Generation and engineering of potent single domain antibody-based bispecific IL-18 mimetics resistant to IL-18BP decoy receptor inhibition
Britta Lipinski,
Laura Unmuth,
Paul Arras,
Stefan Becker,
Christina Bauer,
Lars Toleikis,
Simon Krah,
Achim Doerner,
Desislava Yanakieva,
Ammelie Svea Boje,
Katja Klausz,
Matthias Peipp,
Vanessa Siegmund,
Andreas Evers,
Harald Kolmar,
Lukas Pekar,
Stefan Zielonka
Affiliations
Britta Lipinski
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
Laura Unmuth
Early Protein Supply and Characterization (EPSC), Merck Healthcare KGaA, Darmstadt, Germany
Paul Arras
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
Stefan Becker
Early Protein Supply and Characterization (EPSC), Merck Healthcare KGaA, Darmstadt, Germany
Christina Bauer
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
Lars Toleikis
Early Protein Supply and Characterization (EPSC), Merck Healthcare KGaA, Darmstadt, Germany
Simon Krah
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
Achim Doerner
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
Desislava Yanakieva
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
Ammelie Svea Boje
Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein and Christian-Albrechts-University Kiel, Kiel, Germany
Katja Klausz
Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein and Christian-Albrechts-University Kiel, Kiel, Germany
Matthias Peipp
Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein and Christian-Albrechts-University Kiel, Kiel, Germany
Vanessa Siegmund
Early Protein Supply and Characterization (EPSC), Merck Healthcare KGaA, Darmstadt, Germany
Andreas Evers
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
Harald Kolmar
Institute for Organic Chemistry and Biochemistry, Technical University of Darmstadt, Darmstadt, Germany
Lukas Pekar
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
Stefan Zielonka
Antibody Discovery and Protein Engineering (ADPE), Merck Healthcare KGaA, Darmstadt, Germany
ABSTRACTHere, we generated bispecific antibody (bsAb) derivatives that mimic the function of interleukin (IL)-18 based on single domain antibodies (sdAbs) specific to IL-18 Rα and IL-18 Rβ. For this, camelids were immunized, followed by yeast surface display (YSD)-enabled discovery of VHHs targeting the individual receptor subunits. Upon reformatting into a strictly monovalent (1 + 1) bispecific sdAb architecture, several bsAbs triggered dose-dependent IL-18 R downstream signaling on IL-18 reporter cells, as well as IFN-γ release by peripheral blood mononuclear cells in the presence of low-dose IL-12. However, compared with IL-18, potencies and efficacies were considerably attenuated. By engineering paratope valencies and the spatial orientation of individual paratopes within the overall design architecture, we were able to generate IL-18 mimetics displaying significantly augmented functionalities, resulting in bispecific cytokine mimetics that were more potent than IL-18 in triggering proinflammatory cytokine release. Furthermore, generated IL-18 mimetics were unaffected from inhibition by IL-18 binding protein decoy receptor. Essentially, we demonstrate that this strategy enables the generation of IL-18 mimetics with tailor-made cytokine functionalities.
