Protease-Activation of Fc-Masked Therapeutic Antibodies to Alleviate Off-Tumor Cytotoxicity
The interaction of the Fc region of therapeutic antibodies and antibody-drug conjugates with Fcγ receptors (FcγRs) can lead to unpredictable and severe side effects. Over the last decades several strategies have been developed to overcome this drawback, including extensive Fc- and glycoengineering a...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2021-08-01
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| Series: | Frontiers in Immunology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fimmu.2021.715719/full |
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| author | Adrian Elter Desislava Yanakieva Desislava Yanakieva David Fiebig Kerstin Hallstein Stefan Becker Ulrich Betz Harald Kolmar |
| author_facet | Adrian Elter Desislava Yanakieva Desislava Yanakieva David Fiebig Kerstin Hallstein Stefan Becker Ulrich Betz Harald Kolmar |
| author_sort | Adrian Elter |
| collection | DOAJ |
| description | The interaction of the Fc region of therapeutic antibodies and antibody-drug conjugates with Fcγ receptors (FcγRs) can lead to unpredictable and severe side effects. Over the last decades several strategies have been developed to overcome this drawback, including extensive Fc- and glycoengineering and antibody isotype switching. However, these approaches result in permanently Fc-silenced antibody derivates which partially or completely lack antibody-mediated effector functions. Nevertheless, for a majority of antibody-based drugs, Fc-mediated effector functions, like antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cell-mediated phagocytosis (ADCP) as well as complement-dependent cytotoxicity (CDC), represent the most substantial modes of action. We argued that a new strategy combining the beneficial properties of Fc-silencing and controlled activation of effector functions can pave the way to potent antibody therapeutics, reducing the FcγRs-mediated off-target toxicity. We present a novel Fc-tamed antibody format, where the FcγR-binding sites of antibodies are blocked by anti-isotypic masking units, hindering the association of FcγR and complement component 1 (c1q) to the Fc domain. The masking units were genetically fused to trastuzumab, including a protease-addressable peptide-liker. Our Fc-tamed antibodies demonstrated completely abolished interaction to soluble high-affinity Fcγ-Receptor I and c1q. In reporter cell-based ADCC assays, our Fc-tamed antibodies exhibited a 2,700 to 7,100-fold reduction in activation, compared to trastuzumab. Upon demasking by a tumor-associated protease, the Fc-activated antibodies demonstrated restored FcγR-binding, c1q-binding and the ability to induce potent ADCC activation. Furthermore, cell killing assays using donor-derived NK cells were performed to validate the functionality of the Fc-tamed antibody variants. To our knowledge, this approach represents the first non-permanently Fc-silenced antibody, which can be re-activated by a tumor-associated protease, eventually extending the field of novel antibody formats. |
| first_indexed | 2024-12-17T19:59:10Z |
| format | Article |
| id | doaj.art-e83dda1bb54546cd838edb4dfd165371 |
| institution | Directory Open Access Journal |
| issn | 1664-3224 |
| language | English |
| last_indexed | 2024-12-17T19:59:10Z |
| publishDate | 2021-08-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Immunology |
| spelling | doaj.art-e83dda1bb54546cd838edb4dfd1653712022-12-21T21:34:32ZengFrontiers Media S.A.Frontiers in Immunology1664-32242021-08-011210.3389/fimmu.2021.715719715719Protease-Activation of Fc-Masked Therapeutic Antibodies to Alleviate Off-Tumor CytotoxicityAdrian Elter0Desislava Yanakieva1Desislava Yanakieva2David Fiebig3Kerstin Hallstein4Stefan Becker5Ulrich Betz6Harald Kolmar7Institute for Organic Chemistry and Biochemistry, Technical University of Darmstadt, Darmstadt, GermanyInstitute for Organic Chemistry and Biochemistry, Technical University of Darmstadt, Darmstadt, GermanyProtein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, GermanyInstitute for Organic Chemistry and Biochemistry, Technical University of Darmstadt, Darmstadt, GermanyProtein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, GermanyProtein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, GermanyProtein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, GermanyInstitute for Organic Chemistry and Biochemistry, Technical University of Darmstadt, Darmstadt, GermanyThe interaction of the Fc region of therapeutic antibodies and antibody-drug conjugates with Fcγ receptors (FcγRs) can lead to unpredictable and severe side effects. Over the last decades several strategies have been developed to overcome this drawback, including extensive Fc- and glycoengineering and antibody isotype switching. However, these approaches result in permanently Fc-silenced antibody derivates which partially or completely lack antibody-mediated effector functions. Nevertheless, for a majority of antibody-based drugs, Fc-mediated effector functions, like antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cell-mediated phagocytosis (ADCP) as well as complement-dependent cytotoxicity (CDC), represent the most substantial modes of action. We argued that a new strategy combining the beneficial properties of Fc-silencing and controlled activation of effector functions can pave the way to potent antibody therapeutics, reducing the FcγRs-mediated off-target toxicity. We present a novel Fc-tamed antibody format, where the FcγR-binding sites of antibodies are blocked by anti-isotypic masking units, hindering the association of FcγR and complement component 1 (c1q) to the Fc domain. The masking units were genetically fused to trastuzumab, including a protease-addressable peptide-liker. Our Fc-tamed antibodies demonstrated completely abolished interaction to soluble high-affinity Fcγ-Receptor I and c1q. In reporter cell-based ADCC assays, our Fc-tamed antibodies exhibited a 2,700 to 7,100-fold reduction in activation, compared to trastuzumab. Upon demasking by a tumor-associated protease, the Fc-activated antibodies demonstrated restored FcγR-binding, c1q-binding and the ability to induce potent ADCC activation. Furthermore, cell killing assays using donor-derived NK cells were performed to validate the functionality of the Fc-tamed antibody variants. To our knowledge, this approach represents the first non-permanently Fc-silenced antibody, which can be re-activated by a tumor-associated protease, eventually extending the field of novel antibody formats.https://www.frontiersin.org/articles/10.3389/fimmu.2021.715719/fullFc gamma receptoroff-target cytotoxicityeffector functionFc-silencingmasked therapeutic antibodyMMP-9 |
| spellingShingle | Adrian Elter Desislava Yanakieva Desislava Yanakieva David Fiebig Kerstin Hallstein Stefan Becker Ulrich Betz Harald Kolmar Protease-Activation of Fc-Masked Therapeutic Antibodies to Alleviate Off-Tumor Cytotoxicity Frontiers in Immunology Fc gamma receptor off-target cytotoxicity effector function Fc-silencing masked therapeutic antibody MMP-9 |
| title | Protease-Activation of Fc-Masked Therapeutic Antibodies to Alleviate Off-Tumor Cytotoxicity |
| title_full | Protease-Activation of Fc-Masked Therapeutic Antibodies to Alleviate Off-Tumor Cytotoxicity |
| title_fullStr | Protease-Activation of Fc-Masked Therapeutic Antibodies to Alleviate Off-Tumor Cytotoxicity |
| title_full_unstemmed | Protease-Activation of Fc-Masked Therapeutic Antibodies to Alleviate Off-Tumor Cytotoxicity |
| title_short | Protease-Activation of Fc-Masked Therapeutic Antibodies to Alleviate Off-Tumor Cytotoxicity |
| title_sort | protease activation of fc masked therapeutic antibodies to alleviate off tumor cytotoxicity |
| topic | Fc gamma receptor off-target cytotoxicity effector function Fc-silencing masked therapeutic antibody MMP-9 |
| url | https://www.frontiersin.org/articles/10.3389/fimmu.2021.715719/full |
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