Engineered CH2 Domain Stability via Framework Mutations
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Solution Overview
Problem
Current CH2 domain molecules lack enhanced stability and solubility, and there is no effective method to alter framework residues to reduce immunogenicity or prevent aggregation.
Innovation Solution
Engineered CH2 domain molecules with specific amino acid substitutions in framework regions, such as Val to Phe at position 27 and Leu to Met at position 15, along with modifications in loop regions, are developed to improve stability and solubility, while maintaining FcRn binding characteristics.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If CH2 domain molecules are used as isolated nanobodies, then they can bind to targets and distribute in the body, but they exhibit reduced stability and increased immunogenicity compared to full antibodies
Solution Approach 1:
The patent applies parameter changes by systematically modifying amino acid residues at specific positions within the CH2 domain framework regions. Multiple point mutations were introduced to optimize stability, solubility, and reduce aggregation propensity while preserving FcRn binding functionality. This approach directly addresses the contradiction by changing the molecular parameters of the CH2 domain to achieve both stability and binding function.
2Stability of the object's composition
If additional disulfide bonds are added to CH2 molecules to improve stability, then stability increases, but the molecular complexity and potential aggregation increase
Solution Approach 1:
The patent applies local quality by making targeted, localized modifications to specific framework regions of the CH2 domain rather than global structural changes. Point mutations were introduced at specific positions (e.g., framework residues) to locally enhance stability and solubility properties without adding global structural complexity like additional disulfide bonds. This allows stability improvement while maintaining relatively simple molecular architecture.
3Stability of the object's composition
If N-terminal or C-terminal amino acids are deleted to improve stability, then stability improves, but the molecule loses potential functional regions
Solution Approach 1:
The patent applies parameter changes by modifying internal framework residues through point mutations rather than deleting terminal regions. This approach changes the chemical and physical parameters of the CH2 domain (stability, solubility, aggregation propensity) through localized amino acid substitutions that do not compromise the overall functional architecture or potential adaptability of the molecule.
4Stability of the object's composition
If framework residues are modified to enhance stability and reduce immunogenicity, then stability and solubility improve, but the structural integrity may be compromised
Solution Approach 1:
The patent applies local quality by making targeted modifications to specific framework residues that are less critical for overall structural integrity. By selecting specific positions for mutation that maintain the core structural framework while improving local properties (solubility, aggregation resistance, stability), the patent achieves enhanced performance without compromising global structural integrity.
Data Source
AI summary
This invention relates to engineered CH2 domain molecules containing amino acids in the framework regions that confer enhanced stability and/or solubility. In particular, the invention provides engineered CH2 domain molecules containing amino acid residues that differ from a wild type CH2 domain or a template CH2 domain molecule within one or more framework regions and that result in improved stability and/or solubility.