Cc chemokine receptor 5 dna, new animal models and therapeutic agents for hiv infection

Abstract

The susceptibility of human macrophages to human immunodeficiency virus (HIV) infection depends on cell surface expression of the human CD4 molecule and CC cytokine receptor 5. CCR5 is a member of the 7-transmembrane segment superfamily of G-protein-coupled cell surface molecules. CCR5 plays an essential role in the membrane fusion step of infection by some HIV isolates. The establishment of stable, nonhuman cell lines and transgenic mammals having cells that coexpress human CD4 and CCR5 provides valuable tools for the continuing research of HIV infection. In addition, antibodies which bind to CCR5, CCR5 variants, and CCR5-binding agents, capable of blocking membrane fusion between HIV and target cells represent potential anti-HIV therapeutics for macrophage-tropic strains of HIV.

1 . A method of inhibiting membrane fusion between HIV and a target cell that expresses CCR5 or between an HIV-infected cell and a CD4 positive uninfected cell that expresses CCR5, comprising contacting the target or CD4 positive cell with a fusion-inhibiting effective amount of a CCR5 binding or blocking agent. 
     
     
         2 . The method of  claim 1 , wherein the agent is an anti-CCR5 antibody or epitope binding fragment thereof. 
     
     
         3 . The method of  claim 2 , wherein the antibody is a monoclonal antibody or a polyclonal antibody. 
     
     
         4 . The method of  claim 1 , wherein the contacting is by in vivo administration to a subject. 
     
     
         5 . The method of  claim 2 , wherein the anti-CCR5 antibody is administered by intravenous, intramuscular or subcutaneous injections. 
     
     
         6 . The method of  claim 4 , wherein the anti-CCR5 antibody is administered within a dose range of 0.1 μg/kg to 100 mg/kg. 
     
     
         7 . The method of  claim 5 , wherein the antibody is formulated in a pharmaceutically acceptable carrier. 
     
     
         8 . A method for identifying a compound which blocks HIV-envelope mediated membrane fusion, comprising:
 a) incubating components comprising the compound and a CCR5 polypeptide under conditions sufficient to allow the components to interact; and   b) measuring the binding of the compound to CCR5 polypeptide.   
     
     
         9 . The method of  claim 8 , wherein the compound is a peptide. 
     
     
         10 . The method of  claim 8 , wherein the compound is a peptidomimetic. 
     
     
         11 . The method of  claim 8 , wherein the CCR5 polypeptide is expressed in a cell. 
     
     
         12 . The method of  claim 11  wherein the cell is a recombinant cell line that expresses CCR5 polypeptide. 
     
     
         13 . The method of  claim 8 , wherein the compound comprises an antibody. 
     
     
         14 . The method of  claim 13 , wherein the antibody comprises a polyclonal antibody, monoclonal antibody, or fragment thereof. 
     
     
         15 . The method of  claim 13 , wherein the antibody is specific for an extracellular region of the CCR5 polypeptide. 
     
     
         16 . The method of  claim 13 , wherein the antibody comprises a humanized monoclonal antibody. 
     
     
         17 . The method of  claim 8 , wherein the compound comprises an antisense molecule. 
     
     
         18 . The method of  claim 8 , wherein the method further comprises:
 (c) incubating components comprising the compound with a CCR5 positive cell under conditions sufficient to allow the components to interact with the CCR5 positive cell;   (d) contacting the components of (c) with HIV or an HIV-infected cell; and   (e) measuring the ability of the compound to block membrane fusion between the HIV and the CCR5 positive cell or between the HIV-infected cell and the CCR5 cell.   
     
     
         19 . The method of  claim 8 , wherein the method further comprises:
 (c) incubating a first eukaryotic cell expressing CCR5 and CD4 with a second eukaryotic cell expressing an HIV envelope protein under conditions that permit fusion of the first eukaryotic and second eukaryotic cells;   (d) contacting components comprising the compound with the first eukaryotic and second eukaryotic cells; and   (e) detecting fusion between the first eukaryotic and second eukaryotic cells.   
     
     
         20 . The method of  claim 19 , wherein contacting components comprising the compound with the first eukaryotic and second eukaryotic cells is performed before the first eukaryotic and second eukaryotic cells are incubated. 
     
     
         21 . The method of  claim 19 , wherein contacting components comprising the compound with the first eukaryotic and second eukaryotic cells is performed after the first eukaryotic and second eukaryotic cells are incubated. 
     
     
         22 . A method for identifying a compound which blocks HIV-envelope mediated membrane fusion, comprising:
 incubating components comprising the compound with a CCR5 positive cell under conditions sufficient to allow the components to interact with the CCR5 positive cell;   contacting the components and the CCR5 positive cells with HIV or an HIV-infected cell; and   measuring the ability of the compound to block membrane fusion between the HIV and the CCR5 positive cell or between the HIV-infected cell and the CCR5 cell.   
     
     
         23 . The method of  claim 22 , wherein the CCR5 positive cell expresses CD4. 
     
     
         24 . A method for identifying a compound which blocks HIV-envelope mediated membrane fusion, comprising:
 incubating a first eukaryotic cell expressing CCR5 and CD4 with a second eukaryotic cell expressing an HIV envelope protein under conditions that permit fusion of the first eukaryotic and second eukaryotic cells;   contacting components comprising the compound with the first eukaryotic and second eukaryotic cells; and   detecting fusion between the first eukaryotic and second eukaryotic cells.   
     
     
         25 . A method of treating a subject having an HIV-related disorder associated with expression of CCR5 comprising administering to an HIV infected or susceptible cell of the subject, an agent that suppresses CCR5. 
     
     
         26 . The method of  claim 25 , wherein the agent is an anti-CCR5 antibody. 
     
     
         27 . The method of  claim 25 , wherein the agent is an antisense nucleic acid that hybridizes to a CCR5 nucleic acid. 
     
     
         28 . The method of  claim 25 , wherein the agent is introduced into the cell using a carrier. 
     
     
         29 . The method of  claim 25 , wherein the carrier is a vector. 
     
     
         30 . The method of  claim 25 , wherein the administering is ex vivo. 
     
     
         31 . The method of  claim 25 , wherein the administering is in vivo.