Thermostable ISG65 Mutant for Complement Dysregulation
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Solution Overview
Problem
Current treatments for complement dysregulation disorders, such as atypical hemolytic uremic syndrome and autoimmune diseases, are costly and require targeting downstream effector pathways, limiting their comprehensive approach and potency.
Innovation Solution
A thermostable variant of the ISG65 protein with increased melting temperature, derived from the extracellular domain of Trypanosoma brucei gambiense, is developed, which binds to central components of the complement cascade like C3b, offering a more comprehensive and potent treatment by targeting multiple interaction points within the cascade.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If Eculizumab (monoclonal antibody targeting C5) is used to treat complement dysregulation disorders, then treatment effectiveness is achieved, but cost becomes prohibitively high
Solution Approach 1:
The patent creates a simplified copy of the natural ISG65 protein's extracellular domain, which naturally binds to multiple complement components (C3, C3b, C3(H2O), C3d). This copied functional domain provides comprehensive complement inhibition at a fraction of the cost of full monoclonal antibodies like Eculizumab, while maintaining treatment effectiveness through multi-target binding
Solution Approach 2:
The patent extracts only the essential extracellular domain of ISG65 that contains the complement-binding functionality, removing unnecessary portions of the native protein. This extracted domain retains the ability to bind multiple complement components while being more cost-effective to produce than full-length monoclonal antibodies
2Reliability
If downstream effector pathways (C5 or C9 polymerization) are targeted, then treatment is provided, but comprehensive approach and potency are limited
Solution Approach 1:
The ISG65 extracellular domain exhibits universal binding capability to multiple complement components (C3, C3b, C3(H2O), C3d) simultaneously. This multi-functional binding allows a single protein to intervene at multiple points in the complement cascade, providing comprehensive coverage without requiring multiple separate therapeutic agents
Solution Approach 2:
The ISG65 domain binds to C3 and its activation products early in the complement cascade, preventing downstream activation before it occurs. By acting upstream at the C3 level rather than downstream at C5 or C9, the protein comprehensively blocks multiple pathways before they can proceed to effector functions
3Reliability
If native ISG65 protein is used for treatment, then complement dysregulation can be addressed, but stability for production and storage is insufficient
Solution Approach 1:
The patent systematically modifies amino acid parameters of the ISG65 extracellular domain through site-directed mutagenesis, changing specific residues to improve thermostability. These parameter changes in the protein sequence maintain complement-binding functionality while significantly enhancing stability for production and storage
Solution Approach 2:
The patent applies localized modifications at specific positions within the ISG65 sequence rather than global changes. By targeting specific residues for mutation while preserving the overall structure and key binding interfaces, the protein maintains its therapeutic function while gaining enhanced thermostability in critical regions
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 thermostable ISG65 mutant significantly decreases hemolysis in a concentration-dependent manner, providing a cost-effective and stable protein for treating complement dysregulation disorders with improved stability and broader intervention points within the complement cascade.
Implementation Method 1
The extracellular domain of ISG65 binds to native C3, to its reactive activation products C3b and C3(H2O), and to C3d
Implementation Method 2
The new protein carries several mutations and shows a significant and unexpected increase in the melting temperature (ISG65 M2 by 14.7 °C), compared to the native ISG65 sequence
Data Source
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AI summary
The present invention relates to a thermostable ISG65 mutant derived from the extracellular domain of the surface protein ISG65 (invariant surface glycoprotein 65(Tbg.972.2.1600)) of the human parasite Trypanosoma brucei gambiense. The thermostable ISG65 mutant shows enhanced thermostability in comparison with the wild type extracellular domain of ISG65 and is useful in the treatment of complement dysregulation diseases.