Compositions useful in treatment of spinal muscular atrophy
A rAAV vector is described herein which has an AAVhu68 capsid and at least one expression cassette in the capsid. The at least one expression cassette comprises nucleic acid sequences encoding a functional SMN protein and expression control sequences that direct expression of the SMN sequences in a host cell. Also provided are compositions containing this rAAVhu68.SMN vector and methods of using same for spinal muscular atrophy in a patient.
1. A recombinant adeno-associated viral (rAAV) vector comprising an AAVhu68 capsid and at least one expression cassette, wherein the at least one expression cassette comprises a nucleic acid sequences encoding a functional survival of motor neuron (SMN) 1 protein and expression control sequences that direct expression of the SMN1 protein in a host cell, wherein the rAAV has an AAVhu68 capsid comprising: AAVhu68 VP1 proteins, AAVhu68 VP2 proteins and AAVhu68 VP3 proteins expressed from a nucleic acid sequence encoding SEQ ID NO: 8 (amino acids 1 to 736).
2. The rAAV vector of claim 1 , wherein the encoded SMN1 protein is an SMN1 isoform D protein.
3. The rAAV vector of claim 2 , wherein the encoded SMN1 isoform D protein has the amino acid sequence of SEQ ID NO: 2.
4. The rAAV vector of claim 1 , wherein the nucleic acid sequence is encoding SMN1 isoform D is selected from:
(a) SEQ ID NO: 1 or a sequence at least 70% identity therewith which encodes an SMN1 protein having the amino acid sequence of SEQ ID NO:2;
(b) SEQ ID NO: 4;
(c) SEQ ID NO: 5,
(d) SEQ ID NO: 6, or
(e) SEQ ID NO: 3 or a sequence at least 99% identical to SEQ ID NO: 3 which encodes functional SMN1 isoform D protein.
5. The rAAV vector of claim 1 , wherein the expression control sequences comprise a promoter.
6. The rAAV vector of claim 5 , wherein the promoter is a chicken β-actin (CB) promoter.
7. The rAAV vector of claim 1 , wherein the promoter is a CB7 promoter comprising a cytomegalovirus enhancer and a chicken beta actin promoter.
8. The rAAV vector of claim 1 , further comprising one or more of an enhancer, an intron, a Kozak sequence, a polyA, a post-transcriptional regulatory element.
9. The AAV vector of claim 1 , further comprising AAV inverted terminal repeat (ITRs) sequences from an AAV different from the AAV supplying the capsid.
10. The AAV vector of claim 9 , wherein the ITRs are from AAV2.
11. A recombinant adeno-associated virus (rAAV) vector comprising an AAVhu68 capsid which has packaged therein a vector genome comprising an AAV 5′ ITR, a CB7 promoter which comprises a cytomegalovirus enhancer and a chicken beta actin promoter, an intron, a nucleic acid sequences of SEQ ID NO:1, a polyA, and an AAV 3′ inverted terminal repeat sequence, wherein the AAVhu68 capsid comprises AAVhu68 VP1 proteins, AAVhu68 VP2 proteins and AAVhu68 VP3 proteins expressed from a nucleic acid sequence encoding SEQ ID NO: 8 (amino acids 1 to 736).
12. The rAAV of claim 11 , wherein the vector genome has the sequence of SEQ ID NO: 15 or SEQ ID NO: 25.
13. A pharmaceutical composition comprising a pharmaceutically acceptable carrier, excipient and/or preservative and the rAAV vector of claim 1 .
14. A pharmaceutical composition comprising a pharmaceutically acceptable carrier, excipient and/or preservative and the rAAV vector of claim 11 .