SaCas9 AAV Vector Genome Editing Delivery
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Solution Overview
Problem
Current genome-editing techniques face challenges in efficiently delivering CRISPR-Cas systems for therapeutic genome engineering, particularly in vivo, due to low delivery efficiency and high requirements for viral vectors, as well as inefficiencies in homology-directed repair (HDR) and co-delivery processes.
Innovation Solution
The use of SaCas9 from Staphylococcus aureus, packaged into a single Adeno-associated virus (AAV) vector, to enhance the delivery and targeting specificity of the CRISPR-Cas system, reducing the number of viral vectors needed and improving HDR efficiency.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If CRISPR-Cas system components are packaged into multiple viral vectors for delivery, then the completeness of genome editing components is improved, but the complexity of delivery and number of vectors required increases
Solution Approach 1:
The patent combines multiple CRISPR-Cas system components (Cas9 nuclease, guide RNA, and homology-directed repair template) into a single AAV viral vector. This merging of components that were previously delivered via separate vectors reduces the complexity of the delivery system while maintaining the completeness of all necessary elements for functional genome editing.
2Reliability
If traditional CRISPR-Cas systems are used with larger Cas9 enzymes, then the genome editing capability is achieved, but the packaging efficiency into AAV vectors decreases
Solution Approach 1:
The patent extracts and utilizes a smaller Cas9 enzyme variant that can be efficiently packaged into AAV vectors. By selecting a compact Cas9 enzyme that retains full genome editing functionality, the system achieves effective packaging within the limited AAV capacity while maintaining complete genome editing capability.
3Reliability
If multiple viral vectors are used for co-delivery of CRISPR components, then the functionality of the system is improved, but the delivery efficiency decreases
Solution Approach 1:
The patent merges all essential CRISPR system components into a single AAV vector, eliminating the need for coordinated co-delivery of multiple vectors. This single-vector approach simplifies the delivery process and improves delivery efficiency by avoiding the complexities of simultaneous delivery and coordination of multiple viral vectors.
4Manufacturing precision
If homology-directed repair (HDR) is performed with multiple vectors, then the precision of genome editing is improved, but the complexity of the process increases
Solution Approach 1:
The patent combines the homology-directed repair template with the CRISPR-Cas components in a single AAV vector. This integration simplifies the HDR process by eliminating the need for separate vector delivery, thereby maintaining precision while reducing procedural complexity.
Data Source
AI summary
The invention provides for delivery, engineering and optimization of systems, methods, and compositions for manipulation of sequences and/or activities of target sequences. Provided are delivery systems and tissues or organ which are targeted as sites for delivery. Also provided are vectors and vector systems some of which encode one or more components of a CRISPR complex, as well as methods for the design and use of such vectors. Also provided are methods of directing CRISPR complex formation in eukaryotic cells to ensure enhanced specificity for target recognition and avoidance of toxicity and to edit or modify a target site in a genomic locus of interest to alter or improve the status of a disease or a condition.