Adeno-associated virus serotype 1 nucleic acid sequences, vectors and host cells containing same

Abstract

The nucleic acid sequences of adeno-associated virus (AAV) serotype 1 are provided, as are vectors and host cells containing these sequences and functional fragments thereof. Also provided are methods of delivering genes via AAV-1 derived vectors.

1 . An isolated AAV-1 nucleic acid molecule comprising a sequence selected from the group consisting of: 
 (a) SEQ ID NO: 1;    (b) a DNA sequence complementary to SEQ ID NO: 1;    (c) cDNA complementary to (a) or (b); and    (d) RNA complementary to any of (a) to (c).    
     
     
         2 . A nucleic acid molecule comprising an AAV-1 inverted terminal repeat (ITR) sequence selected from the group consisting of: 
 (a) nt 1 to 143 of SEQ ID NO: 1;    (b) nt 4576 to 4718of SEQ ID NO: 1;    (c) a nucleic acid sequence complementary to (a) or (b); and    (d) a functional fragment of (a), (b), or (c).    
     
     
         3 . A recombinant vector comprising a 5′ AAV-1 inverted terminal repeat (ITR) and a selected transgene, wherein said ITR has the sequence selected from the group consisting of: 
 (a) nt 1 to 143 of SEQ ID NO: 1;    (b) a nucleic acid sequence complementary to (a); and    (c) a functional fragment of (a) or (b).    
     
     
         4 . The recombinant vector according to  claim 3 , wherein said vector further comprises a 3′ AAV-1 ITR.  
     
     
         5 . A recombinant vector comprising a 3′ AAV-1 inverted terminal repeat (ITR) and a selected transgene, wherein said ITR has the sequence selected from the group consisting of: 
 (a) nt 4576 to 4718 of SEQ ID NO: 1;    (b) a nucleic acid sequence complementary to (a); and    (c) a functional fragment of (a) or (b).    
     
     
         6 . The recombinant vector according to  claim 5 , wherein said vector further comprises a 5′ AAV-1 ITR.  
     
     
         7 . A pharmaceutical composition comprising a carrier and a virus comprising the vector according to  claim 5 .  
     
     
         8 . A method for producing a selected gene product comprising the steps of transfecting a mammalian cell with the molecule according to  claim 1  or a functional fragment thereof and culturing said cell under conditions suitable to express said gene product.  
     
     
         9 . The recombinant vector according to  claim 3 , wherein said vector further comprises AAV-1 capsid proteins having the sequence of SEQ ID NO: 13, 15 or 17 or functional fragments thereof.  
     
     
         10 . The recombinant vector according to  claim 3 , wherein said vector further comprises adenovirus sequences.  
     
     
         11 . The host cell transduced with a recombinant viral vector according to  claim 3 .  
     
     
         12 . The host cell transduced with a nucleic acid molecule according to  claim 1 .  
     
     
         13 . The host cell transduced with a nucleic acid molecule according to  claim 2 .  
     
     
         14 . The pharmaceutical composition comprising a carrier and a virus comprising the vector according to  claim 3 .  
     
     
         15 . The method for delivery of a transgene comprising the step of delivering to a host cell a recombinant virus comprising a recombinant vector according to  claim 3 .  
     
     
         16 . A recombinant host cell transformed with a nucleic acid sequence expressing one or more AAV-1 rep proteins selected from among rep78 having the amino acid sequence of SEQ ID NO:7, rep 68 having the amino acid sequence of SEQ ID NO:7, rep 52 having the amino acid sequence of SEQ ID NO:9, and rep 40 having the amino acid sequence of SEQ ID NO: 11.  
     
     
         17 . A composition comprising a recombinant virus having an AAV-1 capsid comprising an AAV-1 protein selected from among AAV-1 vp1 having the amino acid sequence of SEQ ID No: 13; AAV-1 vp2 having the amino acid sequence of SEQ ID NO: 15 and AAV-1 vp3 having the amino acid sequence of SEQ ID NO:  17  and a heterologous molecule which comprises an AAV 5′ inverted terminal repeat sequence (ITR), a transgene, and an AAV 3′ ITR.  
     
     
         18 . The composition of  claim 17  wherein the AAV-1 protein vp1 is encoded by a nucleic acid having at least about 98% identity to nucleotides 2223-4431 of SEQ ID NO: 1, as measured by MacVector 6.0.  
     
     
         19 . The composition of  claim 17  wherein the AAV-1 protein vp2 is encoded by a nucleic acid having at least about 98% identity to nucleotides 2634-4432 of SEQ ID NO: 1, as measured by MacVector 6.0.  
     
     
         20 . The composition of  claim 17  wherein the AAV-1 protein vp3 is encoded by a nucleic acid having at least about 98% identity to nucleotides 2829-4432 of SEQ ID NO: 1 as measured by MacVector6.0.  
     
     
         21 . The composition of  claim 17  wherein the AAV 5′ ITR and 3′ ITR are of AAV serotype 2.  
     
     
         22 . The composition of  claim 21  wherein the recombinant virus further comprises a regulatable promoter which directs expression of the transgene.