Recombinant PMHC Molecules with Engineered Cavities
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Solution Overview
Problem
Current methods for producing soluble peptide-major histocompatibility complex class II (pMHCII) monomers face challenges such as low yields, instability, and the need for foreign affinity tags, which hinder their use in therapeutic applications due to immunogenicity and purification difficulties.
Innovation Solution
The development of peptide-tethered and non-peptide-tethered MHC Class I and Class II monomers with high stability and receptor-signaling activity, using engineered protuberances and cavities for heterodimerization, allowing for high-yield production and purification without affinity tags, and their conjugation to nanoparticles for therapeutic use.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Stability of the object's composition
If leucine zipper domains are used for heterodimerization of pMHCII molecules, then solubility is improved, but production yield remains low and foreign tags are required
Solution Approach 1:
The patent changes the chemical structure of the heterodimerization domain from leucine zipper to IgG Fc domain, fundamentally altering the molecular parameters to achieve both high solubility and high production yield without foreign tags
Solution Approach 2:
The patent removes the need for foreign affinity tags by using the endogenous IgG Fc domain for both heterodimerization and purification purposes, extracting the dual-function requirement into a single native domain
2Ease of manufacture
If foreign affinity tags are used for purification of pMHCII molecules, then purification efficiency is improved, but immunogenicity increases and therapeutic use is precluded
Solution Approach 1:
The patent enables the pMHCII molecule to purify itself through the Protein A binding property of the endogenous IgG Fc domain, eliminating the need for foreign affinity tags and thereby removing the source of immunogenicity
Solution Approach 2:
The IgG Fc domain serves multiple functions simultaneously: heterodimerization, solubility enhancement, and purification via Protein A binding, eliminating the need for separate foreign tags
3Device complexity
If pMHCII molecules are produced without heterodimerization domains, then simplicity is improved, but stability and receptor-signaling activity are reduced
Solution Approach 1:
The patent merges the heterodimerization function with the endogenous IgG Fc domain structure, combining stability enhancement with the native MHCII molecule rather than adding separate destabilizing elements
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 approach results in stable, high-yield production of pMHC monomers that can be efficiently purified and used therapeutically, enhancing receptor-signaling potency and enabling the detection and regulation of autoantigen-specific T-cells, effectively addressing organ-specific autoimmunity.
Implementation Method 1
the interface of the first polypeptide comprises an engineered protuberance which is positionable in an engineered cavity in the interface of the second polypeptide
Implementation Method 2
the disease-relevant antigen is covalently connected to the MHC class II α1 domain or the MHC class II β1 domain by a disulfide bond formed between a cysteine amino acid associated with the antigenic peptide and a cysteine amino acid of the MHC class II α1 domain or the MHC class II β1 domain
Data Source
AI summary
Provided are peptide-MHC class I and class II molecules having improved stability and high potency, and that can be produced in high yield. Also provided are receptor-signaling nanoparticles comprising the improved peptide-MHC molecules.