Retroviral vector having immune-stimulating activity

Abstract

The disclosure provides vectors for treating cancers, method of producing such vectors and methods of use of the vectors.

1 . A recombinant retroviral vector comprising at least one expression cassette comprising at least one agent that down regulates an immune inhibitory agent. 
     
     
         2 . The recombinant retroviral vector of  claim 1 , comprising at least two cassettes. 
     
     
         3 . The recombinant retroviral vector of  claim 1 , wherein the vector comprises two agents that down regulate an immune inhibitory agent. 
     
     
         4 . The recombinant retroviral vector of  claim 1 , wherein the at least one agent comprises a polynucleotide encoding a single-chain antibody and/or an inhibitory nucleic acid sequence. 
     
     
         5 . The recombinant retroviral vector of  claim 1 , wherein the at least one agent comprises an inhibitory nucleic acid sequence and a polynucleotide encoding a polypeptide having cytosine deaminase activity. 
     
     
         6 . The retroviral vector of  claim 1  wherein the at least one agent comprises an inhibitory nucleic acid sequence and wherein the vector further comprises a second agent selected from the group consisting of (i) a second inhibitory nucleic acid sequence, (ii) a polynucleotide encoding a single-chain antibody, (iii) a polynucleotide encoding an polypeptide the converts a prodrug to a cytotoxic drug, and (iv) a polynucleotide encoding a cytokine or chemokine. 
     
     
         7 . The retroviral vector of  claim 2 , wherein a first cassette comprises a polIII promoter operably linked to an inhibitory nucleic acid sequence; and a second cassette comprises a mini-promoter operably linked to a polynucleotide encoding a polypeptide having cytosine deaminase or a single chain antibody. 
     
     
         8 . The retroviral vector of  claim 2 , wherein a first cassette comprises a polIII promoter operably linked to a first inhibitory nucleic acid sequence; and a second cassette comprises a mini-promoter operably linked to a second inhibitory nucleic acid sequence. 
     
     
         9 . The recombinant retroviral vector of  claim 1  wherein the retroviral vector is replication competent. 
     
     
         10 . The recombinant retroviral vector of  claim 1 , wherein the retroviral vector is replication defective. 
     
     
         11 . The recombinant retroviral vector of  claim 9 , wherein the vector has the general structure from 5′ to 3′ comprising a long terminal repeat (LTR)-gag sequence-pol sequence-env sequence-(at least one expression cassette)-LTR. 
     
     
         12 . The recombinant retroviral vector of  claim 11 , wherein the vector comprises a retroviral polynucleotide sequence derived from murine leukemia virus (MLV), Moloney murine leukemia virus (MoMLV), Feline leukemia virus (FeLV) or Gibbon ape leukemia virus (GALV). 
     
     
         13 . The recombinant retroviral vector of  claim 12 , wherein the MLV is an amphotropic MLV. 
     
     
         14 . The recombinant retroviral vector of  claim 11 , wherein vector comprises:
 a retroviral GAG protein;   a retroviral POL protein;   a retroviral envelope;   a retroviral polynucleotide comprising:   Long-Terminal Repeat (LTR) sequences at the 3′ end of the retroviral polynucleotide sequence,   a promoter sequence at the 5′ end of the retroviral polynucleotide, said promoter being suitable for expression in a mammalian cell,   a gag nucleic acid domain, a pol nucleic acid domain; and   an env nucleic acid domain;   at least one cassette comprising a heterologous nucleic acid sequence, wherein the cassette is positioned 5′ to the 3′ LTR and 3′ to the env nucleic acid domain encoding the retroviral envelope; and   cis-acting sequences necessary for reverse transcription, packaging and integration in a target cell.   
     
     
         15 . The recombinant retroviral vector of  claim 14 , wherein the promoter comprises a CMV promoter having a sequence as set forth in SEQ ID NO: 1-15 or 16 from nucleotide 1 to about nucleotide 582 and may include modification to one or more nucleic acid bases and which is capable of directing and initiating transcription. 
     
     
         16 . The recombinant retroviral vector of  claim 14 , wherein the promoter comprises a sequence as set forth in SEQ ID NO: 1-15 or 16 from nucleotide 1 to about nucleotide 582. 
     
     
         17 . The recombinant retroviral vector of  claim 15 , wherein the promoter comprises a CMV-R-U5 domain polynucleotide. 
     
     
         18 . The recombinant retroviral vector of  claim 17 , wherein the CMV-R-U5 domain comprises the immediately early promoter from human cytomegalovirus linked to an MLV R-U5 region. 
     
     
         19 - 44 . (canceled) 
     
     
         45 . A method of treating a cancer or cell proliferative disorder comprising contacting a subject with the cancer of cell proliferative disorder with a recombinant retroviral vector of  claim 1 . 
     
     
         46 . A method of treating a cancer or cell proliferative disorder comprising contacting a subject with the cancer of cell proliferative disorder with a recombinant retroviral vector of  claim 1 , that comprises a cytosine deaminase gene, and additionally treating the subject with 5-fluorocytosine.