Common Light Chain Mouse for In Vivo Bispecific Antibody Pairing
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Solution Overview
Problem
Existing methods for creating bispecific antibodies face challenges in selecting a suitable light chain component that can associate with both heavy chains, often relying on in vitro manipulations that impose sequence identity and are not biologically sensible, and there is a need for compositions and methods that employ more biologically sensible approaches.
Innovation Solution
Genetically modified mice are engineered to express human immunoglobulin heavy and light chain variable domains with a limited repertoire, allowing the mice to select suitable affinity-matured human immunoglobulin heavy and light chain variable domains, wherein the mice have a limited repertoire, allowing the mice to select suitable affinity-matured human immunoglobulin heavy chain variable domains derived from a repertoire of unrearranged human heavy chain variable region gene segments, which associate and express with a single human light chain variable domain.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If in vitro methods are used to select light chains by surveying usage statistics or testing in phage display libraries, then a light chain can be identified that associates with heavy chains, but the process relies on artificial manipulations that impose sequence identity constraints and are not biologically sensible
Solution Approach 1:
The mouse immune system naturally performs the selection function that in vitro methods attempt to replicate. By immunizing the transgenic mouse with the target antigen, the mouse's B cells automatically select and produce the appropriate light chain that pairs with the heavy chain variable region, eliminating the need for artificial in vitro screening and manipulation processes
Solution Approach 2:
The patent replaces the mechanical in vitro selection process (phage display library screening, in vitro association assays) with a biological in vivo system. The mouse's immune system naturally performs the selection and pairing function through normal immunological processes, substituting artificial mechanical manipulation with biological self-organization
2Manufacturing precision
If a light chain is selected by aligning with cognate light chains and making modifications to match sequence characteristics, then sequence identity is improved, but the process requires extensive in vitro manipulation and modification
Solution Approach 1:
The transgenic mouse is pre-engineered with human heavy chain variable region gene segments in its genome before immunization. This preliminary genetic setup ensures that when the mouse is immunized, it will naturally produce heavy chains with the desired human variable regions that can be paired with appropriate light chains, eliminating the need for subsequent in vitro alignment and modification work
Solution Approach 2:
The mouse's immune system naturally performs the sequence optimization function. Through normal V(D)J recombination and somatic hypermutation processes, the mouse generates heavy chain variable regions with appropriate sequence characteristics that are already optimized for pairing with light chains, eliminating the need for artificial sequence alignment and modification procedures
3Reliability
If in vitro association assays are performed to test light chain compatibility with heavy chains, then binding capability is confirmed, but the process is time-consuming and requires multiple testing iterations
Solution Approach 1:
The transgenic mouse's immune system naturally performs the binding compatibility test during the immune response. When immunized with the antigen, the mouse's B cells automatically produce the correct light chain that binds to the heavy chain variable region, confirming binding capability through natural physiological function rather than requiring separate in vitro association assays
Solution Approach 2:
The patent extracts the binding capability confirmation function from the separate in vitro association assay step and integrates it into the in vivo immunization process. The binding capability is confirmed naturally during the immune response in the mouse, eliminating the need for time-consuming separate in vitro testing iterations
Data Source
AI summary
A genetically modified mouse is provided, wherein the mouse is incapable of rearranging and expressing an endogenous mouse immunoglobulin light chain variable sequence, wherein the mouse expresses only one or two human light chain variable domains encoded by human immunoglobulin sequences operably linked to the mouse kappa (κ) constant gene at the endogenous mouse κ locus, wherein the mouse expresses a reverse chimeric antibody having a light chain variable domain derived from one of only two human light chain variable region gene segments and a mouse κ constant domain, and a human heavy chain variable domain and a mouse heavy chain constant domain, from an endogenous mouse heavy chain locus. Bispecific epitope-binding proteins that are fully human are provided, comprising two different heavy chains that associate with an identical light chain that comprises a variable domain derived from one of two different human light chain variable region gene segments.