Antibody humanization-the Influence of the antibody framework on the CDR-H3 loop ensemble in solution
© The Author(s) 2020. Published by Oxford University Press..
Antibody engineering of non-human antibodies has focused on reducing immunogenicity by humanization, being a major limitation in developing monoclonal antibodies. We analyzed four series of antibody binding fragments (Fabs) and a variable fragment (Fv) with structural information in different stages of humanization to investigate the influence of the framework, point mutations and specificity on the complementarity determining region (CDR)-H3 loop dynamics. We also studied a Fv without structural information of the anti-idiotypic antibody Ab2/3H6, because it completely lost its binding affinity upon superhumanization, as an example of a failed humanization. Enhanced sampling techniques in combination with molecular dynamics simulations allow to access micro- to milli-second timescales of the CDR-H3 loop dynamics and reveal kinetic and thermodynamic changes involved in the process of humanization. In most cases, we observe a reduced conformational diversity of the CDR-H3 loop when grafted on a human framework and find a conformational shift of the dominant CDR-H3 loop conformation in solution. A shallow side minimum of the conformational CDR-H3 loop ensemble attached to the murine framework becomes the dominant conformation in solution influenced by the human framework. Additionally, we observe in the case of the failed humanization that the potentially binding competent murine CDR-H3 loop ensemble in solution shows nearly no kinetical or structural overlap with the superhumanized variant, thus explaining the loss of binding.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2019 |
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| Erschienen: |
2019 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:32 |
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| Enthalten in: |
Protein engineering, design & selection : PEDS - 32(2019), 9 vom: 31. Dez., Seite 411-422 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Fernández-Quintero, Monica L [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 30.06.2020 Date Revised 14.02.2024 published: Print Citation Status MEDLINE |
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| doi: |
10.1093/protein/gzaa004 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM307207722 |
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| 245 | 1 | 0 | |a Antibody humanization-the Influence of the antibody framework on the CDR-H3 loop ensemble in solution |
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| 500 | |a Date Revised 14.02.2024 | ||
| 500 | |a published: Print | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a © The Author(s) 2020. Published by Oxford University Press. | ||
| 520 | |a Antibody engineering of non-human antibodies has focused on reducing immunogenicity by humanization, being a major limitation in developing monoclonal antibodies. We analyzed four series of antibody binding fragments (Fabs) and a variable fragment (Fv) with structural information in different stages of humanization to investigate the influence of the framework, point mutations and specificity on the complementarity determining region (CDR)-H3 loop dynamics. We also studied a Fv without structural information of the anti-idiotypic antibody Ab2/3H6, because it completely lost its binding affinity upon superhumanization, as an example of a failed humanization. Enhanced sampling techniques in combination with molecular dynamics simulations allow to access micro- to milli-second timescales of the CDR-H3 loop dynamics and reveal kinetic and thermodynamic changes involved in the process of humanization. In most cases, we observe a reduced conformational diversity of the CDR-H3 loop when grafted on a human framework and find a conformational shift of the dominant CDR-H3 loop conformation in solution. A shallow side minimum of the conformational CDR-H3 loop ensemble attached to the murine framework becomes the dominant conformation in solution influenced by the human framework. Additionally, we observe in the case of the failed humanization that the potentially binding competent murine CDR-H3 loop ensemble in solution shows nearly no kinetical or structural overlap with the superhumanized variant, thus explaining the loss of binding | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 4 | |a CDR-H3 loop dynamics | |
| 650 | 4 | |a conformational selection | |
| 650 | 4 | |a humanization | |
| 650 | 4 | |a immunogenicity | |
| 650 | 4 | |a interdomain dynamics | |
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| 650 | 7 | |a Complementarity Determining Regions |2 NLM | |
| 650 | 7 | |a Solutions |2 NLM | |
| 700 | 1 | |a Heiss, Martin C |e verfasserin |4 aut | |
| 700 | 1 | |a Liedl, Klaus R |e verfasserin |4 aut | |
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