Binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates.
An increasingly appreciated conundrum in the discovery of antibody drug conjugates (ADCs) is that an antibody that was selected primarily for strong binding to its cancer target may not serve as an optimal ADC. In this study, we performed mechanistic cell-based experiments to determine the correlati...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2019-01-01
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| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0226593 |
| _version_ | 1818919161485590528 |
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| author | John C Zwaagstra Traian Sulea Jason Baardsnes Stevo Radinovic Yuneivy Cepero-Donates Alma Robert Maureen D O'Connor-McCourt Ilia A Tikhomirov Maria Luz Jaramillo |
| author_facet | John C Zwaagstra Traian Sulea Jason Baardsnes Stevo Radinovic Yuneivy Cepero-Donates Alma Robert Maureen D O'Connor-McCourt Ilia A Tikhomirov Maria Luz Jaramillo |
| author_sort | John C Zwaagstra |
| collection | DOAJ |
| description | An increasingly appreciated conundrum in the discovery of antibody drug conjugates (ADCs) is that an antibody that was selected primarily for strong binding to its cancer target may not serve as an optimal ADC. In this study, we performed mechanistic cell-based experiments to determine the correlation between antibody affinity, avidity, internalization and ADC efficacy. We used structure-guided design to assemble a panel of antibody mutants with predicted Her2 affinities ranging from higher to lower relative to the parent antibody, Herceptin. These antibodies were ranked for binding via SPR and via flow-cytometry on high-Her2 SKOV3 cells and low-Her2 MCF7 cells, the latter acting as a surrogate for low-Her2 normal cells. A subpanel of variants, representative of different Her2-binding affinities (2 strong, 2 moderate and 3 weak), were further screened via high-content imaging for internalization efficacies in high versus low-Her2 cells. Finally, these antibodies were evaluated in ADC cytotoxicity screening assays (using DM1 and MMAE secondary antibodies) and as antibody-drug conjugates (DM1 and PNU159682). Our results identified specific but weak Her2-binding variants as optimal candidates for developing DM1 and PNU ADCs since they exhibited high potencies (low to sub-nM) in high-Her2 SKOV3 cells and low toxicities in low-Her2 cells. The 2 strong-affinity variants were highly potent in SKOV3 cells but also showed significant toxicities in low-Her2 cells and therefore are predicted to be toxic in normal tissues. Our findings show that pharmacological profiling of an antibody library in multiple binding and functional assays allows for selection of optimal ADCs. |
| first_indexed | 2024-12-20T01:01:27Z |
| format | Article |
| id | doaj.art-3ffbe2edbb7b4027ac905174dfae341a |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-20T01:01:27Z |
| publishDate | 2019-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-3ffbe2edbb7b4027ac905174dfae341a2022-12-21T19:58:58ZengPublic Library of Science (PLoS)PLoS ONE1932-62032019-01-011412e022659310.1371/journal.pone.0226593Binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates.John C ZwaagstraTraian SuleaJason BaardsnesStevo RadinovicYuneivy Cepero-DonatesAlma RobertMaureen D O'Connor-McCourtIlia A TikhomirovMaria Luz JaramilloAn increasingly appreciated conundrum in the discovery of antibody drug conjugates (ADCs) is that an antibody that was selected primarily for strong binding to its cancer target may not serve as an optimal ADC. In this study, we performed mechanistic cell-based experiments to determine the correlation between antibody affinity, avidity, internalization and ADC efficacy. We used structure-guided design to assemble a panel of antibody mutants with predicted Her2 affinities ranging from higher to lower relative to the parent antibody, Herceptin. These antibodies were ranked for binding via SPR and via flow-cytometry on high-Her2 SKOV3 cells and low-Her2 MCF7 cells, the latter acting as a surrogate for low-Her2 normal cells. A subpanel of variants, representative of different Her2-binding affinities (2 strong, 2 moderate and 3 weak), were further screened via high-content imaging for internalization efficacies in high versus low-Her2 cells. Finally, these antibodies were evaluated in ADC cytotoxicity screening assays (using DM1 and MMAE secondary antibodies) and as antibody-drug conjugates (DM1 and PNU159682). Our results identified specific but weak Her2-binding variants as optimal candidates for developing DM1 and PNU ADCs since they exhibited high potencies (low to sub-nM) in high-Her2 SKOV3 cells and low toxicities in low-Her2 cells. The 2 strong-affinity variants were highly potent in SKOV3 cells but also showed significant toxicities in low-Her2 cells and therefore are predicted to be toxic in normal tissues. Our findings show that pharmacological profiling of an antibody library in multiple binding and functional assays allows for selection of optimal ADCs.https://doi.org/10.1371/journal.pone.0226593 |
| spellingShingle | John C Zwaagstra Traian Sulea Jason Baardsnes Stevo Radinovic Yuneivy Cepero-Donates Alma Robert Maureen D O'Connor-McCourt Ilia A Tikhomirov Maria Luz Jaramillo Binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates. PLoS ONE |
| title | Binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates. |
| title_full | Binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates. |
| title_fullStr | Binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates. |
| title_full_unstemmed | Binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates. |
| title_short | Binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates. |
| title_sort | binding and functional profiling of antibody mutants guides selection of optimal candidates as antibody drug conjugates |
| url | https://doi.org/10.1371/journal.pone.0226593 |
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