Single-vector gene construct comprising insulin and glucokinase genes

Abstract

The invention relates to a viral expression construct and related viral vector and composition and to their use wherein said construct and vector comprise elements a) and b): a) a nucleotide sequence encoding an insulin operably linked to a first promoter, b) a nucleotide sequence encoding a glucokinase operably linked to a second promoter and said viral expression construct and related viral vector comprise at least one of elements c), d) and e): c) the first and the second promoters are positioned in reverse orientation within the expression construct, d) the first and the second promoters are positioned in reverse orientation within the expression construct and are located adjacent to each other and e) the first promoter is a CMV promoter, preferably a mini CMV promoter.

1 . A method for preventing, delaying, reverting, curing and/or treating a diabetes using a viral expression construct wherein said viral expression construct comprises the elements a) and b):
 a) a nucleotide sequence encoding an insulin operably linked to a first promoter,   b) a nucleotide sequence encoding a glucokinase operably linked to a second promoter and said viral expression construct comprising at least one of elements c), d) and e):   c) the first and the second promoters are positioned in reverse orientation within the expression construct,   d) the first and the second promoters are positioned in reverse orientation within the expression construct and are located adjacent to each other and   e) the first promoter is a CMV promoter.   
     
     
         2 . The method of  claim 1 , wherein said CMV promoter is a mini CMV promoter. 
     
     
         3 . The method of  claim 1 , wherein said viral expression construct comprises elements a), b) and d) or wherein said construct comprises elements a), b) and e) wherein the first promoter is a mini CMV promoter. 
     
     
         4 . The method of  claim 1 , wherein said viral expression construct is such that, the first promoter is a CMV promoter, and/or the second promoter is a RSV promoter. 
     
     
         5 . The method of  claim 1 , wherein said viral expression construct comprises an additional sequence selected from the group consisting of: ITRs, SV40 polyadenylation signal, SV40 enhancer sequence, bGH polyadenylation signal and SV40 polyadenylation signal and enhancer sequence. 
     
     
         6 . The method of  claim 1 , wherein the viral expression construct is represented by a nucleotide sequence comprising SEQ ID NO: 8, 9, 10, 11, 12, 13, 14, 15, 16, 27 or 29 or a sequence having at least 60% identity with SEQ ID NO: 8, 9, 10, 11, 12, 13, 14, 15, 16, 27 or 29. 
     
     
         7 . The method of  claim 1 , wherein said viral expression construct is comprised in a viral vector and said viral vector is a retrovirus vector, an adenovirus vector, an adeno-associated virus vector, a herpesvirus vector, a polyoma virus vector or a vaccinia virus vector. 
     
     
         8 . The method of  claim 7 , wherein said viral vector is an adeno-associated virus vector. 
     
     
         9 . The method of  claim 8 , wherein the adeno-associated virus vector is an AAV1 vector. 
     
     
         10 . The method of  claim 1 , wherein the viral expression construct or viral vector is comprised in a composition. 
     
     
         11 . The method of  claim 10 , wherein said composition is a pharmaceutical composition.