Novel targets and compositions for use in decontamination, immunoprophylaxis, and post-exposure therapy against anthrax

Abstract

The present invention relates to the decontamination of anthrax spores, prophylaxis and treatment of anthrax infections and, more particularly, to compounds that act as specific inhibitors of B. anthracis germination/outgrowth-associated proteins, methods and means for making such inhibitors and their use as pharmaceuticals and/or vaccines. The invention also relates to the prophylaxis and treatment of anthrax infections and, more particularly, to vaccines and compositions that comprise B. anthracis antigens, epitopes, proteins, or nucleic acid molecules, including anthrax protective antigen, anthrax lethal factor, anthrax edema factor and anthrax proteins associated with spore germination and outgrowth, as well as methods and means for making such compositions and their use pharmaceuticals and/or vaccines.

1 . A  Bacillus anthracis  germination/outgrowth-associated spore protein corresponding to: 
 (i) Spot  12  (Accession # NP — 843763), wherein the protein has a peptide fingerprint mass spectra as depicted in  FIG. 3A ;    (ii) Spot  13  (Accession #: NP — 655179), wherein the protein has a peptide fingerprint mass spectra as depicted in  FIG. 3B ;    (iii) Spot  19  (Accession # NP — 846769), wherein the protein has a peptide fingerprint mass spectra as depicted in  FIG. 3C ; or    (iv) Spot  15  (Accession # NP — 655132), wherein the protein has a peptide fingerprint mass spectra as depicted in  FIG. 3D .    
     
     
         2 . A  Bacillus anthracis  germination/outgrowth-associated spore protein that is up- or down-regulated during germination by at least 30%, wherein the protein corresponds to Spot  21  (Accession # NP — 654182),  24  (Accession # NP — 846918),  27  (Accession # NP — 654198),  41  (Accession # NP — 847705),  47  (Accession # NP — 653587), or  48  (Accession # NP — 846171).  
     
     
         3 . Novel  Bacillus anthracis  spore proteins corresponding to Spots  17  (Accession # NP — 657537),  18  (Accession # NP — 658120),  20  (Accession # NP — 653598),  28  (Accession # NP — 242212),  49  (Accession # NP — 654492),  50  (Accession # NP — 654728).  
     
     
         4 . A method of immunization in an animal and/or a method of inducing a systemic immune response or systemic therapeutic response to a gene product, in an animal, comprising administering a vector that contains and expresses a nucleic acid molecule encoding the gene product, in an amount effective to induce the response, wherein the immunization and/or systemic immune response and/or systemic therapeutic response affords the animal protection against challenge with  B. anthracis.    
     
     
         5 . A method of immunization in an animal and/or a method of inducing a systemic immune response or systemic therapeutic response to a gene product, in an animal, comprising administering a vector that contains and expresses a nucleic acid molecule encoding the gene product, in an amount effective to induce the response, wherein the nucleic acid molecule encodes the  Bacillus anthracis  spore protein of any of claims  1 - 3  or a portion thereof alone or in combination.  
     
     
         6 . A vaccine or immunogenic composition against  Bacillus anthracis  comprising a spore protein as described in any of claims  1 - 3 , or a fragment thereof, or a genetic vector encoding a spore protein as described in any of claims  1 - 3 , or a fragment thereof, and optionally an excipient and/or adjuvant and/or a suitable carrier or diluent.  
     
     
         7 . A method of preparing the vaccine or immunogenic composition of  claim 6 .  
     
     
         8 . A method of treating an animal after suspected or real exposure to  B. anthracis  comprising administering an effective amount of one or more inhibitors to one or more of the spore proteins of claims  1 - 3 .  
     
     
         9 . The method of any of claims  4 ,  5  or  8 , wherein the animal is a vertebrate.  
     
     
         10 . The method of  claim 9  wherein the vertebrate is a mammal.  
     
     
         11 . The method of  claim 10  wherein the mammal is a human or a companion or domesticated or food- or feed-producing or livestock or game or racing or sport animal.  
     
     
         12 . A method of immunization, therapy, or decontamination of anthrax spores by inducing pre-mature spore germination with agents targeting the spore proteins of claims  1 - 3 .  
     
     
         13 . The  Bacillus anthracis  spore protein of any of claims  1 - 3  wherein the protein is obtained by a process comprising the steps of expressing the protein in a vector and purifying the protein.  
     
     
         14 . The spore protein of  claim 1 , wherein the protein comprises the sequences depicted in  FIG. 3A ,  FIG. 3B ,  FIG. 3C  or  FIG. 3D .  
     
     
         15 . A method of immunization in an animal and/or a method of inducing a systemic immune response or systemic therapeutic response to any of the 31  B. anthracis  spore proteins described in Table 1.  
     
     
         16 . A method of non-invasive immunization in an animal and/or a method of inducing a systemic immune response or systemic therapeutic response to a gene product from  Bacillus anthracis , in an animal, comprising contacting skin, or nasal, oral, perlingual or buccal mucosa of the animal with a vector that contains and expresses a nucleic acid molecule encoding the gene product, in an amount effective to induce the response.  
     
     
         17 . The method of  claim 16  wherein the method comprises contacting the skin of the animal with the vector.  
     
     
         18 . The method of  claim 16  wherein the method comprises contacting the nasal mucosa with the vector.  
     
     
         19 . The method of  claim 16  wherein the method comprises contacting the oral or buccal or perlingual mucosa with the vector.  
     
     
         20 . The method of  claim 16  wherein the nucleic acid molecule is exogenous or heterologous to the vector.  
     
     
         21 . The method of  claim 16  wherein the response comprises a systemic immune response.  
     
     
         22 . The method of  claim 16  wherein the vector comprises and expresses an exogenous nucleic acid molecule encoding an epitope of interest.  
     
