Methods of use of viral vectors to deliver antigen to dendritic cells

Abstract

This invention relates to methods and compositions useful for delivering antigens to dendritic cells which are then useful for inducing T antigen specific cytotoxic T lymphocytes. This invention also provides assays for evaluating the activity of cytotoxic T lymphocytes. According to the invention, antigens are provided to dendritic cells using a viral vector such as influenza virus which may be modified to express non-native antigens for presentation to the dendritic cells. The dendritic cells which are infected with the vector are then capable of presenting the antigen and inducing cytotoxic T lymphocyte activity or may also be used as vaccines.

We claim:  
     
         1 . A method of delivering antigens to dendritic cells comprising: 
 a. providing a viral vector comprising a gene sequence encoding for said antigen; and    b. exposing said dendritic cells to said viral vector for a time sufficient to allow said antigen to be expressed on the surface of said dendritic cells.    
     
     
         2 . The method according to  claim 1  wherein the dendritic cells are human.  
     
     
         3 . The method according to  claim 1 , wherein said vector is an influenza viral vector.  
     
     
         4 . The method according to  claim 1 , wherein said vector comprises recombinant nucleic acid.  
     
     
         5 . The method according to  claim 1  wherein said antigen is selected from the group consisting of tumor antigens, viral antigens, microbial antigens and autoimmune antigens.  
     
     
         6 . The method according to  claim 5  wherein said antigen comprises amino acid sequences from at least one strain of influenza virus.  
     
     
         7 . The method according to  claim 1  wherein acid dendritic cells are proliferating.  
     
     
         8 . The method according to  claim 1  wherein said dendritic cells are non-proliferating.  
     
     
         9 . The method according to  claim 2  wherein said influenza virus is PR8.  
     
     
         10 . The method according to  claim 2  wherein said vector is a replicating influenza virus.  
     
     
         11 . The method according to  claim 2  wherein said vector is a non-replicating influenza virus.  
     
     
         12 . The method according to  claim 1  wherein said dendritic cells are mature.  
     
     
         13 . The method according to  claim 2  wherein said antigen comprises amino acid sequences from at least one strain of influenza virus.  
     
     
         14 . A method of generating antigen specific cytotoxic T lymphocytes, comprising: 
 a. providing a viral vector comprising a nucceic acid sequence encoding for said antigen;    b. exposing dendritic cells to said viral vector for a time sufficient to allow said antigen to be expressed on the surface of said dendritic cells; and    c. exposing T lymphocytes precursor to said dendritic cells for a sufficient time to induce them to become activated to antigen specific T lymphocytes.    
     
     
         15 . The method according to  claim 12  wherein said viral vector is an influenza viral vector.  
     
     
         16 . The method according to  claim 1  wherein the ratio of vector to dendritic cells is in the range of about 1 to 100 vectors per dendritic cell.  
     
     
         17 . The method according to  claim 9  further comprising administering said antigen specific cytotoxic killer T lymphocytes to an individual afflicted with a disease.  
     
     
         18 . An antigen presenting dendritic cell prepared according to the method of  claim 1 .  
     
     
         19 . The antigen presenting dendritic cells according to  claim 18  wherein said dendritic cells are infected with influenza virus.  
     
     
         20 . The method according to  claim 19  wherein the dendritic cells are human.  
     
     
         21 . The antigen presenting dendritic cells according to  claim 20  wherein the influenza virus is nonreplicating.  
     
     
         22 . The antigen presenting dendritic cells according to  claim 20  the influenza virus is replicating.  
     
     
         23 . Cytotoxic T lymphocytes prepared according to the method of  claim 14 .  
     
     
         24 . The cytotoxic T lymphocytes according to  claim 23  wherein cytotoxic T lymphocyte precursors are exposed to antigen presenting dendritric cells infected with influenza virus.  
     
     
         25 . The cytotoxic T lymphocytes according to  claim 24  wherein the antigen presenting dendritic cells are human.  
     
     
         26 . The cytotoxic T lymphocytes according to  claim 25  wherein the T lymphocytes exposed to antigen presenting dendritic cells are human.  
     
     
         27 . A method of assessing cytotoxic T lymphocyte activity comprising: 
 a. providing antigen presenting dendritic cells prepared according to method of  claim 1;     b. exposing the-antigen presenting dendritic cells to population of T lymphocytes to be assayed for their ability to exhibit killer cell activity; and    c. assaying the cytotoxic activity of the T lymphocytes exposed to the antigen presenting dendritic cells.    
     
     
         28 . The method according to  claim 27  wherein the antigen presented by the dendritic cells is a tumor antigen and the T lymphocytes are assayed for their cytotoxic activity against tumor cells.  
     
     
         29 . The method according to  claim 27  wherein the antigen presented by the dendritic cells is a viral antigen and the T lymphocytes are assayed for their activity against viral infected cells.