Single-Chain Insulin Analogues via Side Chain Covalent Link
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Solution Overview
Problem
Current insulin analogues have low affinity to the insulin receptor due to blocked essential residues, leading to reduced therapeutic efficacy and stability issues such as fibrillation and precipitation.
Innovation Solution
Development of single-chain insulin analogues where the A-chain and B-chain are connected through the functional group of an amino acid side chain of the B-chain, forming an additional direct covalent link, which maintains essential terminal groups free for biological activity and reduces chain flexibility.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of manufacture
If the A-chain and B-chain are connected through a connecting peptide (as in proinsulin and known single-chain analogues), then the insulin analogue can be produced as a single chain, but the essential terminal residues become blocked leading to reduced affinity for the insulin receptor
Solution Approach 1:
The connecting peptide is segmented and removed, leaving only the essential disulfide bonds to connect the A-chain and B-chain. This segmentation allows the terminal residues to remain free while maintaining single-chain structure through the disulfide bridges alone, resolving the contradiction between single-chain production and receptor affinity.
Solution Approach 2:
The connecting peptide is extracted and removed from the single-chain structure. By taking out the connecting peptide that was blocking the terminal residues, the invention maintains the single-chain configuration through disulfide bonds only, thereby restoring full receptor affinity while preserving ease of single-chain production.
2Reliability
If the connecting peptide is shortened or modified to improve receptor affinity, then the insulin activity increases, but the stability and solubility properties deteriorate
Solution Approach 1:
The disulfide bonds are positioned at specific locations (CysA7-CysB7 and CysA20-CysB19) to provide local structural stability and solubility while allowing the terminal residues to remain free for optimal receptor interaction. This local quality assignment resolves the contradiction between insulin activity and stability.
Solution Approach 2:
The invention changes the structural parameters by eliminating the connecting peptide and relying solely on disulfide bonds for chain connection. This parameter change optimizes both the stability/solubility properties and the insulin activity by allowing terminal residue freedom while maintaining structural integrity.
3Ease of manufacture
If conventional single-chain insulin analogues are used with modified C-peptide, then they exhibit some insulin activity, but the activity is significantly reduced (up to 42% of native insulin)
Solution Approach 1:
The connecting peptide is completely extracted and removed from the structure. This extraction eliminates the steric hindrance and blocking effects of the connecting peptide, allowing the terminal residues to freely interact with the insulin receptor and restore full insulin activity while maintaining the single-chain configuration through disulfide bonds.
Solution Approach 2:
The invention creates a composite structure where the A-chain and B-chain are connected solely through disulfide bonds without any additional connecting peptide. This composite approach combines the stability of disulfide-linked chains with the full bioactivity of native insulin terminal residues, achieving both ease of single-chain manufacture and high insulin activity.
Data Source
AI summary
A first aspect of the invention relates to a single chain insulin analogue comprising: (A) the A-chain of human or animal insulin, or an analogue or derivative thereof; (B) the B-chain of human or animal insulin, or an analogue or derivative thereof; (C) one or more disulfide bonds between said A-chain and said B-chain; and (D) a further covalent link, L, between a functional group of an amino acid in the A-chain and a functional group of an amino acid in the B-chain, at least one of said functional groups being an amino acid side chain functional group. Further aspects of the invention relate to pharmaceutical compositions comprising said single chain insulin derivatives, and therapeutic uses thereof.