Suppressor of cytokine signaling (SOCS)-3 promoter and methods for its use in genetic therapy in humans

Abstract

Disclosed is a nucleic acid construct comprising a murine SOCS-3 promoter sequence having SEQ. ID. NO.: 1, or an operative fragment or derivative. The construct can also contain, operatively linked to the SOCS-3 promoter, a gene encoding any preselected protein, and optionally contains a reporter gene to facilitate detection and/or selection of successfully transfected cells. Also disclosed are a transgenic vertebrate cell containing the nucleic acid construct and transgenic non-human vertebrates comprising such cells. The nucleic acid construct is useful in methods of treating a growth retardation or growth acceleration disorder in a human subject and in a method of treating an autoimmune disease, immune disease, or inflammatory condition in a human subject. A kit for genetically modifying a vertebrate cell includes a polynucleotide comprising the murine SOCS-3 promoter sequence is also disclosed.

We claim:  
     
         1 . A nucleic acid construct, comprising a murine SOCS-3 promoter having a nucleotide sequence SEQ ID NO: 1, an operative fragment of SEQ ID NO: 1 having promoter activity that is inducible by a cytokine selected from the group consisting of LIF, IL-6, and IL-11, or an operative derivative of either of these, said derivative being selected from the group consisting of a mutation, a polymorphism, and a variant allele with respect to any nucleotide position of SEQ ID NO: 1, and said derivative being a derivative that does not fully eliminate promoter activity that is inducible by a cytokine selected from the group consisting of LIF, IL-6, and IL-11.  
     
     
         2 . The nucleic acid construct of  claim 1 , further comprising said SOCS-3 promoter operatively linked, in a transcriptional unit, to a DNA sequence encoding a protein of interest.  
     
     
         3 . The nucleic acid construct of  claim 2 , wherein the protein of interest is a SOCS-3 protein, or a functional fragment thereof.  
     
     
         4 . The nucleic acid construct of  claim 2 , wherein said SOCS-3 promoter is operatively linked to a DNA sequence encoding a reporter protein.  
     
     
         5 . The nucleic acid construct of  claim 4 , wherein the reporter protein is a fluorescent or light-emitting protein.  
     
     
         6 . The nucleic acid construct of  claim 5 , wherein the fluorescent or light-emitting protein is a green fluorescent protein, yellow fluorescent protein, blue fluorescent protein, phycobiliprotein, luciferase, or apoaequorin.  
     
     
         7 . The nucleic acid construct of  claim 1 , wherein the operative fragment is (SEQ ID NO: 2), (SEQ ID NO: 3), (SEQ ID NO: 4), (SEQ ID NO: 5), (SEQ ID NO: 6), (SEQ ID NO: 7), (SEQ ID NO: 8), (SEQ ID NO: 9), (SEQ ID NO: 10), (SEQ ID NO: 11), (SEQ ID NO: 12), (SEQ ID NO: 35), (SEQ ID NO: 36), (SEQ ID NO. 37), or (SEQ ID NO: 39), or operative derivative of any of these, and said derivative being a derivative that does not fully eliminate promoter activity that is inducible by a cytokine selected from the group consisting of LIF, IL-6, and IL-11.  
     
     
         8 . The nucleic acid construct of  claim 2 , further comprising at least one insulator element flanking said transcriptional unit.  
     
     
         9 . The nucleic acid construct of  claim 8 , wherein said at least one insulator element is a chicken β-globin insulator element.  
     
     
         10 . A kit for genetically modifying a vertebrate cell, containing a polynucleotide comprising a murine SOCS-3 promoter sequence having SEQ ID NO: 1, an operative fragment of SEQ ID NO: 1 having promoter activity that is inducible by a cytokine selected from the group consisting of LIF, IL-6, and IL-11, or an operative derivative of either of these, said derivative being selected from the group consisting of a mutation, a polymorphism, and a variant allele with respect to any nucleotide position of SEQ ID NO: 1, and said derivative being a derivative that does not fully eliminate promoter activity that is inducible by a cytokine selected from the group consisting of LIF, IL-6, and IL-11, and a suitable storage means for said polynucleotide.  
     
     
         11 . The kit of  claim 10 , further comprising said SOCS-3 promoter operatively linked, in a transcriptional unit, to a DNA sequence encoding a protein of interest.  
     
     
         12 . The kit of  claim 11 , wherein the protein of interest is a SOCS-3 protein, or a functional fragment thereof.  
     
