Efficient spontaneous site-selective cysteine-mediated toxin attachment within a structural loop of antibodies.
| Autor: | Stadlmayr G; Christian Doppler Laboratory for Innovative Immunotherapeutics, Institute of Molecular Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Muthgasse 18, 1190 Vienna, Austria., Stracke F; Christian Doppler Laboratory for Innovative Immunotherapeutics, Institute of Molecular Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Muthgasse 18, 1190 Vienna, Austria., Stadlbauer K; Christian Doppler Laboratory for Innovative Immunotherapeutics, Institute of Molecular Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Muthgasse 18, 1190 Vienna, Austria., Rybka J; Institute of Molecular Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Muthgasse 18, 1190 Vienna, Austria., Dickgiesser S; Antibody Drug Conjugates and Targeted NBE Therapeutics, Merck Healthcare KGaA, Frankfurter Strasse 250, 64293 Darmstadt, Germany., Rasche N; Antibody Drug Conjugates and Targeted NBE Therapeutics, Merck Healthcare KGaA, Frankfurter Strasse 250, 64293 Darmstadt, Germany., Becker S; Protein Engineering & Antibody Technologies, Global Research and Development, Merck Healthcare KGaA, Frankfurter Straße 250, 64293 Darmstadt, Germany., Toleikis L; Protein Engineering & Antibody Technologies, Global Research and Development, Merck Healthcare KGaA, Frankfurter Straße 250, 64293 Darmstadt, Germany., Rüker F; Christian Doppler Laboratory for Innovative Immunotherapeutics, Institute of Molecular Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Muthgasse 18, 1190 Vienna, Austria., Knopp GW; Christian Doppler Laboratory for Innovative Immunotherapeutics, Institute of Molecular Biotechnology, Department of Biotechnology, University of Natural Resources and Life Sciences, Vienna (BOKU), Muthgasse 18, 1190 Vienna, Austria. Electronic address: gordana.wozniak@boku.ac.at. |
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| Jazyk: | angličtina |
| Zdroj: | Biochimica et biophysica acta. General subjects [Biochim Biophys Acta Gen Subj] 2022 Jul; Vol. 1866 (7), pp. 130155. Date of Electronic Publication: 2022 Apr 22. |
| DOI: | 10.1016/j.bbagen.2022.130155 |
| Abstrakt: | Background: Site-specific coupling of toxin entities to antibodies has become a popular method of synthesis of antibody-drug conjugates (ADCs), as it leads to a homogenous product and allows a free choice of a convenient site for conjugation. Methods: We introduced a short motif, containing a single cysteine surrounded by aromatic residues, into the N-terminal FG-loop of the C Results: Antibody mutants were amenable for rapid coupling with maleimide-based linker endowed toxin payload and the modifications did not impair their reactivity with target cell lines or negatively impact their biophysical properties. Without any previous reduction, up to 50% of the antibody preparation was found to be coupled with two toxins per molecule. After the isolation of this fraction with preparative hydrophobic interaction chromatography, the ADC could elicit a potent cytotoxic effect on the target cell lines. Conclusion: By fine-tuning the microenvironment of the reactive cysteine residue, this strategy offers a simplified protocol for production of site-selectively coupled ADCs. General Significance: Our unique approach allows the generation of therapeutic ADCs with controlled chemical composition, which facilitates the optimization of their pharmacological activity. This strategy for directional coupling could in the future simplify the construction of ADCs with double payloads ("dual warheads") introduced with orthogonal techniques. (Copyright © 2022 Elsevier B.V. All rights reserved.) |
| Databáze: | MEDLINE |
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