AAV Capsid Tropism Redirection via Cell-Type-Specific RNA Screening
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Solution Overview
Problem
Current methods for improving the transduction efficiency of adeno-associated virus (AAV) capsids in the central nervous system (CNS) are limited by the need for transgenic animals and biased library distribution, making it difficult to achieve targeted gene delivery in complex tissues like the CNS.
Innovation Solution
The TRACER platform uses cell-type-specific promoters to drive AAV capsid mRNA expression, allowing for the generation and selection of capsid variants with enhanced tropism through RNA-driven screening in non-transgenic animals, enabling the identification of capsids with high specificity for neuronal, astrocyte, or other cell types without the need for transgenic animals or helper virus co-infection.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If transgenic animals and helper virus co-infection are used to improve AAV capsid tropism, then capsid variants with enhanced transduction can be obtained, but the complexity of the experimental system increases and the method becomes less broadly applicable
Solution Approach 1:
The invention extracts and eliminates the requirement for transgenic animals and helper virus co-infection from the AAV capsid evolution system. By using a standard AAV production system without these additional components, the method simplifies the experimental setup while maintaining the ability to select for capsid variants with enhanced tropism through RNA-driven screening.
Solution Approach 2:
The invention creates a universally applicable platform for AAV capsid evolution that can be used across different cell types and tissue targets without requiring specific transgenic models or helper viruses. The cell-type-specific promoter-driven RNA screening approach provides a multi-functional system that works for neuronal, astrocyte, and other cell type targeting.
2Manufacturing precision
If cell-type-specific library selection is applied to improve CNS tropism, then capsid variants with enhanced specificity can be recovered, but the requirement for transgenic CRE mice limits applicability to other species and large animal studies
Solution Approach 1:
The invention removes the dependency on transgenic CRE mice from the cell-type-specific selection process. By using cell-type-specific promoters to drive AAV capsid mRNA expression directly in the target cells, the method achieves cell-type-specific library selection without requiring transgenic animals, thereby enabling application across multiple species including large animals and non-human primates.
Solution Approach 2:
The invention introduces cell-type-specific promoters as intermediaries between the AAV capsid expression system and the target cell types. These promoters act as mediators that enable selective expression and recovery of capsid variants in specific cell types (neuronal, astrocyte, etc.) without requiring transgenic modification of the host animals.
3Ease of manufacture
If standard AAV production systems are used, then the process is simple and broadly applicable, but capsid variants cannot be selected with high cell-type specificity in complex tissues
Solution Approach 1:
The invention introduces dynamic, cell-type-specific promoter-driven expression into the standard AAV production system. This dynamic control allows the system to adapt to different cell type requirements during the selection process, enabling high tropism specificity while maintaining the simplicity and broad applicability of standard AAV production methods.
Solution Approach 2:
The invention changes the expression parameter of AAV capsid mRNA by controlling it with cell-type-specific promoters. This parameter change enables the standard AAV production system to achieve cell-type-specific selection of capsid variants, thereby maintaining production simplicity while achieving high tropism specificity for neuronal, astrocyte, and other cell types.
Data Source
AI summary
The disclosure relates to compositions, methods, and processes for the preparation, use, and/or formulation of adeno-associated virus capsid proteins, wherein the capsid proteins comprise targeting peptide inserts for enhanced tropism to a target tissue.