Improvement of Biophysical Properties and Affinity of a Human Anti-L1CAM Therapeutic Antibody through Antibody Engineering Based on Computational Methods
The biophysical properties of therapeutic antibodies influence their manufacturability, efficacy, and safety. To develop an anti-cancer antibody, we previously generated a human monoclonal antibody (Ab417) that specifically binds to L1 cell adhesion molecule with a high affinity, and we validated it...
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| Veröffentlicht in: | International journal of molecular sciences 2021-07, Vol.22 (13), p.6696 |
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| creator | Chae, Heesu Cho, Seulki Jeong, Munsik Kwon, Kiyoung Choi, Dongwook Lee, Jaeyoung Nam, Woosuk Hong, Jisu Lee, Jiwoo Yoon, Seonjoo Hong, Hyojeong |
| description | The biophysical properties of therapeutic antibodies influence their manufacturability, efficacy, and safety. To develop an anti-cancer antibody, we previously generated a human monoclonal antibody (Ab417) that specifically binds to L1 cell adhesion molecule with a high affinity, and we validated its anti-tumor activity and mechanism of action in human cholangiocarcinoma xenograft models. In the present study, we aimed to improve the biophysical properties of Ab417. We designed 20 variants of Ab417 with reduced aggregation propensity, less potential post-translational modification (PTM) motifs, and the lowest predicted immunogenicity using computational methods. Next, we constructed these variants to analyze their expression levels and antigen-binding activities. One variant (Ab612)—which contains six substitutions for reduced surface hydrophobicity, removal of PTM, and change to the germline residue—exhibited an increased expression level and antigen-binding activity compared to Ab417. In further studies, compared to Ab417, Ab612 showed improved biophysical properties, including reduced aggregation propensity, increased stability, higher purification yield, lower pI, higher affinity, and greater in vivo anti-tumor efficacy. Additionally, we generated a highly productive and stable research cell bank (RCB) and scaled up the production process to 50 L, yielding 6.6 g/L of Ab612. The RCB will be used for preclinical development of Ab612. |
| doi_str_mv | 10.3390/ijms22136696 |
| format | Article |
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To develop an anti-cancer antibody, we previously generated a human monoclonal antibody (Ab417) that specifically binds to L1 cell adhesion molecule with a high affinity, and we validated its anti-tumor activity and mechanism of action in human cholangiocarcinoma xenograft models. In the present study, we aimed to improve the biophysical properties of Ab417. We designed 20 variants of Ab417 with reduced aggregation propensity, less potential post-translational modification (PTM) motifs, and the lowest predicted immunogenicity using computational methods. Next, we constructed these variants to analyze their expression levels and antigen-binding activities. One variant (Ab612)—which contains six substitutions for reduced surface hydrophobicity, removal of PTM, and change to the germline residue—exhibited an increased expression level and antigen-binding activity compared to Ab417. In further studies, compared to Ab417, Ab612 showed improved biophysical properties, including reduced aggregation propensity, increased stability, higher purification yield, lower pI, higher affinity, and greater in vivo anti-tumor efficacy. Additionally, we generated a highly productive and stable research cell bank (RCB) and scaled up the production process to 50 L, yielding 6.6 g/L of Ab612. The RCB will be used for preclinical development of Ab612.</description><identifier>ISSN: 1422-0067</identifier><identifier>ISSN: 1661-6596</identifier><identifier>EISSN: 1422-0067</identifier><identifier>DOI: 10.3390/ijms22136696</identifier><identifier>PMID: 34206616</identifier><language>eng</language><publisher>Basel: MDPI AG</publisher><subject>Affinity ; Antibodies ; Antigens ; Binding ; Cell adhesion ; Cell adhesion molecules ; Cholangiocarcinoma ; Chromatography ; Computer applications ; Design ; Engineering ; Hydrophobicity ; Immunogenicity ; Manufacturability ; Manufacturing ; Monoclonal antibodies ; Post-translation ; Proteins ; Xenografts ; Xenotransplantation</subject><ispartof>International journal of molecular sciences, 2021-07, Vol.22 (13), p.6696</ispartof><rights>2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). 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In further studies, compared to Ab417, Ab612 showed improved biophysical properties, including reduced aggregation propensity, increased stability, higher purification yield, lower pI, higher affinity, and greater in vivo anti-tumor efficacy. Additionally, we generated a highly productive and stable research cell bank (RCB) and scaled up the production process to 50 L, yielding 6.6 g/L of Ab612. The RCB will be used for preclinical development of Ab612.