Characterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library

A panel of potent neutralizing antibodies are protective against orthopoxvirus (OPXV) infections. For the development of OPXV-specific recombinant human single-chain antibodies (scFvs), the IgG repertoire of four vaccinated donors was amplified from peripheral B-lymphocytes. The resulting library co...

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Autores principales:	Ulrike S. Diesterbeck, Henrike P. Ahsendorf, André Frenzel, Ahmad Reza Sharifi, Thomas Schirrmann, Claus-Peter Czerny
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	scFv vaccinia virus recombinant antibody D8 Medicine R
Acceso en línea:	https://doaj.org/article/46ac0221921e4932ad936734ce702782
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spelling	oai:doaj.org-article:46ac0221921e4932ad936734ce7027822021-11-25T19:11:12ZCharacterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library10.3390/vaccines91113082076-393Xhttps://doaj.org/article/46ac0221921e4932ad936734ce7027822021-11-01T00:00:00Zhttps://www.mdpi.com/2076-393X/9/11/1308https://doaj.org/toc/2076-393XA panel of potent neutralizing antibodies are protective against orthopoxvirus (OPXV) infections. For the development of OPXV-specific recombinant human single-chain antibodies (scFvs), the IgG repertoire of four vaccinated donors was amplified from peripheral B-lymphocytes. The resulting library consisted of ≥4 × 10<sup>8</sup> independent colonies. The immuno-screening against vaccinia virus (VACV) Elstree revealed a predominant selection of scFv clones specifically binding to the D8 protein. The scFv-1.2.2.H9 was engineered into larger human scFv-Fc-1.2.2.H9 and IgG1-1.2.2.H9 formats to improve the binding affinity and to add effector functions within the human immune response. Similar binding kinetics were calculated for scFv-1.2.2.H9 and scFv-Fc-1.2.2.H9 (1.61 nM and 7.685 nM, respectively), whereas, for IgG1-1.2.2.H9, the Michaelis-Menten kinetics revealed an increased affinity of 43.8 pM. None of the purified recombinant 1.2.2.H9 formats were able to neutralize VACV Elstree in vitro. After addition of 1% human complement, the neutralization of ≥50% of VACV Elstree was achieved with 0.0776 µM scFv-Fc-1.2.2.H9 and 0.01324 µM IgG1-1.2.2.H9, respectively. In an in vivo passive immunization NMRI mouse model, 100 µg purified scFv-1.2.2.H9 and the IgG1-1.2.2.H9 partially protected against the challenge with 4 LD<sub>50</sub> VACV Munich 1, as 3/6 mice survived. In contrast, in the scFv-Fc-1.2.2.H9 group, only one mouse survived the challenge.Ulrike S. DiesterbeckHenrike P. AhsendorfAndré FrenzelAhmad Reza SharifiThomas SchirrmannClaus-Peter CzernyMDPI AGarticlescFvvaccinia virusrecombinant antibodyD8MedicineRENVaccines, Vol 9, Iss 1308, p 1308 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	scFv vaccinia virus recombinant antibody D8 Medicine R
spellingShingle	scFv vaccinia virus recombinant antibody D8 Medicine R Ulrike S. Diesterbeck Henrike P. Ahsendorf André Frenzel Ahmad Reza Sharifi Thomas Schirrmann Claus-Peter Czerny Characterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library
description	A panel of potent neutralizing antibodies are protective against orthopoxvirus (OPXV) infections. For the development of OPXV-specific recombinant human single-chain antibodies (scFvs), the IgG repertoire of four vaccinated donors was amplified from peripheral B-lymphocytes. The resulting library consisted of ≥4 × 10<sup>8</sup> independent colonies. The immuno-screening against vaccinia virus (VACV) Elstree revealed a predominant selection of scFv clones specifically binding to the D8 protein. The scFv-1.2.2.H9 was engineered into larger human scFv-Fc-1.2.2.H9 and IgG1-1.2.2.H9 formats to improve the binding affinity and to add effector functions within the human immune response. Similar binding kinetics were calculated for scFv-1.2.2.H9 and scFv-Fc-1.2.2.H9 (1.61 nM and 7.685 nM, respectively), whereas, for IgG1-1.2.2.H9, the Michaelis-Menten kinetics revealed an increased affinity of 43.8 pM. None of the purified recombinant 1.2.2.H9 formats were able to neutralize VACV Elstree in vitro. After addition of 1% human complement, the neutralization of ≥50% of VACV Elstree was achieved with 0.0776 µM scFv-Fc-1.2.2.H9 and 0.01324 µM IgG1-1.2.2.H9, respectively. In an in vivo passive immunization NMRI mouse model, 100 µg purified scFv-1.2.2.H9 and the IgG1-1.2.2.H9 partially protected against the challenge with 4 LD<sub>50</sub> VACV Munich 1, as 3/6 mice survived. In contrast, in the scFv-Fc-1.2.2.H9 group, only one mouse survived the challenge.
format	article
author	Ulrike S. Diesterbeck Henrike P. Ahsendorf André Frenzel Ahmad Reza Sharifi Thomas Schirrmann Claus-Peter Czerny
author_facet	Ulrike S. Diesterbeck Henrike P. Ahsendorf André Frenzel Ahmad Reza Sharifi Thomas Schirrmann Claus-Peter Czerny
author_sort	Ulrike S. Diesterbeck
title	Characterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library
title_short	Characterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library
title_full	Characterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library
title_fullStr	Characterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library
title_full_unstemmed	Characterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library
title_sort	characterization of an in vivo neutralizing anti-vaccinia virus d8 single-chain fragment variable (scfv) from a human anti-vaccinia virus-specific recombinant library
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/46ac0221921e4932ad936734ce702782
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Characterization of an In Vivo Neutralizing Anti-Vaccinia Virus D8 Single-Chain Fragment Variable (scFv) from a Human Anti-Vaccinia Virus-Specific Recombinant Library

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