Development of a Human IgG4 Bispecific Antibody for Dual Targeting of Interleukin-4 (IL-4) and Interleukin-13 (IL-13) Cytokines

Justin M. Scheer,Dorothea Reilly,Heleen Scheerens,Gerald Nakamura,Michael A. Dillon ,Glen Giese,Hongbin Liu,Enzo Palma,P. Molina ,Jack Bevers,Prateek Gupta,Josefine Persson,Eric Stefanich,Mark S. Dennis,James Giulianotti,Janet Jackman,Yin Zhang,Christoph Spiess,J. Michael Elliott,Lawren C. Wu,Jianyong Wang,Whitney Shatz,Nancy Y. Chiang ,Germaine Fuh

doi:10.1074/jbc.m113.480483

Abstract

Human bispecific antibodies have great potential for the treatment of human diseases. Although human IgG1 bispecific antibodies have been generated, few attempts have been reported in the scientific literature that extend bispecific antibodies to other human antibody isotypes. In this paper, we report our work expanding the knobs-into-holes bispecific antibody technology to the human IgG4 isotype. We apply this approach to generate a bispecific antibody that targets IL-4 and IL-13, two cytokines that play roles in type 2 inflammation. We show that IgG4 bispecific antibodies can be generated in large quantities with equivalent efficiency and quality and have comparable pharmacokinetic properties and lung partitioning, compared with the IgG1 isotype. This work broadens the range of published therapeutic bispecific antibodies with natural surface architecture and provides additional options for the generation of bispecific antibodies with differing effector functions through the use of different antibody isotypes.

Highlights

Dual neutralization of IL-4 and IL-13 is a promising therapeutic approach for asthma and allergy
Monoclonal antibodies to human IL-4 were generated from BALB/c mice immunized with human IL-4
Clone 19C11 bound to human IL-4 with an affinity of 10 pM, as determined by surface plasmon resonance analysis. 19C11 blocked binding of biotinylated IL-4 to IL-4R␣ and inhibited IL4-induced proliferation of TF-1 cells (Fig. 1), suggesting an epitope on IL-4 that overlaps with a region critical for binding to IL-4R␣. 19C11 was subsequently humanized by grafting the hypervariable region into a human V␬ 1/VHIII consensus framework along with two mouse vernier positions at 71 and 78 in the variable heavy domain

Summary

Background

Dual neutralization of IL-4 and IL-13 is a promising therapeutic approach for asthma and allergy. We report our work expanding the knobs-into-holes bispecific antibody technology to the human IgG4 isotype We apply this approach to generate a bispecific antibody that targets IL-4 and IL-13, two cytokines that play roles in type 2 inflammation. Most of these bispecific formats contain non-natural or non-human sequences that serve as linkers, binding sites, or heterodimerization domains These sequences can lead to poor stability, aggregation, and large scale manufacturing challenges. Human IgG4 Anti-IL-4/IL-13 Bispecific Antibody dimer interface This technology has been used previously to generate bispecific antibodies of IgG1 subclass with a common light chain [11, 12] and has subsequently been extended to bispecific antibodies with two different light chains through the use of linker sequences [13] or domain swaps [14]. In order to match the bispecific antibody isotype to that of lebrikizumab, we used knobs-intoholes technology to create human IgG4 bispecifics and showed that such IgG4 bispecific antibodies can be generated with equivalent efficiency and quality and have activities comparable with those of IgG1 bispecific antibodies

EXPERIMENTAL PROCEDURES

RESULTS

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DISCUSSION