Novel polynucleotides and polypeptides in pathogenic mycobacteria and their use as diagnostics, vaccines and targets for chemotherapy

Abstract

The invention provides a nucleotide sequence representing a pathogenicity island found in species of pathogenic mycobacteria. The islands are shown as SEQ ID NOS: 3 and 4 and comprises several open reading frames encoding polypeptides. These polypeptides and their use in diagnosis and therapy form a further aspect of the invention.

1 - 23 . (Canceled).  
     
     
         24 . A substantially isolated polynucleotide comprising: 
 (a) a polynucleotide sequence according to any one of SEQ ID NOS: 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25 and 27;    (b) a polynucleotide capable of selectively hybridising to a polynucleotide of (a);    (c) a polynucleotide having at least 80% sequence homology to a polynucleotide of (a) over 30 contiguous amino acids;    (d) a polynucleotide encoding a polypeptide of any one of SEQ ID NOS: 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26 and 28; or    (e) a polynucleotide encoding a fragment of at least 10 amino acids of a polypeptide of any one of SEQ ID NOS: 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26 and 28;    wherein said polynucleotide encodes a polypeptide having the ability to stimulate an immune response against a mycobacterium.    
     
     
         25 . A substantially isolated polynucleotide according to  claim 24  comprising: 
 (a) a polynucleotide sequence according to SEQ ID NO: 23;  
 (b) a polynucleotide capable of selectively hybridising to a polynucleotide of SEQ ID NO: 23;  
 (c) a polynucleotide having at least 80% sequence homology to the polynucleotide of SEQ ID NO: 23 over 30 contiguous amino acids;  
 (d) a polynucleotide encoding the polypeptide of SEQ ID NO: 24; or  
 (e) a polynucleotide encoding a fragment of at least 10 amino acids of the polypeptide of SEQ ID NO: 24;  
 wherein said polynucleotide encodes a polypeptide having the ability to stimulate an immune response against a mycobacterium.  
 
     
     
         26 . A polynucleotide according to  claim 24  which further comprises a label.  
     
     
         27 . A vector carrying a polynucleotide according to  claim 24 .  
     
     
         28 . A vector according to  claim 27  which is an expression vector.  
     
     
         29 . A vector according to  claim 28  wherein said polynucleotide is operably linked to a control sequence which is capable of providing for the expression of the coding sequence of the polynucleotide.  
     
     
         30 . A vector according to  claim 27  which comprises one or more components selected from the group consisting of an origin of replication, a promoter for expression of the polypeptide encoded by said polynucleotide, a regulator of a promoter for expression of the polypeptide encoded by said polypeptide, an enhancer and a selectable marker gene.  
     
     
         31 . A vector according to  claim 30  wherein said promoter is a mammalian, viral, yeast or bacterial promoter.  
     
     
         32 . A vector according to  claim 31  wherein said promoter is selected from the group consisting of: a metallothionien promoter, an adenovirus promoter, the SV40 large T promoter, a retroviral LTR promoter, the polyhedrin promoter, an alcohol dehydrogenase promoter and a β-galactosidase promoter.  
     
     
         33 . A vector according to  claim 27  which is adapted for use in vivo.  
     
     
         34 . A vector according to  claim 27  which is a plasmid, virus or phage vector.  
     
     
         35 . A vector according to  claim 34  wherein said viral vector is selected from the group consisting of retroviral vectors, adenoviral vectors, adeno-associated viral vectors, vaccinia virus vectors, herpes virus vector and alpha virus vectors.  
     
     
         36 . A host cell comprising, transformed with or transfected by a vector according to  claim 27 .  
     
     
         37 . A host cell according to  claim 36  which is a bacterial, yeast, insect or mammalian cell.  
     
     
         38 . A host cell according to  claim 37  which is selected from the group consisting of  M. bovis  BCG,  M. smegmatis , a mycobacterium,  Corynebacteria  and  Salmonella.    
     
     
         39 . A pharmaceutical composition comprising a polynucleotide according to  claim 24  and a pharmaceutically acceptable carrier or diluent.  
     
     
         40 . A pharmaceutical composition comprising a vector according to  claim 27  and a pharmaceutically acceptable carrier or diluent.  
     
     
         41 . A pharmaceutical composition comprising a host cell according to  claim 36  and a pharmaceutically acceptable carrier or diluent.  
     
     
         42 . A method of raising an immune response in an animal or human against a mycobacterium, which method comprises administering an effective amount of a polynucleotide capable of expressing a polypeptide selected from: 
 (i) a polypeptide according to any one of SEQ ID NOS: 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28 and 29;    (ii) a polypeptide comprising a polypeptide according to (i);    (iii) a polypeptide having at least 70% amino acid identity to a polypeptide of (i) over 30 or more contiguous amino acids, which retains the ability to stimulate an immune response against said mycobacterium; or    (iv) a fragment of a polypeptide of (i) comprising at least 10 amino acids which retains the ability to stimulate an immune response against said mycobacterium to said human or animal and allowing said polypeptide to be expressed.    
     
     
         43 . A method according to claim  19  wherein said polynucleotide is selected from: 
 (a) a polynucleotide sequence according to any one of SEQ ID NOS: 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25 and 27;  
 (b) a polynucleotide capable of selectively hybridising to a polynucleotide of (a);  
 (c) a polynucleotide having at least 80% sequence homology to a polynucleotide of (a) over 30 contiguous amino acids;  
 (d) a polynucleotide encoding a polypeptide of any one of SEQ ID NOS: 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26 and 28; or  
 (e) a polynucleotide encoding a fragment of at least 10 amino acids of a polypeptide of any one of SEQ ID NOS: 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26 and 28.  
 
     
     
         44 . A method of raising an immune response in an animal or human against a mycobacterium, which method comprises administering an effective amount of a polynucleotide capable of expressing a polypeptide selected from: 
 (ii) a polypeptide according to SEQ ID NO: 24;    (ii) a polypeptide comprising a polypeptide according to (i);    (iii) a polypeptide having at least 70% amino acid identity to a polypeptide of (i) over 30 or more contiguous amino acids, which retains the ability to stimulate an immune response against said mycobacterium; or    (iv) a fragment of a polypeptide of (i) comprising at least 10 amino acids which retains the ability to stimulate an immune response against said mycobacterium to said human or animal and allowing said polypeptide to be expressed.    
     
     
         45 . A method according to claim  19  wherein said polynucleotide is selected from: 
 (a) a polynucleotide sequence according to SEQ ID NO: 23;  
 (b) a polynucleotide capable of selectively hybridising to a polynucleotide of SEQ ID NO: 23;  
 (c) a polynucleotide having at least 80% sequence homology to the polynucleotide of SEQ ID NO: 23 over 30 contiguous amino acids;  
 (d) a polynucleotide encoding the polypeptide of SEQ ID NO: 24; or  
 (e) a polynucleotide encoding a fragment of at least 10 amino acids of the polypeptide of SEQ ID NO: 24.  
 
     
     
         46 . A method according to  claim 42  wherein said polynucleotide is provided in a vector, operably linked to a control sequence which is capable of providing for the expression of said polypeptide from said vector.  
     
     
         47 . A method according to  claim 43  wherein said vector is a plasmid, virus or phage vector.  
     
     
         48 . A method of enhancing the response of an animal or human infected with a mycobacterium to treatment with an antimycobacterial drug, which comprises raising an immune response in said animal or human according to  claim 42.