Modified heat shock protein-antigenic peptide complex

Abstract

The present invention relates to methods for purifying immunogenic, prophylactically and therapeutically effective complexes of modified heat shock proteins noncovalently associated with antigenic peptides of cancer or infected cells. The claimed methods comprise the constructing of a nucleotide sequence encoding a secretable modified heat shock protein, expressing the sequence in an appropriate host cell, recovering the immunogenic complexes from the cell culture and the cells, and purifying the immunogenic complexes by affinity chromatography. Large amounts of such immunogenic complexes can be obtained by large-scale culturing of host cells containing the genetic sequence. The complexes can be used as a vaccine to elicit specific immune responses against cancer or infected cells, and to treat or prevent cancer or infectious diseases.

1 - 73 . (canceled) 
     
     
         74 . An isolated human cancer cell transfected with a construct which expresses a modified heat shock protein in the cell, wherein the modified heat shock protein: (i) is secreted by the cell and is non-covalently associated with an antigenic peptide; (ii) lacks an endoplasmic reticulum retention sequence present in an unmodified heat shock protein; and (iii) comprises a peptide tag. 
     
     
         75 . The isolated human cancer cell of  claim 74 , wherein the modified heat shock protein further comprises a leader peptide not present in the unmodified heat shock protein. 
     
     
         76 . The isolated human cancer cell of  claim 74 , wherein the unmodified heat shock protein is human gp96. 
     
     
         77 . The isolated human cancer cell of  claim 74 , wherein the cell is a lung carcinoma cell. 
     
     
         78 . The isolated human cancer cell of  claim 74 , wherein the cell is a small cell lung carcinoma cell. 
     
     
         79 . The isolated human cancer cell of  claim 74 , wherein the cell is a bladder carcinoma cell. 
     
     
         80 . The isolated human cancer cell of  claim 74 , wherein the peptide tag is selected from the group consisting of one or more constant regions of an immunoglobulin, a polyhistidine tag, glutathione S-transferase, maltose binding protein, a Myc tag, and a FLAG epitope. 
     
     
         81 . The isolated cancer cell of  claim 74 , wherein the peptide tag comprises the CH2 and CH3 domains of an immunoglobulin heavy chain constant region. 
     
     
         82 . The isolated cancer cell of  claim 74 , wherein the peptide tag comprises the hinge, CH2 and CH3 domains of an immunoglobulin heavy chain constant region. 
     
     
         83 . A method for eliciting an immune response to a tumor antigen in an individual having a cancer and in need of treatment for the cancer, the method comprising administering to the individual a composition comprising a human cancer cell comprising the tumor antigen; wherein:
 (a) the human cancer cell comprises an expression vector comprising a nucleic acid sequence that encodes a modified heat shock protein; and   (b) the encoded modified heat shock protein: (i) is secreted by the human cancer cell; (ii) lacks an endoplasmic reticulum retention sequence present in the unmodified heat shock protein; and (iii) comprises a peptide tag.   
     
     
         84 . The method of  claim 83 , wherein the secreted heat shock protein is non-covalently associated with an antigenic peptide derived from the tumor antigen as a modified heat shock protein-antigenic peptide complex. 
     
     
         85 . The method of  claim 83 , wherein the encoded modified heat shock protein comprises a leader peptide not present in the unmodified heat shock protein. 
     
     
         86 . The method of  claim 83 , wherein the unmodified heat shock protein is human gp96. 
     
     
         87 . The method of  claim 83 , wherein the human cancer cell is a lung carcinoma cell. 
     
     
         88 . The method of  claim 83 , wherein the human cancer cell is a small cell lung carcinoma cell. 
     
     
         89 . The method of  claim 83 , wherein the human cancer cell is a bladder carcinoma cell. 
     
     
         90 . The method of  claim 83 , wherein the peptide tag is selected from the group consisting of one or more constant regions of an immunoglobulin heavy chain, a polyhistidine tag, glutathione S-transferase, maltose binding protein, a Myc tag, and a FLAG epitope. 
     
     
         91 . The method of  claim 83 , wherein the peptide tag comprises the CH2 and CH3 domains of an immunoglobulin heavy chain constant region. 
     
     
         92 . The method of  claim 83 , wherein the peptide tag comprises the hinge, CH2 and CH3 domains of an immunoglobulin heavy chain constant region. 
     
     
         93 . The method of  claim 83 , wherein the nucleic acid sequence encoding the modified heat shock protein is operably linked to at least one regulatory region that controls expression of the modified heat shock protein nucleic acid sequence in the cancer cell. 
     
     
         94 . The method of  claim 83 , wherein the nucleic acid sequence encoding the modified heat shock protein is expressed by a vector based on bovine papilloma virus. 
     
     
         95 . The method of  claim 83 , wherein the human cancer cell is obtained from an individual other than the one to whom composition is administered. 
     
     
         96 . The method of  claim 83 , wherein the composition, when administered to the individual, is effective for causing activation of a CD4-independent, CD8 T lymphocyte response against the tumor antigen. 
     
     
         97 . The method of  claim 84 , wherein the human cancer cell has been irradiated to inhibit its proliferation, while maintaining its ability to secrete the modified heat shock protein-antigenic peptide complexes for 7 to 10 days. 
     
     
         98 . The method of  claim 83 , wherein the human cancer cell is allogeneic to the individual. 
     
     
         99 . The method of  claim 83 , wherein the human cancer cell is administered to the individual by subcutaneous administration.