Multispecific Antibody Chain Pairing via Interface Engineering
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Solution Overview
Problem
Developing IgG-like bispecific antibodies is challenging due to mispaired chain combinations during co-expression, leading to low yield and stability issues, and the selection of common light chains that maintain binding affinity is inefficient, requiring extensive optimization and empirical testing.
Innovation Solution
A method involving competition and non-competition Chain Selectivity Assessment (CSA) to identify optimal pairings of heavy and light chain complementarity-determining regions (CDRs) for multispecific antibody constructs, using high-throughput screening to select for minimal cross-pairing and optimal binding affinities.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If IgG-like bisspecific antibodies are developed using co-expression of multiple heavy and light chains, then the stability and cell production are improved, but mispaired chain combinations occur leading to low purification yield
Solution Approach 1:
The invention divides the antibody chains into modular domains (VH, VL, CH1, CL) that can be independently engineered and assembled. By segmenting the chains and introducing specific interface modifications, the system enables controlled pairing while maintaining the benefits of co-expression.
Solution Approach 2:
The invention applies localized modifications at the chain interface regions (CH1-CL interface) rather than throughout the entire chain. Specific amino acid substitutions are introduced at the pairing interface to enhance cognate interactions while leaving the rest of the chain structure unchanged, thus improving specificity without compromising overall stability.
2Manufacturing precision
If engineering strategies are applied to favor cognate HC/LC pairing, then pairing specificity is improved, but sequence diversity limits the success when applied as a rigid platform
Solution Approach 1:
The invention creates a universal platform with standardized interface modifications that can be applied across diverse antibody sequences. The CH1-CL interface engineering approach serves multiple functions: it enhances pairing specificity, maintains compatibility with various CDR sequences, and works across different antibody formats and specificities.
Solution Approach 2:
The invention modifies physical-chemical parameters at the chain interface (charge distribution, hydrophobicity, steric complementarity) to create favorable pairing conditions. By changing these parameters through targeted amino acid substitutions, the system achieves high pairing specificity that is adaptable to diverse sequence backgrounds without requiring sequence-specific optimization.
3Device complexity
If common light chains are used to avoid pairing engineering, then the need for HC/LC pairing optimization is reduced, but identifying suitable cLC is challenging and requires significant investment
Solution Approach 1:
The invention performs preliminary engineering of the CH1-CL interface during the early design stage, creating a platform that inherently favors correct pairing. This preliminary action eliminates the need for later optimization efforts and avoids the time-consuming process of screening for suitable common light chains, as the interface modifications work universally with diverse LC sequences.
Data Source
AI summary
The ability to generate a single antibody-based construct that can recognize multiple targets simultaneously, is paramount to advance many therapeutics candidates to clinic. Often, this implies extensive protein design with vary degrees of success. In the case of multispecific antibody constructs, there are multiple modalities from which to choose and often multiple antigen binders as well. Described here is the discovery of new methods to optimally pair antigen binders with the proper format, including the selection of common light chains.