Variable heavy-variable light domain and Fab-arm CrossMabs with charged residue exchanges to enforce correct light chain assembly

Protein Engineering, Design & Selection : PEDS
Joerg Thomas RegulaChristian Klein

Abstract

Technologies for the production of bispecific antibodies need to overcome two major challenges. The first one is correct heavy chain assembly, which was solved by knobs-into-holes technology or charge interactions in the CH3 domains. The second challenge is correct light chain assembly. This can be solved by engineering the Fab-arm interfaces or applying the immunoglobulin domain crossover approach. There are three different crossovers possible, namely Fab-arm, constant domain and variable domain crossovers. The CrossMabCH1-CL exchange does not lead to the formation of unexpected side products, whereas the CrossMabFab and the CrossMabVH-VL formats result in the formation of typical side products. Thus, CrossMabCH1-CL was initially favored for therapeutic antibody development. Here, we report a novel improved CrossMab design principle making use of site-specific positional exchanges of charged amino acid pairs in the constant domain of these CrossMabs to enable the correct light chain assembly in the CrossMabVH-VL and improvements for the CrossMabFab design.

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Citations

Jun 27, 2019·Expert Opinion on Therapeutic Patents·Irma Herrera-CamachoGerardo Landeta
Sep 24, 2019·Antibodies·Qiong WangMichael J Betenbaugh
Jun 14, 2019·Science Translational Medicine·Christina ClausPablo Umaña
May 25, 2021·Frontiers in Immunology·Jiabing MaCheng Qian

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Methods Mentioned

BETA
size exclusion chromatography
electrophoresis
Chip
Gel Filtration
surface plasmon resonance
X-ray

Software Mentioned

BIOVIA Discovery Studio
PHASER
COOT
CrossMabs
CrossMab
CCP4
XDS
Discovery Studio

Related Concepts

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Antibody Engineering

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