FVIII Variant Engineering for Lower Immunogenicity and Gene Delivery
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Solution Overview
Problem
Current hemophilia A treatments, including protein replacement therapy and gene therapy, are costly, inconvenient, and face challenges from immune responses and toxic side effects due to high-dosage vectors, necessitating improved FVIII variants with reduced immunogenicity and increased expression/stability for effective gene therapy.
Innovation Solution
Development of FVIII variants with mutations such as N2118Q, BDD deletion, and other specific mutations to enhance secretion, expression, stability, and functional activity, along with optimized expression cassettes for targeted delivery, particularly to liver sinusoidal endothelial cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If high-dosage vectors are used for gene therapy, then therapeutic effect is improved, but immune responses and toxic side effects increase
Solution Approach 1:
The patent applies parameter changes by modifying the FVIII protein structure through specific mutations (N2118Q, BDD deletion, and other mutations) to reduce immunogenicity. These structural parameter changes allow the protein to elicit fewer immune responses while maintaining therapeutic function, thereby enabling effective gene therapy at lower vector doses.
2Reliability
If native FVIII is used for protein replacement therapy, then coagulation function is provided, but immunogenicity and treatment cost are high
Solution Approach 1:
The patent modifies the FVIII protein parameters through specific mutations including N2118Q substitution and BDD domain deletion. These changes reduce the protein's immunogenicity while preserving its coagulation function, making it suitable for repeated administration in protein replacement therapy without triggering strong immune responses.
Solution Approach 2:
The patent extracts and removes the B domain (BDD deletion) from the native FVIII protein structure. This extraction reduces the overall size and immunogenicity of the protein while maintaining the essential coagulation activity provided by the remaining domains, thereby reducing immune responses.
3Reliability
If full-length FVIII is packaged into vectors, then complete protein function is achieved, but vector packaging efficiency decreases
Solution Approach 1:
The patent removes the B domain from the full-length FVIII protein through BDD deletion, creating a truncated version that is smaller in size. This extracted form fits more efficiently into viral vectors, improving packaging efficiency and enabling higher expression levels while retaining sufficient coagulation function.
Solution Approach 2:
The patent applies local quality changes by selectively deleting only the B domain while preserving the essential A and C domains that contain the critical coagulation activity. This localized modification maintains necessary protein function while reducing overall size for improved vector packaging.
Data Source
AI summary
Variants of coagulation factor VIII (FVIII) and expression cassettes encoding the FVIII variants thereof are described. A variant FVIII includes a glycoepitope of the FVIII protein including an N2118Q mutation. The N2118Q mutation can be combined with other mutations including a BDD-FVIII, N6, V3, RH, furin-cleavage site deletion. X10, K12, and/or F309S mutation to form additional FVIII variants. The FVIII variants with the N2118Q mutation and expression cassettes thereof can result in reduced immunogenicity of the resulting protein. When combined with other FVIII mutations, higher gene expression, increased secretion, increased stability, and higher FVIII functional activity can be achieved by the expressed FVIII variants. The variant FVIII and expression cassettes described here can be useful in protein replacement therapy and/or gene therapy for the treatment of hemophilia A.