POLYPEPTIDES FOR ENGINEERING INTEGRASE CHIMERIC PROTEINS AND THEIR USE IN GENE THERAPY
The present invention relates to polypeptide for engineering integrase chimeric proteins and their use in gene therapy. In particular, the present invention relates to a polypeptide which comprises the amino acid sequence ranging from the amino acid residue at position 617 to the amino acid residue at position 622 in SEQ ID NO: 1 or a function conservative thereof.
1 . A polypeptide which comprises the amino acid sequence ranging from the amino acid residue at position 617 to the amino acid residue at position 622 in SEQ ID NO:1 or a function conservative thereof.
2 . The polypeptide of claim 1 wherein the function-conservative variant comprises an amino acid sequence having 50; 51; 52; 53; 54; 55; 56; 57; 58; 59; 60; 61; 62; 63; 64; 65; 66; 67; 68; 69; 70; 71; 72; 73; 74; 75; 76; 77; 78; 79; 80; 81; 82; 83; 84; 85; 86; 87; 88; 89; 90; 91; 92; 93; 94; 95; 96; 97; 98; or 99% identity with the amino acid sequence ranging from the amino acid residue at position 617 to the amino acid residue at position 622 in SEQ ID NO:1.
3 . The polypeptide of claim 1 wherein the amino acid at position 618 or 620 is substituted by any amino acid.
4 . The polypeptide of claim 1 wherein the amino acid residue at position 618 is substituted by an alanine (A).
5 . The polypeptide of claim 1 wherein the amino acid residue at position 620 is substituted by an aspartic acid residue (D).
6 . The polypeptide of claim 1 which comprises the amino acid sequence as set forth in SEQ ID NO:3 (KEMDSL).
7 . The polypeptide of claim 1 which comprises the amino acid sequence selected from the group consisting of SEQ ID NO:2 to SEQ ID NO:12 and amino acid sequences having at least 50% of identity with SEQ ID NO:2 to SEQ ID NO:12.
8 . The polypeptide of claim 1 which is fused to a heterologous polypeptide.
9 . A chimeric integrase which comprises the polypeptide of claim 1 .
10 . The chimeric integrase of claim 9 which comprises a retroviral integrase selected from the group consisting of feline immunodeficiency virus (FIV) integrases, Foamy virus (FV) integrases, murine leukemia virus (MLV) integrases, and lentivirus integrases.
11 . The chimeric integrase of claim 10 wherein the retroviral integrase has a sequence having at least 80% of identity with SEQ ID NO: 13 or SEQ ID NO:14.
12 . The chimeric integrase of claim 9 wherein the polypeptide is fused at the C-terminal end of the integrase protein or at the N-terminal end of the integrase protein.
13 . The chimeric integrase of claim 9 wherein the polypeptide is made part of a retroviral integrase.
14 . A nucleic acid molecule which encodes a polypeptide of claim 1 or a chimeric integrase comprising the polypeptide.
15 . A retroviral vector which comprises a nucleic acid molecule encoding the chimeric integrase of claim 9 .
16 . The retroviral vector of claim 15 which is derived from a retrovirus selected from the group consisting of alpharetroviruses, betaretroviruses, gammaretroviruses, deltaretroviruses, epsilonretroviruses, lentiviruses and spumaviruses.
17 . The retroviral vector of claim 15 which comprises a transgene.
18 . A method for expressing a transgene from the genome of a plurality of cells comprising infecting the plurality of cells with the retroviral vector of claim 17 under conditions which i) result in integration of the retroviral vector into the genome of the plurality of cells and ii) permit expression of the transgene from the integrated retroviral vector.
19 . A method of performing gene therapy in a subject in need thereof, comprising
providing the subject with a transgene by administering to the subject the retroviral vector of claim 17 , under conditions that allow expression of the transgene within the subject.