ABCA4 Trans-Splicing Molecules for Large-Gene Exon Correction
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Solution Overview
Problem
Current treatments for ABCA4-related retinopathies, such as Stargardt disease, are lacking, and existing strategies like AAV vectors and base-editing approaches are insufficient due to the large size of the ABCA4 gene and numerous disease-causing mutations, leaving a significant unmet need for effective therapeutic interventions.
Innovation Solution
Development of nucleic acid trans-splicing molecules, including RNA exon editors, that can trans-splice functional ABCA4 exons into endogenous pre-mRNA to correct mutations, utilizing specific domains for binding and splicing, and are delivered via vectors like AAV to target photoreceptor and retinal pigment epithelial cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Device complexity
If a single AAV vector is used to deliver ABCA4 gene therapy, then delivery simplicity is maintained, but the large size of ABCA4 gene (6882-bp coding sequence) cannot be accommodated
Solution Approach 1:
The ABCA4 gene is divided into multiple exons (22 exons total), and the invention delivers specific exon combinations through separate AAV vectors. This segmentation allows the large gene to be broken into deliverable units that can be individually packaged and then assembled in the target cells through trans-splicing.
Solution Approach 2:
Multiple exon sequences are nested within a single AAV vector construct, where the vector contains organized arrangements of exons that will be sequentially delivered and assembled. The nested structure allows efficient packaging of multiple gene segments within the limited AAV capacity.
2Ease of operation
If a single base-editing approach is used, then treatment simplicity is maintained, but it cannot address the hundreds of disease-causing mutations distributed throughout the ABCA4 gene
Solution Approach 1:
The AAV vector system is designed with universal applicability to address multiple different mutations. By delivering wild-type exon sequences that can replace any mutated exon through trans-splicing, the same vector platform can treat various mutation types and locations throughout the ABCA4 gene, providing a versatile solution for hundreds of different disease-causing mutations.
Solution Approach 2:
The treatment approach is made dynamic by allowing selection of different exon combinations in the AAV vectors based on the patient's specific mutation profile. The system can be adapted to deliver the appropriate exon segments needed to correct the particular mutations present in each patient's ABCA4 gene.
3Reliability
If trans-splicing molecules are used to correct ABCA4 mutations, then therapeutic effectiveness is improved, but trans-splicing efficiency has been historically poor
Solution Approach 1:
The invention introduces specific intermediary sequences (such as splice acceptor and donor sites, linker regions) that facilitate the trans-splicing process. These intermediary elements act as mediators between the delivered exon sequences and the endogenous ABCA4 pre-mRNA, enabling efficient recognition and splicing by the cellular machinery.
Solution Approach 2:
The trans-splicing efficiency is improved by optimizing various parameters of the trans-splicing molecules, including the sequence composition of exon boundaries, the length and sequence of linker regions, and the structural organization of the delivered RNA. These parameter optimizations enhance the recognition and processing efficiency by cellular splicing machinery.
Data Source
AI summary
Provided herein are nucleic acid trans-splicing molecules (e.g., pre-mRNA trans-splicing molecules (RTMs); RNA exon editing molecules) capable of correcting mutations in the ABCA4 gene. Such molecules are useful in the treatment of disorders such as ABCA4-associated retinal dystrophies (e.g., Stargardt Disease or cone-rod dystrophy). Also described herein are methods of using the nucleic acid trans-splicing molecules described herein to correct mutations in ABCA4, thereby treating disorders associated with mutations in ABCA4 and use of the nucleic acid trans-splicing molecules described herein for treating disorders associated with mutations in ABCA4 and in the preparation of medicaments for the treatment of disorders associated with mutations in ABCA4.