Generation and engineering of potent single domain antibody-based bispecific IL-18 mimetics resistant to IL-18BP decoy receptor inhibition

ABSTRACT

Here, 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.

KEYWORDS:

• Antibody engineering
• bispecific antibody
• cytokine mimetic
• IL-18
• IL-18 binding protein
• IL-18BP
• single domain antibody
• surrogate agonist
• valencies
• VHH
• yeast surface display

Acknowledgments

We thank Hannah-Melina Mayer, Kerstin Hallstein, Marion Wetter, Pia Stroh, Sigrid Auth, Gernot Musch, Markus Fleischer and Dirk Mueller-Pompalla for experimental support.

Disclosure statement

BL, PA, LU, LP and SZ filed a patent application based on this work. In addition, BL, PA, LU, LP, SK, SB, LT, VS, AE, and SZ are employees at Merck Healthcare KGaA. Besides, this work was conducted in the absence of any further commercial interest.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/19420862.2023.2236265

Additional information

Funding

The author(s) reported that there is no funding associated with the work featured in this article.