XTEN Binding Fusion Proteins for Stable Bacterial Production
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Solution Overview
Problem
Current methods for producing antibodies and antibody fragments in bacterial hosts often result in the formation of inclusion bodies, leading to complexity and increased costs, and are associated with insufficient stability and production yields.
Innovation Solution
Development of novel binding fusion proteins comprising extended recombinant polypeptides (XTEN) linked to targeting moieties with specific binding affinity, which exhibit enhanced pharmacokinetic properties such as prolonged terminal half-life and improved production yields.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If antibodies or antibody fragments are expressed in bacterial hosts, then production can be achieved, but inclusion bodies form in the cytoplasm leading to complexity and increased costs
Solution Approach 1:
The patent changes the expression conditions by using periplasmic expression instead of cytoplasmic expression, and by optimizing culture conditions (aerobic vs anaerobic, temperature, pH) to prevent inclusion body formation and improve soluble protein yield
Solution Approach 2:
The patent uses the periplasm as an intermediary compartment between the cytoplasm and the extracellular environment, providing a protected space for protein folding and assembly that prevents aggregation while maintaining production efficiency
2Reliability
If scFv or Fab fragments are produced in host cells, then antigen binding capability is achieved, but stability and production yields are insufficient
Solution Approach 1:
The patent optimizes multiple parameters including temperature (37°C vs lower temperatures), pH (7.0-7.5 range), aeration, and nutrient composition to enhance both the stability of the protein fragments and the production yield simultaneously
Solution Approach 2:
The patent incorporates chaperone proteins and folding assistants in the expression system to ensure proper protein folding and stability before the fragments are produced, preventing misfolding and aggregation that would reduce both stability and yield
3Ease of manufacture
If periplasmic extraction is used, then some protein can be recovered, but the extraction procedure is not as robust as cytoplasmic extraction contributing to low yields
Solution Approach 1:
The patent converts the previously problematic periplasmic extraction difficulty into an advantage by optimizing the extraction protocol specifically for periplasmic proteins, using enzyme treatments and pH adjustments that selectively solubilize periplasmic proteins while leaving cytoplasmic contents behind, thereby achieving high purity and acceptable yield
Data Source
AI summary
The present invention relates to binding fusion protein compositions comprising targeting moieties linked to extended recombinant polypeptide (XTEN), binding fusion protein-drug conjugate compositions, and XTEN-drug conjugate compositions, isolated nucleic acids encoding the compositions and vectors and host cells containing the same, and methods of using such compositions in treatment of diseases, disorders, and conditions.