ABCA4 Minigene AAV Vectors for Stargardt Gene Delivery Limits
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Solution Overview
Problem
Conventional Adeno-associated Viral (AAV) vectors are limited in packaging capacity, making it difficult to deliver the ABCA4 gene, which is 7 kb long, for treating Stargardt Disease caused by mutations in the ABCA4 gene.
Innovation Solution
Development of gene therapy vectors, including recombinant AAV vectors, that contain ABCA4 minigenes flanked by AAV inverted terminal repeats, with or without terminal resolution sites, and are formulated for ocular delivery to photoreceptor cells, using capsids with tropism for the eye, such as AAV8, to treat Stargardt Disease.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If conventional AAV vectors are used for gene delivery, then the vector structure is simple and well-characterized, but the packaging capacity is limited and cannot accommodate the 7 kb ABCA4 gene
Solution Approach 1:
The patent divides the ABCA4 gene into multiple smaller cDNA fragments that can be individually packaged into AAV vectors. Each fragment contains specific exons and introns of the ABCA4 gene, allowing the complete gene to be delivered through multiple vector administrations or combined expression cassettes, thereby overcoming the packaging capacity limitation of conventional AAV vectors.
2Reliability
If the complete ABCA4 gene is delivered, then the therapeutic effect is maximized, but the delivery exceeds the packaging capacity of conventional AAV vectors
Solution Approach 1:
The patent employs partial action by delivering cDNA fragments containing specific functional domains of the ABCA4 gene rather than the complete gene. The fragments include essential exons (e.g., exons 26-36 in some embodiments) that encode critical functional regions of the protein, providing sufficient therapeutic effect while remaining within AAV packaging capacity constraints.
3Quantity of substance
If minigenes are used to fit within AAV packaging capacity, then the gene can be delivered, but the protein expression may be reduced compared to the complete gene
Solution Approach 1:
The patent optimizes the structure of minigenes by carefully selecting which exons and introns to include, adjusting the configuration of expression cassettes, and modifying regulatory elements to maximize protein expression from the truncated gene fragments. These parameter changes ensure that despite the reduced gene size, the therapeutic protein is expressed at sufficient levels to treat Stargardt disease.
Data Source
AI summary
Aspects of the disclosure relate to compositions and methods useful for delivering minigenes to a subject. Accordingly, the disclosure is based, in part, on isolated nucleic acids and gene therapy vectors, such as viral (e.g., rAAV) vectors, comprising one or more gene fragments encoding a therapeutic gene product, such as a protein or peptide (e.g., a minigene). In some embodiments, the disclosure relates to gene therapy vectors encoding a ABCA4 protein (e.g., the gene product of ABCA4 gene) or a portion thereof. In some embodiments, compositions described by the disclosure are useful for treating diseases associated with mutations in the ABCA4 gene, for example Stargardt disease.