Covalent Arginine Deiminase Dimer for Stability and Low Immunogenicity
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Solution Overview
Problem
Current arginine deiminase enzymes face challenges with poor stability, short in vivo half-life, and high immunogenicity, limiting their effectiveness in arginine deprivation therapy for cancer treatment.
Innovation Solution
A covalent dimeric arginine deiminase mutant is created through site-directed mutagenesis to form inter-subunit covalent disulfide bonds, combined with PEGylation to enhance structural stability and reduce immunogenicity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If arginine deiminase is used for arginine deprivation therapy, then cancer cells can be targeted effectively, but the enzyme exhibits poor stability and short in vivo half-life
Solution Approach 1:
The patent creates a covalent dimeric structure by introducing inter-subunit disulfide bonds, combining multiple enzyme subunits into a stable composite entity. This dimeric structure provides enhanced stability and prolonged in vivo half-life while maintaining the arginine deprivation therapeutic effect on cancer cells
2Reliability
If arginine deiminase is used to deplete arginine in tumors, then cancer cell growth is inhibited, but the enzyme exhibits high immunogenicity
Solution Approach 1:
The patent employs PEGylation technology, conjugating polyethylene glycol chains to the arginine deiminase enzyme. This creates a composite structure where the PEG layer acts as an immunomodulatory coating, reducing the immune system's recognition and attack on the enzyme while preserving its anti-tumor activity
Solution Approach 2:
The patent introduces PEG as an intermediary substance between the immune system and the arginine deiminase enzyme. The PEG layer serves as a protective mediator that prevents direct interaction between the immune system and the foreign enzyme, thereby reducing immunogenicity and antibody formation
3Reliability
If arginine deiminase is administered therapeutically, then arginine levels are reduced in target tumors, but the enzyme suffers from poor structural stability
Solution Approach 1:
The patent creates a covalent dimeric structure by introducing inter-subunit disulfide bonds, combining multiple enzyme subunits into a stable composite entity. This dimeric structure provides enhanced stability and prolonged in vivo half-life while maintaining the arginine deprivation therapeutic effect on cancer cells
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 mutant maintains enzymatic activity, reduces immunogenicity, and prolongs in vivo half-life, effectively depleting arginine levels in tumors like hepatocellular carcinoma and melanoma.
Implementation Method 1
an arginine deiminase mutant is provided. The mutant includes an inter-subunit covalent disulfide bond formed between arginine deiminase subunits via mutation at an amino acid site on the arginine deiminase
Implementation Method 2
Arginine deiminase (ADI, EC 3.5.3.6) is another microbial enzyme capable of degrading arginine into citrulline and ammonia
Implementation Method 3
Covalent conjugation of PEG and arginine deiminase facilitates shielding epitopes on the surface, reducing the immunogenicity, and prolonging the in vivo half-life
Data Source
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AI summary
Provided are an arginine deiminase mutant, a covalent dimer and a conjugate and a use thereof. The provided arginine deiminase mutant forms a covalent disulfide bond between arginine deiminase subunits by means of mutation of an amino acid site on arginine deiminase. Also provided are a covalent conjugate and a use in the field of cancer treatment. Site-directed mutagenesis of an amino acid site promotes formation of a covalent disulfide bond between non-covalent homodimeric arginine deiminase subunits, and a formed dimer has a more stable structure; in addition, the exposure level of an epitope on an interaction interface can be reduced, the in vivo immunogenicity is reduced, and a more excellent anti-tumor effect is exhibited.