Taming interleukin-12: Engineering of bispecific antibody-based IL-12 mimetics with biased agonism capacities

被引：0

作者：

Lipinski, Britta ^{[1
,2
]}

Unmuth, Laura ^{[1
,2
]}

Arras, Paul ^{[1
]}

Endruszeit, Ron ^{[1
]}

Becker, Stefan ^{[2
]}

Roedel, Jonathan Mathias ^{[3
,4
]}

Scheller, Juergen ^{[3
,4
]}

Pudewell, Silke ^{[3
,4
]}

Floss, Doreen M. ^{[3
,4
]}

Krah, Simon ^{[2
]}

Harwardt, Julia ^{[2
]}

Doerner, Achim ^{[2
]}

Helming, Laura ^{[5
]}

Xu, Chunxiao ^{[5
]}

Menrad, Andreas ^{[2
]}

Evers, Andreas ^{[2
]}

Kolmar, Harald ^{[6
]}

Elter, Desislava ^{[2
]}

Pekar, Lukas ^{[2
]}

Zielonka, Stefan ^{[1
,2
]}

机构：

[1] Tech Univ Darmstadt, Inst Organ Chem & Biochem, Biomol Immunotherapy, Peter Grunberg Str 4, D-64287 Darmstadt, Germany

[2] Merck Healthcare KGaA, Antibody Discovery & Prot Engn, Darmstadt, Germany

[3] Heinrich Heine Univ Dusseldorf, Med Fac, Inst Biochem & Mol Biol 2, Dusseldorf, Germany

[4] Heinrich Heine Univ Dusseldorf, Univ Hosp Dusseldorf, Dusseldorf, Germany

[5] EMD Serono Res Ctr, Res Unit Oncol, Billerica, MA USA

[6] Tech Univ Darmstadt, Inst Organ Chem & Biochem, Appl Biochem, Darmstadt, Germany

来源：

PROTEIN SCIENCE | 2025年 / 34卷 / 03期

关键词：

antibody engineering; bispecific antibody; cytokine mimetic; IL-12; NK cell; single-domain antibody; surrogate cytokine; T cell; VHH; yeast surface display; SINGLE-DOMAIN ANTIBODIES; EXPRESSION; CYTOKINE; DELIVERY; CELL; PURIFICATION; MURINE; IL-23; NK;

D O I：

10.1002/pro.70072

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

In this work, we have generated bispecific interleukin (IL)-12 surrogate agonists based on camelid-derived single-domain antibodies (sdAbs) targeting the IL-12 receptor (IL-12R) subunits IL-12R beta 1 and IL-12R beta 2. Following immunization and antibody display-based paratope isolation, respective sdAbs were combinatorially reformatted into a monovalent bispecific architecture by grafting resulting paratopes onto the hinge region of a heterodimeric Fc region. Functional characterization using NK-92 cells enabled the identification of multiple different sdAb-based bispecifics displaying divergent IL-12R agonism capacities as analyzed by STAT4 phosphorylation. Further investigations by harnessing peripheral blood mononuclear cells (PBMCs) from healthy donors revealed attenuated pSTAT4 activation compared to recombinant human (rh) wild-type IL-12 regarding both natural killer (NK)-cell and T-cell activation but robust IL-12R agonism on stimulated T cells. While several sdAb-based IL-12 mimetics were nearly inactive on NK cells as well as T cells obtained from PBMCs, they elicited significant STAT4 phosphorylation and interferon (IFN)-gamma release on stimulated T cells as well as an IL-12-like transcriptional signature. Furthermore, we demonstrate that the activity of receptor agonism of generated bispecific IL-12 mimetics can also be biased towards stimulated T cells by changing the spatial orientation of the individual sdAbs within the molecular design architecture. Taken together, we present an alternative strategy to generate IL-12-like biologics with tailor-made characteristics.

引用

页数：18

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[1] Expression and function of IL-12 and IL-18 receptors on human tonsillar B cells [J].