     
         23 . The method of  claim 16  wherein the vector comprises and expresses an antigen.  
     
     
         24 . The method of  claim 16  wherein the vector comprises and expresses a therapeutic product.  
     
     
         25 . The method of  claim 16  wherein the nucleic acid molecule encodes an epitope of interest and/or an antigen of interest and/or a nucleic acid molecule that stimulates and/or modulates an immunological response and/or stimulates and/or modulates expression comprising transcription and/or translation an endogenous and/or exogenous nucleic acid molecule.  
     
     
         26 . The method of  claim 20  wherein the exogenous nucleic acid molecule encodes one or more of an antigen or portion thereof, or one or more of an epitope of interest, from  Bacillus anthracis.    
     
     
         27 . The method of  claim 20  wherein the exogenous nucleic acid molecule encodes one or more of: anthrax protective antigen, anthrax lethal factor, and anthrax edema factor.  
     
     
         28 . The method of  claim 25  wherein the response is induced by the vector expressing the nucleic acid molecule in the animal's cells.  
     
     
         29 . The method of  claim 28  wherein the cells comprise epidermal cells.  
     
     
         30 . The method of  claim 25  wherein the response comprises an immune response against  Bacillus anthracis.    
     
     
         31 . The method of claims  16  wherein the animal is a vertebrate.  
     
     
         32 . The method of  claim 31  wherein the vertebrate is a mammal.  
     
     
         33 . The method of  claim 32  wherein the mammal is a human or a companion or domesticated or food- or feed-producing or livestock or game or racing or sport animal.  
     
     
         34 . The method of  claim 33  wherein the animal is a cow, a horse, a dog, a cat, a goat, a sheep, or a pig.  
     
     
         35 . The method of  claim 16  wherein the vector comprises one or more of a viral vector, a viral coat comprising a virus having some or all viral genes deleted therefrom, a bacterial vector, protozoan vector, a transposon, a retrotransposon, and a DNA vector.  
     
     
         36 . The method of  claim 35  wherein the vector comprises a recombinant vector.  
     
     
         37 . The method of  claim 36  wherein the vector comprises an adenovirus.  
     
     
         38 . The method of  claim 37  wherein the adenovirus is defective or deleted in its E1 and/or E3 and/or E4 region(s).  
     
     
         39 . The method of  claim 35 , wherein the vector is an  Escherichia  bacterial vector.  
     
     
         40 . The method of  claim 39  wherein the  Escherichia  vector is  Escherichia coli.    
     
     
         41 . The method of  claim 40  wherein the  Escherichia coli  is replication defective.  
     
     
         42 . The method of  claim 41  wherein the  Escherichia coli  is rendered replication defective by irradiation, antibiotics, fixatives, genetic substitutions or deletions, or gentle heat.  
     
     
         43 . A method of inducing a systemic immune response or systemic therapeutic response to a gene product from  Bacillus anthracis , in an animal, comprising topical and/or intranasal and/or mucosal and/or buccal and/or perlingual and/or oral administration of a vector comprising: 
 (i) an adenovirus defective in its E1 and/or E3 and/or E4 region(s), wherein the adenovirus contains and expresses a nucleic acid molecule encoding the gene product for the response; or,    (ii) replication defective  Escherichia coli , wherein the vector contains and expresses a nucleic acid molecule encoding the gene product for the response.    
     
     
         44 . The method of  claim 43  comprising topical administration.  
     
     
         45 . The method of  claim 43  comprising intranasal administration.  
     
     
         46 . The method of  claim 43  wherein the adenovirus is defective or deleted in its E1 and E3 regions.  
     
     
         47 . The method of claims  43  wherein the vector comprises an exogenous or heterologous nucleic acid molecule encoding the gene product for the response.  
     
     
         48 . The method of  claim 43  wherein the nucleic acid molecule is exogenous or heterologous and encodes an epitope of interest and the method is for inducing a systemic immunological response.  
     
     
         49 . The method of  claim 43  wherein the adenovirus is a human adenovirus.  
     
     
         50 . The method of  claim 43  wherein the adenovirus comprises a non-human adenovirus.  
     
     
         51 . The method of  claim 16  or  43  wherein the vector is matched to, or a natural pathogen of, the animal.  
     
     
         52 . The method of  claim 16  or  43  comprising applying a delivery device including the vector to the skin of the animal.  
     
     
         53 . The method of  claim 52  further comprising disposing the vector in and/or on the delivery device.  
     
     
         54 . A pharmaceutical, vaccine, immunogenic, immunological or therapeutic composition for use in a method according to any one of claims  16  or  43  comprising the vector in a pharmaceutically acceptable carrier or diluent for administering the vector topically or mucosally or intranasally or buccally or perlingually or orally.  
     
     
         55 . A kit for preparing a pharmaceutical, vaccine, immunogenic, immunological or therapeutic composition for use in a method according to any one of claims  16  or  43  comprising in a first container the vector and in a second container a pharmaceutically acceptable carrier or diluent for administering the vector topically or mucosally or intranasally or bucally or perlingually or orally; wherein the containers are optionally present in the same package or are in separate packages; and, the kit optionally contains instructions for admixture of the vector and the carrier or diluent and/or administration.  
     
     
         56 . The kit of  claim 55  further comprising a delivery device in a third container; wherein the third container is optionally present in the same package as one or both of the first and second containers or is in a package separate from the first and second containers; and, wherein kit optionally contains instructions for installing the vector or vector and carrier or diluent into or onto the delivery device and/or for administration or application of the delivery device.  
     
     
         57 . The method of  claim 16  or  43 , wherein application of the vector to the skin is performed in combination with the use of microneedles.