Humanized Anti-CLDN6 Antibodies With Back Mutations for Affinity Recovery
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Solution Overview
Problem
Current methods for humanizing murine antibodies to target CLDN6 suffer from reduced affinity and stability, and there is no guaranteed method to achieve high-affinity humanized antibodies.
Innovation Solution
Developed antigen-binding proteins with specific heavy and light chain variable regions, including back mutations at defined sites, to enhance binding activity to CLDN6, and created antibody-drug conjugates using various linkers and toxins for targeted cancer treatment.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Object-affected harmful factors
If murine antibodies are humanized to reduce immunogenicity, then immunogenicity is reduced, but affinity and stability are reduced
Solution Approach 1:
The patent applies local quality by making specific back mutations at defined positions (e.g., position 44 in heavy chain framework region, positions 66, 70, 85 in light chain framework region) rather than uniform modifications throughout the antibody structure. This localized approach restores critical interactions at specific sites while maintaining the overall humanized structure, thereby recovering affinity and stability without increasing immunogenicity.
Solution Approach 2:
The patent systematically varies parameters by testing multiple combinations of back mutations at different framework region positions. By changing amino acid sequences at specific locations and measuring the resulting affinity and stability, the invention identifies optimal parameter combinations that maximize antibody performance while maintaining humanization benefits.
2Reliability
If antibodies are engineered for high selectivity to CLDN6, then selectivity is improved, but development complexity increases
Solution Approach 1:
The patent achieves high selectivity through local quality by focusing mutations and engineering efforts on specific CDR regions that directly contact the antigen, particularly targeting epitopes unique to CLDN6 versus CLDN9. This localized optimization allows differentiation between similar antigens without requiring complex modifications throughout the entire antibody structure.
Solution Approach 2:
The patent applies partial action by concentrating engineering efforts on the variable regions (VH and VL) that are critical for antigen binding, while leaving the constant regions relatively unchanged. This focused approach achieves the necessary selectivity without the complexity of comprehensive antibody engineering.
Data Source
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AI summary
Provided are an antigen-binding protein for hCLDN6 and the use thereof. Said antigen-binding protein comprises a heavy chain variable region. The heavy chain variable region has HCDR1 having an amino acid sequence of X1YTMS; HCDR2 having an amino acid sequence of TISSGGGX2TYYPDSVKG in which X2 = R, N, Q, D or E; and HCDR3 having an amino acid sequence of GDX4RYDX3FAY in which X4 = Y, N or Q, wherein (1) X1 = Y, M, Q, D or E, and X3 = G; or (2) X1 = S or D, and X3 = A. Provided are novel humanized antibodies targeting CLDN6, and different compositions based on these antibodies, which have better cell proliferation inhibitory activity and tumor inhibitory activity, thus providing novel means for cancer diagnosis and treatment.