Single Domain Antibody Framework Modifications to Reduce Immunogenicity
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Solution Overview
Problem
Existing single domain antibodies (sdAbs) are often immunogenic due to pre-existing antibodies in humans, leading to clinical challenges and adverse events, despite efforts to humanize or engineer them to reduce immunogenicity, particularly affecting the CDR regions and framework regions.
Innovation Solution
Modifications in the framework regions of sdAbs, such as framework 1, framework 2, framework 3, and framework 4, including mutations, truncations, and extensions, are introduced to prevent recognition by pre-existing anti-single domain antibody (ASDA) without compromising binding affinity, specificity, or stability, using specific amino acid changes like Leu11Lys (L11K), Leu11Arg (L11R), Leu11Glu (L11E), Ala88Glu (A88E), and carboxy-terminal extensions or truncations.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If single domain antibodies are used, then binding affinity and specificity are improved, but immunogenicity increases due to pre-existing antibodies
Solution Approach 1:
The patent applies local quality by making specific amino acid substitutions at particular framework region positions (such as positions 11, 44, 45, 46, 88, 112, 113) while leaving the CDR regions unchanged. This targeted modification approach reduces immunogenicity locally without affecting the overall binding function of the antibody.
Solution Approach 2:
The patent changes the amino acid sequence parameters in the framework regions by introducing specific mutations (e.g., L11K, A88E, and combinations thereof). These parameter changes alter the epitopic landscape to reduce recognition by pre-existing human antibodies while preserving the antigen-binding capability.
2Object-affected harmful factors
If framework region modifications are introduced, then ASDA recognition is reduced, but structural stability may be compromised
Solution Approach 1:
The patent applies local quality by selecting specific framework region positions for modification (positions 11, 44, 45, 46, 88, 112, 113) that are less critical for structural stability. This allows reduction of ASDA recognition while preserving the overall structural integrity and functional stability of the sdAb.
Solution Approach 2:
The patent introduces specific amino acid substitutions (L11K, L11R, L11E, A88E, and combinations) that change local chemical properties to reduce immunogenicity. These parameter changes are carefully selected to maintain structural stability while achieving the desired reduction in ASDA recognition.
3Object-affected harmful factors
If multiple framework region modifications are combined, then immunogenicity is further reduced, but complexity of engineering increases
Solution Approach 1:
The patent applies local quality by identifying and modifying specific key positions in the framework regions. By focusing on particular positions (11, 44, 45, 46, 88, 112, 113) rather than进行全面 modification, the patent reduces immunogenicity effectively while keeping the engineering complexity manageable.
Solution Approach 2:
The patent introduces specific amino acid substitutions at defined positions with predictable effects on immunogenicity. This systematic approach to parameter changes allows for controlled reduction of immunogenicity through combinatorial mutations without requiring complex engineering processes.
Data Source
AI summary
The invention provides modifications within human or humanized single domain antibody fragments (sdAbs) that prevent recognition by pre-existing antibodies, to isolated polypeptides that include these modifications, and to methods and uses thereof.