Novel insertion sites in pox vectors

Abstract

The present invention provides novel insertion sites for introducing DNA into pox vectors.

1 . A recombinant poxvirus containing and capable of expressing at least one foreign gene inserted at an insertion site within the poxvirus genome, wherein the insertion site is located at an intergenic region between two naturally occurring, adjacent, open reading frames of the poxvirus genome, and wherein insertion of the foreign gene at said insertion site does not effect translation of the two adjacent open reading frames. 
     
     
         2 . The recombinant poxvirus of  claim 1 , wherein the poxvirus is selected from the group consisting of orthopox, suipox, avipox, capripox, leporipox, and iridoviruses. 
     
     
         3 . The recombinant poxvirus of  claim 2 , wherein the poxvirus is an avipox. 
     
     
         4 . The recombinant poxvirus of  claim 3 , wherein avipox is fowlpox and the insertion site is located between a pair of adjacent open reading frames, wherein the pair is selected from the group consisting of FPV107/FPV108, FPV060/FPV061, FPV006/FPV007, and FPV254/FPV255. 
     
     
         5 . The recombinant poxvirus of  claim 1 , wherein the poxvirus is an orthopoxvirus. 
     
     
         6 . The recombinant poxvirus of  claim 5 , wherein the orthopoxvirus is a vaccinia virus. 
     
     
         7 . The recombinant poxvirus of  claim 6 , wherein the vaccinia virus is vaccinia Copenhagen and the insertion site is located between a pair of adjacent open reading frames, wherein the pair is selected from the group consisting of F14L/F15L, E2L/E3L, and A13L/A14L. 
     
     
         8 . The recombinant poxvirus of  claim 6 , wherein the vaccinia virus is an attenuated vaccinia virus. 
     
     
         9 . The recombinant poxvirus of  claim 8 , wherein the attenuated vaccinia virus is Modified Vaccinia Ankara (MVA) or NYVAC. 
     
     
         10 . The recombinant poxvirus of  claim 9 , wherein the attenuated vaccinia virus is MVA and the insertion site is located between a pair of adjacent open reading frames, wherein the pair is selected from the group consisting of MVA 44/45, MVA 49/50, and MVA 124/125. 
     
     
         11 . The recombinant poxvirus of  claim 1 , wherein the foreign gene encodes a marker, a therapeutic agent, or an antigenic determinant. 
     
     
         12 . The recombinant poxvirus of  claim 11 , wherein the foreign gene codes for an antigenic determinant selected from the group consisting of a pathogenic virus, a bacteria, other microorganism, a parasite, and a tumor cell. 
     
     
         13 . The recombinant poxvirus of  claim 11 , wherein the foreign gene codes for an antigenic determinant expressed in a pathogen, wherein the pathogen is selected from the group consisting of  Plasmodium , Mycobacteriua, Herpes virus, influenza virus, hepatitis, and HIV. 
     
     
         14 . The recombinant poxvirus of  claim 1 , wherein the foreign gene is selected from the group of genes encoding hormones, growth factors, enzymes, cytokines, receptors, and tumor suppressor genes. 
     
     
         15 . The recombinant poxvirus of  claim 1 , wherein said virus is used to induce an immune response in a subject. 
     
     
         16 . The recombinant poxvirus of  claim 15 , wherein said immune response in a subject is humoral. 
     
     
         17 . The recombinant poxvirus of  claim 15 , wherein said immune response in a subject is cellular. 
     
     
         18 . A method of gene delivery of a foreign gene in a target cell, wherein the target cell is used in somatic cell therapy or gene therapy, which comprises introducing into the target cell the recombinant poxvirus of  claim 1 . 
     
     
         19 . The method of  claim 18 , wherein the target cell is used in somatic cell therapy. 
     
     
         20 . The method of  claim 18 , wherein the target cell is a tumor infiltrating lymphocyte. 
     
     
         21 . The method of  claim 18 , wherein the gene is selected from the group of genes encoding hormones, growth factors, enzymes, cytokines, receptors, and tumor suppressor genes. 
     
     
         22 . A pharmaceutical composition, comprising the recombinant poxvirus of  claim 1 . 
     
     
         23 . The pharmaceutical composition of  claim 22 , wherein the pharmaceutical composition is a vaccine. 
     
     
         24 . The pharmaceutical composition of  claim 22 , wherein the pharmaceutical composition is an adjuvant. 
     
     
         25 . The pharmaceutical composition of  claim 22 , wherein the pharmaceutical composition is an activator, suppressor, and/or stabilizer of the immune system. 
     
     
         26 . A method for immunization comprising administering to a subject in need thereof an effective amount of a composition according to  claim 23 . 
     
     
         27 . A method for the activation, suppression, and/or stabilization of the unspecific immune system of a subject in need thereof, comprising administering an effective amount of a composition according to  claim 25 . 
     
     
         28 . A method for enhancing a specific immune response against an antigenic determinant in a vaccine comprising administering the composition of  claim 24 . 
     
     
         29 . A method for inducing an immunological response against at least one antigen in a mammal, said method comprising inoculating the mammal with at least a first recombinant poxvirus containing and capable of expressing at least the gene encoding said antigen inserted at a site within the poxvirus genome, wherein the insertion site is located between two naturally occurring, adjacent, open reading frames of the poxvirus genome. 
     
     
         30 . The method of  claim 29 , wherein said recombinant poxvirus further contains a gene encoding at least one immunostimulatory molecule. 
     
     
         31 . The method of  claim 29 , wherein said method further comprises inoculating the mammal with a second vector containing and capable of expression at least one additional gene. 
     
     
         32 . The method of  claim 31 , wherein said second vector contains a gene encoding at least one immunostimulatory molecule. 
     
     
         33 . The method of  claim 29 , wherein the first recombinant vector comprises a poxvirus selected from the group consisting of orthopox, suipox, avipox, capripox, leporipox, and iridoviruses. 
     
     
         34 . The method of  claim 29 , wherein the first recombinant vector comprises avipox. 
     
     
         35 . The method according to  claim 29 , wherein the first recombinant vector comprises a recombinant vaccinia virus vector. 
     
     
         36 . The method of  claim 35 , wherein the recombinant vaccinia virus is an attenuated vaccinia virus and the insertion site is located between a pair of adjacent open reading frames, wherein the pair is selected from the group corresponding to F14L/F15L, E2L/E3L, and A13L/A14L of vaccinia Copenhagen. 
     
     
         37 . The method of  claim 35 , wherein the recombinant vaccinia virus is MVA and the insertion site is located between a pair of adjacent open reading frames, wherein the pair is selected from the group consisting of MVA 44/45, MVA 49/50, and MVA 124/125. 
     
     
         38 . The method of  claim 29 , wherein the first recombinant vector comprises a recombinant fowlpoxvirus vector. 
     
     
         39 . The method according to  claim 38 , wherein the first recombinant vector is a fowlpox vector and the insertion site is located between a pair of adjacent open reading frames, wherein the pair is selected from the group consisting of FPV107/FPV108, FPV060/FPV061, FPV006/FPV007, and FPV254/FPV255. 
     
     
         40 . The method of  claim 29 , wherein the second vector comprises an adenovirus vector. 
     
     
         41 . The method of  claim 29 , wherein the second vector comprises a recombinant vaccinia virus vector. 
     
     
         42 . The method of  claim 13 , wherein the second recombinant vector comprises a recombinant fowlpoxvirus vector.