Mutant IL-2 Protein Engineering for Selective Receptor Binding
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Solution Overview
Problem
Existing IL-2 proteins face challenges in terms of production and purification efficiency, as well as receptor selectivity, leading to unwanted immune downregulation and toxicity in immunotherapy.
Innovation Solution
A novel IL-2 mutant protein with specific mutations in the glycosylation motif and B′C′ loop region to enhance expression, reduce IL-2Rα binding, and increase IL-2Rβ affinity, thereby improving druggability and receptor selectivity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If wild-type IL-2 protein is used, then it can bind to IL-2Rα receptor with high affinity, but it causes unwanted immune downregulation and toxicity
Solution Approach 1:
The patent introduces specific point mutations at localized positions (N31, K70, N87) in the IL-2 protein structure to alter its binding properties. These localized changes modify the interaction interface with IL-2Rα while preserving overall protein function, achieving selective reduction of harmful binding without compromising beneficial immunostimulation
Solution Approach 2:
The patent changes the biochemical parameters of IL-2 by introducing mutations that alter glycosylation motifs and protein conformation. The N31 and N87 mutations create new N-glycosylation sites that change the protein's physical-chemical properties and receptor interaction characteristics, thereby reducing affinity for IL-2Rα while maintaining stability and immunostimulatory activity
2Reliability
If wild-type IL-2 protein is used, then it has natural biological activity, but it shows poor expression and purification efficiency
Solution Approach 1:
The patent modifies expression parameters by introducing mutations that affect protein solubility, stability, and post-translational modification. The N31 and N87 mutations alter glycosylation patterns that improve protein folding and solubility in mammalian expression systems, while the K70 mutation enhances purification efficiency by modifying charge properties for better chromatographic behavior
Solution Approach 2:
The patent introduces N-glycosylation motifs as intermediary structures that mediate between protein synthesis and functional activity. These glycosylation sites act as intermediaries that improve protein folding, stability, and solubility during expression, while also serving as handles for purification through lectin affinity chromatography
3Strength
If IL-2 binds to IL-2Rα receptor, then high affinity binding occurs, but CD25+ cell activation increases leading to immune suppression
Solution Approach 1:
The patent applies local quality modification by introducing mutations specifically at the IL-2Rα binding interface (N31, K70, N87 positions). These localized changes selectively reduce affinity for IL-2Rα-containing receptors on CD25+ cells, while preserving binding to IL-2Rβγ on CD25- effector cells, thereby achieving cell-type selective immunostimulation
Solution Approach 2:
The patent inverts the normal binding pattern by reducing affinity for the high-affinity IL-2Rαβγ trimeric receptor while maintaining or enhancing affinity for the intermediate-affinity IL-2Rβγ dimeric receptor. This inversion of binding preferences reverses the selectivity from CD25+ to CD25- cell activation, achieving the desired therapeutic effect
Data Source
AI summary
The present invention relates to a novel interleukin-2 (IL-2) mutant protein. The present invention further provides a fusion protein and an immunoconjugate comprising the IL-2 mutant protein, a nucleic acid encoding the IL-2 mutant protein, and a vector and a host cell comprising the nucleic acid. The present invention further provides a method for preparing the IL-2 mutant protein, a pharmaceutical composition comprising the IL-2 mutant protein, and therapeutic use of the mutant protein.