Incapacitated whole-cell immunogenic bacterial compositions produced by recombinant expression

Abstract

The present invention features incapacitated whole-cell bacterial immunogenic compositions and methods of their production, which compositions are useful to deliver antigens in a manner resembling the live infectious organism in terms of elicitation of a robust immune response, but with reduced risk or no risk of disease. The compositions of the invention are produced by rendering a bacterium bacteriostatic through expression of a recombinant promoter in the bacterial cell, which promoter can be operably linked to a polynucleotide encoding a recombinant gene product. In one embodiment, where the bacterium is a gram negative host, the recombinant gene product provides for reduced toxicity of LPS. In one embodiment, the gene product is a bacteriophage protein, such as endolysin, holin, or ndd.

1 . A method of eliciting an immune response to a bacterial pathogen, the method comprising: 
 administering an incapacitated whole cell bacterial composition to a subject susceptible to a disease caused by a pathogenic bacterium, wherein the composition comprises a bacterium incapacitated by expression from a recombinant promoter, said expression being sufficient to incapacitate the bacterium; said administering being in an amount effective to elicit an immune response to the pathogenic bacterium in the host.    
     
     
         2 . The method of  claim 1 , wherein the pathogenic bacterium is of a genus selected from the group consisting of  Mycobacteria, Staphylococci, Vibrio, Enterobacter, Enterococcus, Escherichia, Haemophilus, Neisseria, Pseudomonas, Shigella, Serratia, Salmonella, Streptococcus, Klebsiella  and  Yersinia.    
     
     
         3 . The method of  claim 1 , wherein the recombinant promoter is operably linked to a recombinant polynucleotide encoding a recombinant protein other than a viral protein of a virus that infects a mammalian cell.  
     
     
         4 . The method of  claim 3 , wherein the recombinant protein is a protein that binds lipopolysaccharide, including a lipopolysaccharide-binding protein (LBP) or an LPS-binding domain thereof.  
     
     
         5 . The method of  claim 1 , wherein the recombinant promoter is a strong bacteriophage promoter.  
     
     
         6 . The method of  claim 5 , wherein the bacterium is further modified to express a bacteriophage RNA polymerase for transcription from the bacteriophage promoter.  
     
     
         7 . The method of  claim 6 , wherein the bacteriophage RNA polymerase is operably linked to an inducible promoter.  
     
     
         8 . The method of  claim 7 , wherein the bacteriophage promoter is a T7 promoter and the bacteriophage RNA polymerase is a T7 RNA polymerase.  
     
     
         9 . A method of vaccinating a subject against disease caused by a bacterial pathogen, the method comprising: 
 administering to a subject susceptible to disease caused by a pathogenic bacterium an incapacitated whole cell bacterial vaccine, the vaccine comprising the pathogenic bacterium incapacitated by expression from a recombinant promoter, said administering being in an amount effective to elicit an immune response to the pathogenic bacterium in the subject.    
     
     
         10 . The method of  claim 9 , wherein the recombinant promoter is operably linked to a recombinant polynucleotide encoding a recombinant protein other than a viral protein of a virus that infects a mammalian cell.  
     
     
         11 . The method of  claim 10 , wherein the recombinant protein is a protein that binds lipopolysaccharide.  
     
     
         12 . The method of  claim 11 , wherein the protein comprises a lipopolysaccharide-binding protein (LBP) or an LPS-binding domain thereof.  
     
     
         13 . The method of  claim 9 , wherein the recombinant promoter is a strong bacteriophage promoter and the bacterium is further modified to express a bacteriophage RNA polymerase for transcription from the bacteriophage promoter.  
     
     
         14 . The method of  claim 13 , wherein the bacteriophage RNA polymerase is operably linked to an inducible promoter.  
     
     
         15 . The method of  claim 14 , wherein the bacteriophage promoter is a T7 promoter and the bacteriophage RNA polymerase is a T7 RNA polymerase.  
     
     
         16 . A method for eliciting an immune response to an antigen, the method comprising: 
 administering to a subject an incapacitated whole cell bacterial composition, wherein the composition comprises a bacterium incapacitated by expression from a recombinant promoter, said administering being in an amount effective to elicit an immune response in the subject to an antigen present in or on the bacterium.    
     
     
         17 . The method of  claim 16 , wherein: 
 (a) the recombinant promoter is operably linked to a recombinant polynucleotide encoding a recombinant protein other than a viral protein of a virus that infects a mammalian cell, including wherein the recombinant protein is the antigen;    (b) the antigen is an endogenous bacterial antigen, including wherein the recombinant protein is a protein that binds lipopolysaccharide, such as a lipopolysaccharide-binding protein (LBP) or an LPS-binding domain thereof; or    (c) the antigen is an endogenous bacterial antigen.    
     
     
         18 . The method of  claim 17 , wherein the recombinant promoter is a strong bacteriophage promoter and the bacterium is further modified to express a bacteriophage RNA polymerase for transcription from the bacteriophage promoter.  
     
     
         19 . The method of  claim 18 , wherein the bacteriophage RNA polymerase is operably linked to an inducible promoter.  
     
     
         20 . The method of  claim 19 , wherein the bacteriophage promoter is a T7 promoter and the bacteriophage RNA polymerase is a T7 RNA polymerase.