A disruptive clickable antibody design for the generation of antibody-drug conjugates

Nathanaël Rakotoarinoro; Yan F K Dyck; Simon K Krebs; Miriam-Kousso Assi; Maria K Parr; Marlitt Stech

doi:10.1093/abt/tbad023

A disruptive clickable antibody design for the generation of antibody-drug conjugates

Antib Ther. 2023 Oct 26;6(4):298-310. doi: 10.1093/abt/tbad023. eCollection 2023 Oct.

Authors

Nathanaël Rakotoarinoro^{1

2}, Yan F K Dyck², Simon K Krebs^{1

3}, Miriam-Kousso Assi^{1

4}, Maria K Parr², Marlitt Stech¹

Affiliations

¹ Institute for Cell Therapy and Immunology branch Bioanalytics and Bioprocesses, Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e.V., 14476 Potsdam-Golm, Germany.
² Institute of Pharmacy, Freie Universität Berlin, 14195 Berlin, Germany.
³ Institute of Biotechnology, Technische Universität Berlin, 13355 Berlin, Germany.
⁴ Department of Biotechnology, Hamburg University of Applied Sciences, 21033 Hamburg, Germany.

PMID: 38107665
PMCID: PMC10720948 (available on 2024-10-26)
DOI: 10.1093/abt/tbad023

Abstract

Background: Antibody-drug conjugates are cancer therapeutics that combine specificity and toxicity. A highly cytotoxic drug is covalently attached to an antibody that directs it to cancer cells. The conjugation of the drug-linker to the antibody is a key point in research and development as well as in industrial production. The consensus is to conjugate the drug to a surface-exposed part of the antibody to ensure maximum conjugation efficiency. However, the hydrophobic nature of the majority of drugs used in antibody-drug conjugates leads to an increased hydrophobicity of the generated antibody-drug conjugates, resulting in higher liver clearance and decreased stability.

Methods: In contrast, we describe a non-conventional approach in which the drug is conjugated in a buried part of the antibody. To achieve this, a ready-to-click antibody design was created in which an azido-based non-canonical amino acid is introduced within the Fab cavity during antibody synthesis using nonsense suppression technology. The Fab cavity was preferred over the Fc cavity to circumvent issues related to cleavage of the IgG1 lower hinge region in the tumor microenvironment.

Results: This antibody design significantly increased the hydrophilicity of the generated antibody-drug conjugates compared to the current best-in-class designs based on non-canonical amino acids, while conjugation efficiency and functionality were maintained. The robustness of this native shielding effect and the versatility of this approach were also investigated.

Conclusions: This pioneer design may become a starting point for the improvement of antibody-drug conjugates and an option to consider for protecting drugs and linkers from unspecific interactions.

Keywords: antibody-drug conjugates; fab cavity; hydrophilicity; hydrophobicity; native shielding; non-canonical amino acids.