     
         13 . The kit of  claim 11 , wherein said SOCS-3 promoter is operatively linked to a DNA sequence encoding a reporter protein.  
     
     
         14 . The kit of  claim 13 , wherein the reporter protein is a fluorescent or light-emitting protein.  
     
     
         15 . The kit of  claim 14 , wherein the fluorescent or light-emitting protein is a green fluorescent protein, yellow fluorescent protein, blue fluorescent protein, phycobiliprotein, luciferase, or apoaequorin.  
     
     
         16 . The kit of  claim 11 , wherein said polynucleotide further comprises at least one insulator element flanking said transcriptional unit.  
     
     
         17 . The kit of  claim 16 , wherein said at least one insulator element is a chicken β-globin insulator element.  
     
     
         18 . The kit of  claim 10 , further comprising a transfecting agent.  
     
     
         19 . The kit of  claim 18 , wherein the transfecting agent is a liposome, viral vector, transferrin-polylysine enhanced viral vector, retroviral vector, lentiviral vector, or uptake enhancing DNA segment, or a mixture of any of these.  
     
     
         20 . The kit of  claim 18 , wherein the transfecting agent comprises a retroviral vector, adenoviral vector, transferrin-polylysine enhanced adenoviral vector, human immunodeficiency virus vector, lentiviral vector, Moloney murine leukemia virus-derived vector, mumps vector, a DNA segment that facilitates polynucleotide uptake by and release into the cytoplasm of vertebrate cells, or comprises an operative fragment of or mixture of any of these.  
     
     
         21 . The kit of  claim 18 , wherein the transfecting agent comprises an adenovirus vector having endosomal lytic activity, and the polynucleotide is operatively linked to the vector.  
     
     
         22 . The kit of  claim 18 , wherein the transfecting agent comprises a lipid transfecting agent.  
     
     
         23 . A nucleic acid construct, comprising a murine SOCS-3 promoter having a nucleotide sequence selected from the group consisting of (SEQ ID NO: 1), (SEQ ID NO: 2), (SEQ ID NO: 3), (SEQ ID NO: 4), (SEQ ID NO: 5), (SEQ ID NO: 6), (SEQ ID NO: 7), (SEQ ID NO: 8), (SEQ ID NO: 9), (SEQ ID NO: 10), (SEQ ID NO: 11), (SEQ ID NO: 12), (SEQ ID NO: 35), (SEQ ID NO: 36), (SEQ ID NO: 37), and (SEQ ID NO: 39), or an operative derivative of any of these selected from the group consisting of a mutation, a polymorphism, and a variant allele with respect to any nucleotide position of (SEQ ID NO: 1), (SEQ ID NO: 2), (SEQ ID NO: 3), (SEQ ID NO: 4), (SEQ ID NO: 5), (SEQ ID NO: 6), (SEQ ID NO: 7), (SEQ ID NO: 8), (SEQ ID NO: 9), (SEQ ID NO: 10), (SEQ ID NO: 11), (SEQ ID NO: 12), (SEQ ID NO: 35), (SEQ ID NO: 36), (SEQ ID NO: 37), or (SEQ ID NO: 39), said derivative being a derivative that does not fully eliminate promoter activity that is inducible by a cytokine selected from the group consisting of LIF, IL-6, and IL-11.  
     
     
         24 . The nucleic acid construct of  claim 23 , further comprising said SOCS-3 promoter operatively linked, in a transcriptional unit, to a DNA sequence encoding a protein of interest.  
     
     
         25 . The nucleic acid construct of  claim 24 , wherein the protein of interest is a SOCS-3 protein.  
     
     
         26 . The nucleic acid construct of  claim 24 , wherein said polynucleotide further comprises at least one insulator element flanking said transcriptional unit.  
     
     
         27 . The nucleic acid construct of  claim 26 , wherein said at least one insulator element is a chicken β-globin insulator element.  
     
     
         28 . The nucleic acid construct of  claim 24 , wherein said SOCS-3 promoter is operatively linked to a DNA sequence encoding a reporter protein.  
     
     
         29 . The nucleic acid construct of  claim 28 , wherein the reporter protein is a fluorescent or light-emitting protein.  
     
     
         30 . The nucleic acid construct of  claim 29 , wherein the fluorescent or light-emitting protein is a green fluorescent protein, yellow fluorescent protein, blue fluorescent protein, phycobiliprotein, luciferase, or apoaequorin.