Small G-CSFR Binding Proteins for Thermostable Receptor Inhibition
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Solution Overview
Problem
Current G-CSFR binders, such as antibodies, are large in size, costly, require cold storage, and have low thermostability, limiting their therapeutic applicability due to high production costs and reduced tumor penetration.
Innovation Solution
Development of non-immunoglobulin-derived polypeptides and proteins that specifically bind to G-CSFR, with a size of ≤ 50 kDa, enhanced thermostability, and improved tumor penetration capabilities, allowing for easier production and storage.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If antibodies are used as G-CSFR binders, then binding affinity and specificity are achieved, but molecular size is large (150 kDa) resulting in high production costs and reduced tumor penetration
Solution Approach 1:
The patent segments the antibody structure into a smaller functional binding unit by using single-domain antibodies (nanobodies) that contain only the variable domain (VH) responsible for antigen binding. This segmentation reduces molecular size from 150 kDa to approximately 15-20 kDa while preserving binding affinity and specificity for G-CSFR
Solution Approach 2:
The patent extracts only the essential binding function from the complete antibody structure by isolating the variable heavy domain (VH) that contains the complementarity-determining regions (CDRs) responsible for G-CSFR recognition. This extraction eliminates the constant regions and other antibody components that contribute to molecular weight but are not essential for binding
2Reliability
If antibodies are used as G-CSFR binders, then binding functionality is achieved, but thermostability is low and cold chain storage is required
Solution Approach 1:
The patent applies local quality changes by introducing stabilizing mutations specifically in the VH domain structure, optimizing disulfide bridge positions, and enhancing the framework regions to increase thermal stability. These localized modifications improve thermostability without affecting the CDR regions responsible for binding functionality
Solution Approach 2:
The patent creates a composite molecular structure by fusing the VH domain with stable protein scaffolds or linkers that enhance overall thermostability. This composite approach combines the binding functionality of the VH domain with the structural stability of engineered protein frameworks
3Reliability
If antibodies are used as G-CSFR binders, then binding capability is achieved, but production costs are high due to molecular size and complex manufacturing requirements
Solution Approach 1:
The patent adopts a disposable single-use approach with nanobody formulations that eliminate the need for complex manufacturing infrastructure. The small molecular size and simplified structure allow for cost-effective production in simpler bioreactor systems without requiring the extensive purification and formulation processes needed for full-size antibodies
4Reliability
If antibodies are used as G-CSFR binders, then receptor binding is achieved, but tumor penetration is reduced due to large molecular size
Solution Approach 1:
The patent segments the antibody into a compact nanobody format that can navigate through the extracellular matrix and tumor vasculature more effectively. The reduced molecular dimensions (15-20 kDa versus 150 kDa) enable better tissue penetration while the engineered CDR regions maintain high-affinity binding to G-CSFR on tumor cells
Data Source
Figure 1A~1B
Figure 2
Figure 3A~3I
AI summary
The present invention relates to a polypeptide configured to assemble into a binder of granulocyte-colony stimulating factor receptor (G-CSFR), a protein comprising said polypeptide, a nucleic acid molecule encoding said polypeptide or protein, an expression vector comprising the nucleic acid molecule, a recombinant host cell comprising said polypeptide, protein, nucleic acid molecule and/or expression vector, a pharmaceutical composition comprising the said polypeptide, protein, nucleic acid molecule, expression vector and/or host cell, and to a kit.