date when="2018" type="published">2018</date>
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<profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Animals</term>
<term>Antigens, Viral (genetics)</term>
<term>Antigens, Viral (immunology)</term>
<term>CHO Cells</term>
<term>Coronavirus Infections (immunology)</term>
<term>Coronavirus Infections (prevention & control)</term>
<term>Cricetulus</term>
<term>Epitopes (chemistry)</term>
<term>Epitopes (immunology)</term>
<term>Gene Expression</term>
<term>Genetic Engineering</term>
<term>Genetic Vectors (genetics)</term>
<term>Immunogenicity, Vaccine</term>
<term>Immunoglobulin Fc Fragments (genetics)</term>
<term>Immunoglobulin Fc Fragments (immunology)</term>
<term>Mice</term>
<term>Middle East Respiratory Syndrome Coronavirus (immunology)</term>
<term>Protein Processing, Post-Translational</term>
<term>Recombinant Proteins (genetics)</term>
<term>Recombinant Proteins (immunology)</term>
<term>Viral Vaccines (immunology)</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Epitopes</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Antigens, Viral</term>
<term>Immunoglobulin Fc Fragments</term>
<term>Recombinant Proteins</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="immunology" xml:lang="en"><term>Antigens, Viral</term>
<term>Epitopes</term>
<term>Immunoglobulin Fc Fragments</term>
<term>Recombinant Proteins</term>
<term>Viral Vaccines</term>
</keywords>
<keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Genetic Vectors</term>
</keywords>
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<term>Middle East Respiratory Syndrome Coronavirus</term>
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<term>CHO Cells</term>
<term>Cricetulus</term>
<term>Gene Expression</term>
<term>Genetic Engineering</term>
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<front><div type="abstract" xml:lang="en">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.</div>
</front>
</TEI>
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<DateCompleted><Year>2018</Year>
<Month>09</Month>
<Day>11</Day>
</DateCompleted>
<DateRevised><Year>2020</Year>
<Month>04</Month>
<Day>07</Day>
</DateRevised>
<Article PubModel="Print-Electronic"><Journal><ISSN IssnType="Electronic">1873-2518</ISSN>
<JournalIssue CitedMedium="Internet"><Volume>36</Volume>
<Issue>14</Issue>
<PubDate><Year>2018</Year>
<Month>03</Month>
<Day>27</Day>
</PubDate>
</JournalIssue>
<Title>Vaccine</Title>
<ISOAbbreviation>Vaccine</ISOAbbreviation>
</Journal>
<ArticleTitle>Engineering a stable CHO cell line for the expression of a MERS-coronavirus vaccine antigen.</ArticleTitle>
<Pagination><MedlinePgn>1853-1862</MedlinePgn>
</Pagination>
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<ELocationID EIdType="doi" ValidYN="Y">10.1016/j.vaccine.2018.02.065</ELocationID>
<Abstract><AbstractText>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.</AbstractText>
<CopyrightInformation>Copyright © 2018 Elsevier Ltd. All rights reserved.</CopyrightInformation>
</Abstract>
<AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Nyon</LastName>
<ForeName>Mun Peak</ForeName>
<Initials>MP</Initials>
<AffiliationInfo><Affiliation>Tropical Infectious Diseases Research and Education Centre (TIDREC), University of Malaya, Kuala Lumpur, Malaysia; Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Du</LastName>
<ForeName>Lanying</ForeName>
<Initials>L</Initials>
<AffiliationInfo><Affiliation>Lindsley F. Kimball Research Institute, New York Blood Center, New York, NY, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Tseng</LastName>
<ForeName>Chien-Te Kent</ForeName>
<Initials>CK</Initials>
<AffiliationInfo><Affiliation>Department of Microbiology and Immunology & Center of Biodefense and Emerging Diseases, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Seid</LastName>
<ForeName>Christopher A</ForeName>
<Initials>CA</Initials>
<AffiliationInfo><Affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Pollet</LastName>
<ForeName>Jeroen</ForeName>
<Initials>J</Initials>
<AffiliationInfo><Affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Naceanceno</LastName>
<ForeName>Kevin S</ForeName>
<Initials>KS</Initials>
<AffiliationInfo><Affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Agrawal</LastName>
<ForeName>Anurodh</ForeName>
<Initials>A</Initials>
<AffiliationInfo><Affiliation>Department of Microbiology and Immunology & Center of Biodefense and Emerging Diseases, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Algaissi</LastName>
<ForeName>Abdullah</ForeName>
<Initials>A</Initials>
<AffiliationInfo><Affiliation>Department of Microbiology and Immunology & Center of Biodefense and Emerging Diseases, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Peng</LastName>
<ForeName>Bi-Hung</ForeName>
<Initials>BH</Initials>
<AffiliationInfo><Affiliation>Department of Microbiology and Immunology & Center of Biodefense and Emerging Diseases, University of Texas Medical Branch, Galveston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Tai</LastName>
<ForeName>Wanbo</ForeName>
<Initials>W</Initials>
<AffiliationInfo><Affiliation>Lindsley F. Kimball Research Institute, New York Blood Center, New York, NY, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Jiang</LastName>
<ForeName>Shibo</ForeName>
<Initials>S</Initials>
<AffiliationInfo><Affiliation>Lindsley F. Kimball Research Institute, New York Blood Center, New York, NY, USA; Key Laboratory of Medical Molecular Virology of MOE/MOH, School of Basic Medical Sciences, Fudan University, Shanghai, China.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Bottazzi</LastName>
<ForeName>Maria Elena</ForeName>
<Initials>ME</Initials>
<AffiliationInfo><Affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA. Electronic address: bottazzi@bcm.edu.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Strych</LastName>
<ForeName>Ulrich</ForeName>
<Initials>U</Initials>
<AffiliationInfo><Affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
<Author ValidYN="Y"><LastName>Hotez</LastName>
<ForeName>Peter J</ForeName>
<Initials>PJ</Initials>
<AffiliationInfo><Affiliation>Texas Children's Hospital Center for Vaccine Development, USA; Department of Pediatrics, National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.</Affiliation>
</AffiliationInfo>
</Author>
</AuthorList>
<Language>eng</Language>
<GrantList CompleteYN="Y"><Grant><GrantID>R01 AI098775</GrantID>
<Acronym>AI</Acronym>
<Agency>NIAID NIH HHS</Agency>
<Country>United States</Country>
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<Grant><GrantID>R21 AI128311</GrantID>
<Acronym>AI</Acronym>
<Agency>NIAID NIH HHS</Agency>
<Country>United States</Country>
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