Self-inactivating AAV Vectors for Gene Editing Safety
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current gene editing technologies, such as those using adeno-associated viral (AAV) vectors, face challenges with long-term expression specificity and safety concerns, particularly in post-mitotic cells, due to the persistent presence of CRISPR nucleases, which can lead to immunogenicity and off-target effects.
Innovation Solution
Development of self-inactivating recombinant vectors (SIRV) and self-inactivating adeno-associated virus (siAAV) vectors that temporally control the expression of CRISPR components by incorporating self-editing segments, reducing or eliminating CRISPR expression post-target editing, thereby enhancing safety and specificity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If AAV vectors are used to deliver CRISPR nucleases for long-term gene editing, then editing efficiency is improved, but immunogenicity and off-target effects increase due to persistent nuclease expression
Solution Approach 1:
The patent changes the temporal expression parameter of CRISPR components by using self-inactivating AAV vectors that reduce nuclease expression after a defined period. This is achieved through self-targeting gRNAs that direct Cas9 to cleave and inactivate the transgene following editing, thereby maintaining high initial editing efficiency while reducing long-term immunogenicity and off-target effects
Solution Approach 2:
The patent extracts the harmful persistent expression feature from the AAV vector system by incorporating self-inactivation mechanisms. The self-targeting gRNA system selectively removes the CRISPR nuclease expression capability after completing the editing function, separating the beneficial editing phase from the harmful persistent expression phase
2Reliability
If CRISPR nucleases are continuously expressed in post-mitotic cells, then gene editing capability is maintained, but safety concerns increase due to prolonged presence of editing machinery
Solution Approach 1:
The patent implements periodic action by designing the CRISPR system to be active only during the necessary editing window. The self-inactivating mechanism creates a temporal pattern where nuclease expression is high initially for editing, then automatically reduces after completion, providing reliable editing capability while limiting safety concerns to the necessary time period only
3Object-affected harmful factors
If self-inactivating mechanisms are incorporated into AAV vectors, then safety and specificity are improved, but vector design complexity increases
Solution Approach 1:
The patent applies self-service by designing the AAV vector to automatically inactivate itself after completing the editing function. The self-targeting gRNA system uses the same CRISPR machinery to target and cleave the transgene encoding the nuclease, creating a self-regulating system that improves safety without requiring external control mechanisms or complex regulatory systems
Data Source
AI summary
Provided herein are compositions and methods for use of self-inactivating recombinant vectors (SIRV) encoding Class 2 Type V and guide ribonucleic acid (gRNA) sequences useful for nucleic acid sequence editing, and including self-inactivating components. The SIRV may be delivered to cells as part of an AAV vector to target a gene of interest.