</description><subject>Affinity</subject><subject>Antibodies</subject><subject>Antigens</subject><subject>Binding</subject><subject>Cell adhesion</subject><subject>Cell adhesion molecules</subject><subject>Cholangiocarcinoma</subject><subject>Chromatography</subject><subject>Computer applications</subject><subject>Design</subject><subject>Engineering</subject><subject>Hydrophobicity</subject><subject>Immunogenicity</subject><subject>Manufacturability</subject><subject>Manufacturing</subject><subject>Monoclonal antibodies</subject><subject>Post-translation</subject><subject>Proteins</subject><subject>Xenografts</subject><subject>Xenotransplantation</subject><issn>1422-0067</issn><issn>1661-6596</issn><issn>1422-0067</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2021</creationdate><recordtype>article</recordtype><sourceid>8G5</sourceid><sourceid>BENPR</sourceid><sourceid>GUQSH</sourceid><sourceid>M2O</sourceid><recordid>eNpdkctuEzEUhi1ERUvLjgewxIYFA75MHHuDlEaFVkrVLsracjzHGUcz9mB7KuVReFvci6Cw8tHnT7-Pz0HoPSWfOVfki9-PmTHKhVDiFTqhLWMNIWL5-kV9jN7mvCeEcbZQb9AxbxkRgooT9OtqnFK8hxFCwdHhcx-n_pC9NQO-TXGCVDxkbEKHV8754MvhQTP4ch5NwKtQfLOh69U1vushmQnm4u0j3sbugEuf4rzr_4KLsPMBIPmww-cmQ4djwOs4TnMxxcdQn72G0scun6EjZ4YM757PU_Tj28Xd-rLZ3Hy_Wq82jeVSlaazzC2JXDiwnDoJrOVAGXOSE97aCoXsKqWd6oQgltGKzMI6Jxwz3G35Kfr6lDvN2xE6WweRzKCn5EeTDjoar_-9Cb7Xu3ivJROSLFkN-PgckOLPGXLRo88WhsEEiHPWbNFKrtRSyap--E_dxznVTz9aqqVKSVGtT0-WTTHnBO5PM5Toh53rlzvnvwEk5qGv</recordid><startdate>20210701</startdate><enddate>20210701</enddate><creator>Chae, Heesu</creator><creator>Cho, Seulki</creator><creator>Jeong, Munsik</creator><creator>Kwon, Kiyoung</creator><creator>Choi, Dongwook</creator><creator>Lee, Jaeyoung</creator><creator>Nam, Woosuk</creator><creator>Hong, Jisu</creator><creator>Lee, Jiwoo</creator><creator>Yoon, Seonjoo</creator><creator>Hong, Hyojeong</creator><general>MDPI AG</general><general>MDPI</general><scope>AAYXX</scope><scope>CITATION</scope><scope>3V.</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>8G5</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BENPR</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>GUQSH</scope><scope>K9.</scope><scope>M0S</scope><scope>M1P</scope><scope>M2O</scope><scope>MBDVC</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>Q9U</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20210701</creationdate><title>Improvement of Biophysical Properties and Affinity of a Human Anti-L1CAM Therapeutic Antibody through Antibody Engineering Based on Computational Methods</title><author>Chae, Heesu ; 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To develop an anti-cancer antibody, we previously generated a human monoclonal antibody (Ab417) that specifically binds to L1 cell adhesion molecule with a high affinity, and we validated its anti-tumor activity and mechanism of action in human cholangiocarcinoma xenograft models. In the present study, we aimed to improve the biophysical properties of Ab417. We designed 20 variants of Ab417 with reduced aggregation propensity, less potential post-translational modification (PTM) motifs, and the lowest predicted immunogenicity using computational methods. Next, we constructed these variants to analyze their expression levels and antigen-binding activities. One variant (Ab612)—which contains six substitutions for reduced surface hydrophobicity, removal of PTM, and change to the germline residue—exhibited an increased expression level and antigen-binding activity compared to Ab417. In further studies, compared to Ab417, Ab612 showed improved biophysical properties, including reduced aggregation propensity, increased stability, higher purification yield, lower pI, higher affinity, and greater in vivo anti-tumor efficacy. Additionally, we generated a highly productive and stable research cell bank (RCB) and scaled up the production process to 50 L, yielding 6.6 g/L of Ab612. The RCB will be used for preclinical development of Ab612.</abstract><cop>Basel</cop><pub>MDPI AG</pub><pmid>34206616</pmid><doi>10.3390/ijms22136696</doi><oa>free_for_read</oa></addata></record> |
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| subjects | Affinity Antibodies Antigens Binding Cell adhesion Cell adhesion molecules Cholangiocarcinoma Chromatography Computer applications Design Engineering Hydrophobicity Immunogenicity Manufacturability Manufacturing Monoclonal antibodies Post-translation Proteins Xenografts Xenotransplantation |
| title | Improvement of Biophysical Properties and Affinity of a Human Anti-L1CAM Therapeutic Antibody through Antibody Engineering Based on Computational Methods |
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