Common Light Chain Transgenic Animals Using Gene Conversion Repair
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Solution Overview
Problem
Producing antibodies with the same light chain is challenging due to somatic hyper-mutation, which changes the light chain variable region coding sequence during affinity maturation in B cells, leading to light chain diversity in classical antibodies.
Innovation Solution
Engineering transgenic animals with a functional immunoglobulin light chain gene and operably linked pseudogenes that repair mutations through gene conversion, ensuring the light chain variable region remains consistent across antibodies.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If somatic hyper-mutation is allowed to occur during affinity maturation in B cells, then antibody affinity and diversity are improved, but light chain diversity increases making it difficult to produce antibodies with the same light chain
Solution Approach 1:
The patent introduces a common light chain that acts as an intermediary component, separated from the heavy chain antigen-binding function. This common light chain is engineered to be shared across multiple antibody specificities, while the heavy chains provide the diverse antigen recognition. The light chain is designed with a framework that supports multiple heavy chain variable regions without undergoing somatic hyper-mutation, thus maintaining sequence consistency while enabling antibody diversity through the heavy chains.
Solution Approach 2:
The patent segments the antibody structure into distinct functional components: a common light chain that remains invariant and provides structural support, and multiple heavy chains that undergo diversification for antigen binding. This segmentation allows the light chain to be excluded from somatic hyper-mutation processes that occur in heavy chains, thereby maintaining light chain sequence consistency while achieving the desired antibody diversity and affinity through heavy chain variation.
2Stability of the object's composition
If transgenic animals are engineered with a single light chain sequence at the endogenous light chain locus, then light chain uniformity is improved, but somatic hyper-mutation still diversifies the light chain during affinity maturation
Solution Approach 1:
The patent designs the common light chain as an intermediary element that is intentionally excluded from the somatic hyper-mutation process. The light chain locus is engineered to contain a fixed, non-mutating sequence that serves as a stable platform for heavy chain diversification. This is achieved by placing the light chain under the control of regulatory elements that prevent recombination and mutation, while allowing heavy chains to undergo normal affinity maturation processes.
Solution Approach 2:
The patent inverts the conventional approach by making the light chain the invariant component rather than the heavy chain. Traditionally, the heavy chain is the stable framework and the light chain can vary; this patent reverses that paradigm by creating a system where the light chain is the common, non-mutating element and the heavy chains are the diversified, antigen-specific components. This inversion allows light chain uniformity to be maintained while still achieving antibody affinity maturation through heavy chain variation.
3Adaptability or versatility
If pseudogenes are used for gene conversion to diversify antibodies, then antibody diversity is improved, but light chain diversity increases rather than remaining common
Solution Approach 1:
The patent applies local quality by allowing gene conversion and diversification to occur only in the heavy chain locus while maintaining a fixed, non-diversified light chain locus. The light chain region is engineered with specific properties (lack of pseudogenes, fixed sequence) that prevent gene conversion, whereas the heavy chain region retains normal diversification mechanisms. This localized differentiation enables antibody diversity through heavy chain gene conversion while preserving light chain sequence consistency.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The transgenic animals produce polyclonal antisera with reduced light chain diversity, enabling the production of bispecific antibodies with the same light chain, enhancing antibody specificity and tolerance for human administration.
Implementation Method 1
the pseudogenes should repair many of the mutations by gene conversion, thereby restoring the coding sequence for the variable region back to its original form
Data Source
AI summary
This disclosure provides, among other things, strategies for minimizing antibody diversification in a transgenic animal that uses gene conversion for antibody diversification. In some embodiments, the animal may comprise a genome comprising an endogenous immunoglobulin light chain locus comprising: (a) a functional immunoglobulin light chain gene comprising a nucleic acid encoding a light chain variable region; and (b) a plurality of pseudogenes that are operably linked to the functional immunoglobulin light chain gene and that donate, by gene conversion, nucleotide sequence to the nucleic acid encoding a light chain variable region, wherein the pseudogenes are upstream or downstream of the functional immunoglobulin light chain gene and encode the same amino acid sequence as the light chain variable region of the functional immunoglobulin light chain gene of (a). In other embodiments, the locus may have a tandem array of coding sequences for the light chain.