Akt Nucleic Acids, Polypeptides, and Uses Thereof

Abstract

The present invention relates to human Akt3 proteins and polypeptides. The invention also relates to isolated nucleic acids encoding human Akt3, to vectors containing them and to their therapeutic uses, in particular for gene therapy. Expression of Akt3 inhibits cell death associated with hypoxia, apoptosis or necrosis.

1 . An isolated human Akt3 protein comprising the sequence Cys-Gln-Gln-Ser-Asp-Cys-Gly-Met-Leu-Gly-Asn-Trp-Lys-Lys, or a substantially similar sequence, wherein the protein has a property selected from the following:
 A. it is encoded by a nucleic acid comprising SEQ ID NO:1;   B. it comprises an amino acid sequence as depicted in SEQ ID NO:2, splice variants thereof, or allelic variants thereof; and   C. it specifically binds to an antibody generated against an epitope within a peptide having the sequence Cys-Gln-Gln-Ser-Asp-Cys-Gly-Met-Leu-Gly-Asn-Trp-Lys-Lys, or a substantially similar sequence.   
     
     
         2 . The isolated protein of  claim 1  comprising an amino acid sequence as depicted in SEQ ID NO:2. 
     
     
         3 . The isolated human Akt3 protein according to  claim 1 , further comprising a tag sequence. 
     
     
         4 . The isolated human Akt3 protein according to  claim 3 , further comprising a myristylation sequence. 
     
     
         5 . An antigenic peptide comprising the sequence Cys-Gln-Gln-Ser-Asp-Cys-Gly-Met-Leu-Gly-Asn-Trp-Lys-Lys, or a substantially similar sequence, and is a fragment of human Akt3 protein, wherein the Akt3 protein has a property selected from the following:
 A. it is encoded by the nucleic acid of  claim 1 ;   B. it comprises an amino acid sequence as depicted in SEQ ID NO:2, splice variants thereof, or allelic variants thereof; and   C. it specifically binds to an antibody generated against an epitope within a peptide having the sequence Cys-Gln-Gln-Ser-Asp-Cys-Gly-Met-Leu-Gly-Asn-Trp-Lys-Lys, or a substantially similar sequence.   
     
     
         6 . The antigenic peptide of  claim 5 , consisting essentially of the sequence Cys-Gln-Gln-Ser-Asp-Cys-Gly-Met-Leu-Gly-Asn-Trp-Lys-Lys or an antigenic fragment thereof. 
     
     
         7 . An antibody which specifically binds a human Akt3 protein of  claim 1 . 
     
     
         8 . The antibody of  claim 7  which specifically recognizes an epitope within a peptide having the sequence Cys-Gln-Gln-Ser-Asp-Cys-Gly-Met-Leu-Gly-Asn-Trp-Lys-Lys. 
     
     
         9 . The antibody of  claim 8  which is polyclonal. 
     
     
         10 . A method of treating myocardial infarction or ischemia reperfusion injury, the method comprising administering to a patient suffering therefrom a nucleic acid comprising the nucleotide sequence of SEQ ID NO:1 or a nucleotide sequence encoding a polypeptide comprising the amino acid sequence of SEQ ID NO:2, wherein the nucleic acid is operably linked to a regulatory region. 
     
     
         11 . The method according to  claim 10 , wherein the nucleic acid is administered to cardiac myocytes of said patient. 
     
     
         12 . The method according to  claim 10 , wherein the nucleic acid is in a vector. 
     
     
         13 . The method according to  claim 12 , wherein the vector is a plasmid. 
     
     
         14 . The method according to  claim 12 , wherein the vector is a viral vector. 
     
     
         15 . The method according to  claim 14 , wherein the viral vector is selected from the group consisting of retroviruses, adenoviruses, adeno-associated viruses, vaccinia virus and HSV virus. 
     
     
         16 . The method according to  claim 10 , wherein the ischemia reperfusion injury is myocardial ischemia reperfusion injury or is associated with stroke, liver damage, renal failure, organ transplantation, or coronary artery bypass grafting. 
     
     
         17 . A method of screening for molecules that stimulate or inhibit Akt3 activity in a cell, the method comprising
 contacting an Akt3 protein according to  claim 1  with a candidate molecule; and   detecting Akt3 activity in the presence of the molecule.   
     
     
         18 . The method according to  claim 17 , wherein the Akt3 activity is inhibition of apoptosis. 
     
     
         19 . The method according to  claim 18 , wherein inhibition of apoptosis is measure by the presence of a marker gene. 
     
     
         20 . The method according to  claim 17 , wherein the molecule is an agonist of Akt3. 
     
     
         21 . The method according to  claim 17 , wherein the molecule is an antagonist of Akt3. 
     
     
         22 . A method of increasing Akt3 activity in a cell comprising increasing the level of an Akt3 protein according to  claim 1  in the cell. 
     
     
         23 . The method according to  claim 22 , wherein the cell has been transfected with a vector encoding Akt3 under conditions permitting expression of the Akt3 protein. 
     
     
         24 . A method of inhibiting Akt3 activity in a cell, comprising decreasing the level of an Akt3 protein according to  claim 1  in the cell. 
     
     
         25 . The method according to  claim 24 , wherein the level of Akt3 protein is decreased by introducing an Akt3 antisense nucleic acid into the cell, which antisense nucleic acid hybridizes under intracellular conditions to an Akt3 mRNA. 
     
     
         26 . The method according to  claim 24  wherein the level of Akt3 protein is decreased by introducing a single chain Fv antibody (scFv) that specifically binds Akt3 into the cell at a level sufficient to bind to and inactivate Akt3.