Gene therapies for Stargardt disease (ABCA4)
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.
1 . An isolated nucleic acid comprising a transgene having a nucleic acid sequence encoding a ABCA4 protein, wherein the ABCA4 protein comprises the amino acid sequence as set forth in any one of SEQ ID NOs: 9-14 and 20-24, wherein the ABCA4 protein does not comprise the amino acid sequence set forth in SEQ ID NO: 2.
2 . The isolated nucleic acid of claim 1 , wherein the transgene further comprises a promoter operably linked to the nucleic acid sequence encoding the ABCA4 protein.
3 . The isolated nucleic acid of claim 2 , wherein the promoter is a constitutive promoter, inducible promoter, or a tissue-specific promoter.
4 . The isolated nucleic acid of claim 1 , wherein the transgene is flanked by adeno-associated virus (AAV) inverted terminal repeats (ITRs).
5 . The isolated nucleic acid of claim 4 , wherein at least one of the ITRs is an AAV2 ITR.
6 . The isolated nucleic acid of claim 4 , wherein at least one ITR lacks a terminal resolution site.
7 . A vector comprising the isolated nucleic acid of claim 1 .
8 . A host cell comprising the isolated nucleic acid of claim 1 .
9 . A recombinant adeno-associated virus (rAAV) comprising:
(i) the isolated nucleic acid of claim 1 ; and
(ii) an AAV capsid protein.
10 . The rAAV of claim 9 , wherein the capsid protein has a tropism for ocular cells.
11 . The rAAV of claim 9 , wherein the rAAV is formulated for delivery to the eye.
12 . A composition comprising the isolated nucleic acid of claim 1 , and a pharmaceutically acceptable excipient.
13 . The isolated nucleic acid of claim 1 , wherein the nucleic acid sequence encoding the ABCA4 protein comprises the sequence set forth in any one of SEQ ID NOs: 3-8 and 15-19.
14 . The isolated nucleic acid of claim 3 , wherein the tissue-specific promoter is a photoreceptor-specific promoter.
15 . The vector of claim 7 , wherein the vector is a plasmid DNA, or closed-ended DNA, or a viral vector.
16 . The host cell of claim 8 , wherein the cell is a mammalian cell, bacterial cell, yeast cell, or insect cell.
17 . The rAAV of claim 10 , wherein the capsid protein is AAV8 capsid protein.
18 . The rAAV of claim 11 , wherein the rAAV is formulated for delivery to photoreceptor cells or retinal pigmented epithelium (RPE).