Gene therapy vectors for treatment of danon disease

Abstract

The disclosure relates to optimized polynucleotide sequences for LAMP-2B, expression cassettes, vectors, and methods of use thereof in treating disease, e.g. Danon disease.

1 . A gene therapy vector comprising an expression cassette comprising a transgene encoding an isoform of lysosome-associated membrane protein 2 (LAMP-2) or a functional variant thereof, wherein the transgene is optimized for expression in a human host cell. 
     
     
         2 .- 17 . (canceled) 
     
     
         18 . A pharmaceutical composition comprising the gene therapy vector of  claim 1 . 
     
     
         19 . A method of treating or preventing Danon disease or another autophagy disorder in a subject in need thereof, comprising administering to the subject the gene therapy vector of  claim 1 . 
     
     
         20 . The method of  claim 19 , wherein the vector or pharmaceutical composition is administered via an intravenous route. 
     
     
         21 . The method of  claim 19 , wherein the autophagy disorder is selected from the group consisting of end-stage heart failure, myocardial infarction, drug toxicities, diabetes, end-stage renal failure, and aging. 
     
     
         22 . (canceled) 
     
     
         23 . The method of  claim 19 , wherein the subject is exhibiting symptoms of Danon disease or another autophagy disorder. 
     
     
         24 . The method of  claim 19 , wherein the subject has been identified as having reduced or non-detectable LAMP-2 expression. 
     
     
         25 . The method of  claim 19 , wherein the subject has been identified as having a mutated LAMP-2 gene. 
     
     
         26 . The method of  claim 19 , wherein administration of the gene therapy vector results in increased expression of LAMP-2B polynucleotide in the heart compared to an untreated subject. 
     
     
         27 . The method of  claim 19 , wherein administration of the gene therapy vector results in at least about 1.2-fold increased expression of LAMP-2B polynucleotide in the heart compared to an untreated subject. 
     
     
         28 . (canceled) 
     
     
         29 . A method of expressing LAMP-2B in a subject, comprising systemically administering an adeno-associated viral (AAV) vector to the subject, wherein the AAV vector comprises an expression cassette comprising a transgene sharing at least 95% identity with any one of SEQ ID NOs: 3-5, the transgene operatively linked to an enhancer/promoter region, wherein systemic administration of the AAV vector to the subject results in increased expression of LAMP-2B compared to expression of LAMP-2B prior to administration of the AAV vector. 
     
     
         30 . (canceled) 
     
     
         31 . The method of  claim 29 , wherein the AAV vector is administered at a dose of between about 1×10 12  and 5×10 14  vector genomes (vg) of the AAV vector per kilogram (vg) of total body mass of the subject (vg/kg). 
     
     
         32 .- 34 . (canceled) 
     
     
         35 . The method of  claim 29 , wherein the enhancer/promoter region comprises in the 5′ to 3′ direction a CMV IE Enhancer and a Chicken Beta-Actin Promoter, and optionally wherein the enhancer/promoter region further comprises a first exon and first intron of a chicken beta-actin gene and a splice acceptor of a rabbit beta-globin gene. 
     
     
         36 . The method of  claim 29 , wherein the expression cassette comprises a consensus optimal Kozak sequence operatively linked to the transgene, wherein optionally the consensus optimal Kozak sequence comprises SEQ ID NO: 6. 
     
     
         37 . The method of  claim 29 , wherein the expression cassette comprises a full-length polyA sequence operatively linked to the transgene, wherein optionally the full-length polyA sequence comprises SEQ ID NO: 7. 
     
     
         38 . (canceled) 
     
     
         39 . The method of  claim 29 , wherein the expression cassette comprises operatively linked, in the 5′ to 3′ direction, a first inverted terminal repeat, an enhancer/promoter region, introns, a consensus optimal Kozak sequence, the transgene, a 3′ untranslated region including a full-length polyA sequence, and a second inverted terminal repeat, wherein the expression cassette comprises no start codon 5′ to the start codon of the transgene. 
     
     
         40 . (canceled) 
     
     
         41 . The method of  claim 29 , wherein the subject has been identified as having, or is suspected of having, reduced or non-detectable LAMP-2 expression. 
     
     
         42 . The method of  claim 29 , wherein the subject has a mutated LAMP-2 gene. 
     
     
         43 .- 44 . (canceled) 
     
     
         45 . The method of  claim 29 , wherein administration of the gene therapy vector results in increased expression of LAMP-2B polynucleotide in the heart compared to an untreated subject. 
     
     
         46 . The method of  claim 29 , wherein administration of the gene therapy vector results in at least about 1.2-fold increased expression of LAMP-2B polynucleotide in the heart compared to an untreated subject. 
     
     
         47 .- 74 . (canceled)