Increased packaging efficiency of vector for cardiac gene therapy
View Patent ↗The present disclosure is related to compositions and methods useful in treating heart conditions. The disclosed compositions and methods are based on gene therapies comprising a recombinant AAV vector for delivering two or more transgenes into the heart of a subject, wherein the transgenes encode an S100A1 protein and a cardiac Apoptosis Repressor with caspase recruitment Domain (cARC) apoptotic inhibitor, respectively. In various embodiments, the compositions and methods disclosed herein comprise vectors comprising S100A1 and/or cARC cDNA sequences that are codon-optimized for expression in humans. In various embodiments, the compositions and methods disclosed herein comprise vectors with improved packaging efficiencies. In some aspects, targeting multiple sources of one or more heart conditions can provide synergistic benefits during treatment.
1 . A recombinant adeno-associated virus (rAAV) nucleic acid vector for delivering two or more transgenes into the heart of a subject, wherein said vector comprises, from 5′ to 3′, in order, a first adeno-associated virus (AAV) inverted terminal repeat (ITR) sequence, two or more transgenes and a promoter operably linked to the two or more transgenes, a polyadenylation (polyA) signal, and a second AAV ITR sequence,
wherein the two or more transgenes encode an S100 family protein and an apoptotic inhibitor, respectively, and
wherein the transgene encoding an S100 family protein comprises a sequence that is at least 95%, identical to at least one of the nucleotide sequences of SEQ ID NOs: 25 and 26.
2 . The rAAV nucleic acid vector of claim 1 , wherein the polyA signal comprises a nucleotide sequence that has at least 95% identity to the nucleotide sequence of SEQ ID NO: 28.
3 . The rAAV nucleic acid vector of claim 1 , wherein the transgene encoding an S100 family protein comprises a sequence having a reduced guanosine (G) and cytosine (C) (G/C) content, relative to any one of the nucleotide sequences of SEQ ID NOs: 5, 8, 19, and 20.
4 . The rAAV nucleic acid vector of claim 1 , wherein the apoptotic inhibitor is cardiac Apoptosis Repressor with Caspase Recruitment Domain (ARC) or a variant thereof.
5 . The rAAV nucleic acid vector of claim 1 , wherein the transgene encoding an apoptotic inhibitor comprises a sequence that is at least 90%, at least 95%, or at least 99.5% identical to any one of the nucleotide sequences of SEQ ID NOs: 3, 6, 7, and 15-18.
6 . The rAAV nucleic acid vector of claim 1 , wherein the promoter is a cardiac-restricted promoter.
7 . The rAAV nucleic acid vector of claim 1 , wherein the rAAV nucleic acid vector is single-stranded.
8 . The rAAV nucleic acid vector of claim 1 , wherein the rAAV nucleic acid vector is self-complementary.
9 . The rAAV nucleic acid vector of claim 1 , wherein the rAAV nucleic acid vector comprises a nucleotide sequence that is at least 90%, at least 95% or at least 99.5% identical to either of the nucleotide sequences of SEQ ID NOs: 22 and 23.
10 . An rAAV particle comprising the rAAV nucleic acid vector of claim 1 encapsidated in an AAV capsid.
11 . The rAAV particle of claim 10 , wherein the AAV capsid comprises a capsid protein derived from an AAV1, AAV2, AAV3, AAV6, AAV8, AAVrh.74, AAVrh.10, AAV2/6, or AAV9 serotype.
12 . A composition comprising the rAAV particle of claim 10 .
13 . A method of treatment of a subject suffering from a heart disease comprising administering to the subject the composition of claim 12 .
14 . The method of claim 13 , wherein the composition is administered via injection into the heart of the subject or intravascular injection into the coronary arteries of the subject.
15 . The method of claim 13 , wherein the step of administering results in expression of the two or more transgenes in the subject's heart, improved cardiac function in the subject, or both.
16 . The method of claim 13 , wherein the subject is a mammal.
17 . The method of claim 13 , wherein the step of administering results in improved cardiac function in the subject for more than 10 months.
18 . The rAAV nucleic acid vector of claim 1 , wherein the transgene encoding an S100 family protein is positioned 5′ to the transgene encoding an apoptotic inhibitor.
19 . The rAAV nucleic acid vector of claim 1 , wherein the transgene encoding an apoptotic inhibitor is positioned 5′ to the transgene encoding an S100 family protein.
20 . The rAAV nucleic acid vector of claim 1 , wherein the rAAV nucleic acid vector encodes a protein comprising an amino acid sequence at least 90%, at least 95%, or at least 99.5% identical to the amino acid sequence of any one of SEQ ID NOs: 13, 14, 24 or 29.