Monomeric CCN Fusion Proteins for Proteolytic Stability
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Solution Overview
Problem
Recombinant CCN proteins, particularly full-length CCN proteins, are susceptible to proteolytic degradation, making them unsuitable as therapeutic agents due to instability during manufacturing and unpredictable pharmacokinetics, and existing fusion proteins with CCN proteins face similar issues regarding proteolytic susceptibility and unpredictable activity.
Innovation Solution
Development of monomeric fusion proteins comprising the thrombospondin type 1 repeat homology domain (TSP-1) of CCN family proteins, fused with a monomeric fusion partner and optionally a peptide linker, which are resistant to proteolytic degradation, thereby maintaining biological activity and stability.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If full-length CCN proteins are used as therapeutic agents, then biological activity is achieved, but proteolytic degradation susceptibility increases making them unstable
Solution Approach 1:
The invention segments the full-length CCN protein into individual functional domains (Domain I, II, III, or IV) that can be expressed and used independently. This segmentation allows selection of only the necessary functional domain(s) without the vulnerable full-length structure, maintaining biological activity while reducing proteolytic susceptibility by eliminating unnecessary protein regions that serve as protease targets.
Solution Approach 2:
The invention extracts specific functional domains from the full-length CCN protein structure. By isolating and expressing only the essential domain(s) responsible for the desired therapeutic effect, the invention removes the vulnerable portions of the protein that are prone to proteolytic degradation, thereby achieving both biological activity and improved stability.
2Stability of the object's composition
If fusion proteins with CCN proteins are created, then stability is improved, but proteolytic susceptibility remains unpredictable
Solution Approach 1:
The invention applies segmentation by using already-isolated CCN domains as the basis for fusion proteins rather than full-length CCN proteins. This segmental approach creates fusion proteins with predictable proteolytic resistance because the smaller, defined domain structures have fewer vulnerable sites and more predictable behavior when fused to stable partner proteins like Fc fragments.
Solution Approach 2:
The invention creates composite fusion proteins by combining isolated CCN domains with stable fusion partners such as Fc fragments. This composite approach leverages the stability of the Fc portion while incorporating only the necessary functional CCN domain, resulting in fusion proteins with improved overall stability and predictable proteolytic resistance characteristics.
3Stability of the object's composition
If truncated CCN domain proteins are used, then proteolytic stability is improved, but manufacturing complexity increases
Solution Approach 1:
The invention facilitates segmentation by providing explicitly defined domain boundaries and sequences for CCN domains. This clear delineation of domain structures (with specific amino acid sequences and lengths) simplifies the manufacturing process by enabling precise genetic construct design, straightforward expression system selection, and simplified purification protocols based on known domain characteristics.
Data Source
AI summary
The present invention relates to recombinant proteins having an amino acid sequence corresponding to or related to the thrombospondin type 1 repeat homology domain of a member of the CCN family proteins and the use thereof. Furthermore, the present invention relates to fusion proteins comprising an amino acid sequence corresponding to or related to the thrombospondin type 1 repeat homology domain of a member of the CCN family proteins combined with a fusion partner and optionally a linker region. Also, novel protease resistant Fc-fragments are disclosed herein.