Gene delivery system and method of use

Abstract

A recombinant replication competent retrovirus for gene deliver and gene therapy is provided. The recombinant retrovirus has a heterologous nucleic acid sequence, a sequence encoding a cell- or tissue-specific ligand or a sequence for transcriptional targeting, or a combination of both a cell- or tissue-specific ligand and a cell- or tissue-specific transcriptional targeting sequence.

1 - 62 . (canceled) 
     
     
         63 . A recombinant replication competent mammalian retrovirus, comprising:
 an oncoretroviral polynucleotide sequence comprising:
 a nucleic acid sequence encoding a retroviral GAG protein; 
 a nucleic acid sequence encoding a retroviral POL protein; 
 a nucleic acid sequence encoding a retroviral envelope; 
 Long-Terminal Repeat (LTR) sequences at the 5′ and 3′ end of the oncoretroviral polynucleotide sequence; 
   a cassette comprising a regulatory nucleic acid sequence operably linked to a heterologous nucleic acid sequence, wherein the cassette is positioned 5′ to the U3 region of the 3′ LTR and 3′ to the nucleic acid sequence encoding the retroviral envelope; and   cis-acting sequences for reverse transcription, packaging and integration in a target naturally occurring mammalian cell.   
     
     
         64 . A recombinant replication competent mammalian retrovirus comprising:
 a retroviral GAG protein;   a retroviral POL protein;   a retroviral envelope (ENV);   a oncoretroviral polynucleotide sequence comprising Long-Terminal Repeat (LTR) sequences at the 5′ and/or 3′ end of the retroviral polynucleotide sequence, wherein a mammalian promoter sequence is contained within LTR sequences at the 5′ and/or 3′ end of the retroviral polynucleotide sequence, wherein the oncoretroviral polynucleotide sequence contains a coding sequence for GAG, POL and ENV;   a cassette comprising a regulatory nucleic acid sequence operably linked to a heterologous polynucleotide, wherein the cassette is positioned 3′ to the sequence encoding the retroviral envelope and 5′ to the 3′ LTR; and   cis-acting sequences necessary for reverse transcription, packaging and integration in a target cell.   
     
     
         65 . The recombinant replication competent mammalian retrovirus of  claim 64 , wherein said mammalian promoter is a heterologous mammalian promoter. 
     
     
         66 . The replication competent mammalian retrovirus of  claim 63 , further comprising a heterologous mammalian promoter sequence operably associated with Long-Terminal Repeat (LTR) sequences at the 5′ end of the oncoretroviral polynucleotide sequence. 
     
     
         67 . The replication competent mammalian retrovirus of  claim 65 , wherein the heterologous promoter sequence is a tissue-specific promoter sequence that permits replication of the retrovirus in a targeted tissue. 
     
     
         68 . The replication competent mammalian retrovirus of  claim 66 , wherein the heterologous promoter sequence is a tissue-specific promoter sequence that permits replication of the retrovirus in a targeted tissue. 
     
     
         69 . The replication competent mammalian retrovirus of  claim 63 , wherein the heterologous nucleic acid sequence encodes a biological response modifier. 
     
     
         70 . The replication competent mammalian retrovirus of  claim 69 , wherein the biological response modifier comprises an immunopotentiating cytokine. 
     
     
         71 . The replication competent mammalian retrovirus of  claim 70 , wherein the immunopotentiating cytokine is selected from the group consisting of interleukins 1 through 12, interferon, tumor necrosis factor (TNF), and granulocyte-macrophage-colony stimulating factor (GM-CSF). 
     
     
         72 . The replication competent mammalian retrovirus of  claim 71 , wherein the immunopotentiating cytokine is interferon. 
     
     
         73 . The replication competent mammalian retrovirus of  claim 72 , wherein the interferon is gamma interferon. 
     
     
         74 . The replication competent mammalian retrovirus of  claim 63 , wherein the heterologous nucleic acid sequence encodes a polypeptide that converts a nontoxic prodrug in to a toxic drug. 
     
     
         75 . The replication competent mammalian retrovirus of  claim 64 , wherein the heterologous nucleic acid sequence encodes a polypeptide that converts a nontoxic prodrug in to a toxic drug. 
     
     
         76 . The replication competent mammalian retrovirus of  claim 74 , wherein the polypeptide that converts a nontoxic prodrug in to a toxic drug is thymidine kinase, purine nucleoside phosphorylase (PNP), or cytosine deaminase. 
     
     
         77 . The replication competent mammalian retrovirus of  claim 75 , wherein the polypeptide that converts a nontoxic prodrug in to a toxic drug is thymidine kinase, purine nucleoside phosphorylase (PNP), or cytosine deaminase. 
     
     
         78 . The replication competent mammalian retrovirus of  claim 63 , wherein the regulatory nucleic acid sequence is a promoter. 
     
     
         79 . The replication competent mammalian retrovirus of  claim 64 , wherein the regulatory nucleic acid sequence is a promoter.