Humanized Antibody Animal Models with Variable-Region Gene Integration
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Solution Overview
Problem
Existing methods for producing fully human antibodies in non-human animal models face challenges such as limited diversity, low affinity, and immunogenicity due to endogenous antibody coding lineages, leading to unpredictable modifications and adverse clinical effects.
Innovation Solution
A non-human animal model is developed by operably linking human immunoglobulin variable region genes downstream of the immunoglobulin locus, maintaining endogenous gene expression and regulation, and reversing the transcription direction of the constant region to prevent V(D)J recombination, thereby enhancing antibody diversity and affinity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If engineered non-human animals carrying human antibody genes are used to produce fully human antibodies, then human antibody production is achieved, but antibody diversity is limited and V(D)J recombination efficiency is low
Solution Approach 1:
The patent segments the human antibody coding lineage into specific variable region genes (IGHV1-18, IGHV1-46, IGHV3-23, IGHV3-30-2, IGKV1-39-1, IGKV3-20) and introduces them individually into the non-human animal genome at defined loci, rather than attempting to introduce the entire complex human antibody coding system. This segmentation approach achieves functional human antibody production while avoiding the complexity and diversity limitations of complete human lineage introduction.
2Object-affected harmful factors
If endogenous antibody genes are present in non-human animal models, then the animal model is simpler to construct, but immunogenicity occurs due to endogenous antibody coding infiltration
Solution Approach 1:
The patent extracts and removes the endogenous variable region genes from the non-human animal immunoglobulin loci before introducing human variable region genes. Specifically, the mouse IGHV and IGKV variable region genes are deleted from their respective loci, preventing endogenous antibody coding infiltration that would cause immunogenicity. This extraction approach maintains construction simplicity while eliminating the harmful immunogenicity effect.
Solution Approach 2:
The patent uses human constant region genes (IGHC, IGKC) as intermediaries to bridge the introduction of human variable region genes into the non-human animal system. The human constant regions serve as functional mediators that accept the introduced human variable regions and enable proper antibody assembly and secretion, while the removed endogenous variable regions prevent immunogenicity. This intermediary approach resolves the contradiction between construction ease and immunogenicity prevention.
3Productivity
If display technology is used for antibody screening, then screening speed is rapid, but antibody diversity and affinity are limited due to lack of in-vivo processes
Solution Approach 1:
The patent performs preliminary action by introducing functional human variable region genes with complete V(D)J recombination capability into the non-human animal model before antibody screening. This preliminary genetic engineering enables the animal to perform in-vivo V(D)J recombination and affinity maturation processes naturally, generating high-affinity human antibodies in advance. When screening is performed, both rapid screening and high affinity are achieved because the animal has already performed the necessary biological processes.
Data Source
AI summary
The present application relates to a non-human animal and a method for preparing the non-human animal. The non-human animal is operably linked to a human immunoglobulin variable region gene downstream of an immunoglobulin locus. The present application further provides a method for generating an antibody through the non-human animal.