Bacterial superantigen vaccines

• US 20050064526A1
• Filed: 01/29/2004
• Published: 03/24/2005
• Est. Priority Date: 06/25/1997
• Status: Active Grant

• Current Assignee: United States Army Medical Research and Materiel Command
• Original Assignee: United States Army Medical Research and Materiel Command
• Inventors: Bavari, Sina, Ulrich, Robert G., Olson, Mark A.
• Granted Patent: US 7,378,257 B2
• Time in Patent Office: Days
• US Class Current: 435/7.320
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: C07K 14/31 from Staphylococcus (G)

Abstract

The present invention relates to genetically attenuated superantigen toxin vaccines altered such that superantigen attributes are absent, however the superantigen is effectively recognized and an appropriate immune response is produced. The attenuated superantigen toxins are shown to protect animals against challenge with wild type toxin. Methods of producing and using the altered superantigen toxins are described.

Claims

1. An isolated and purified superantigen toxin DNA fragment which has been altered such that binding of the encoded altered toxin to either the MHC class II or T cell antigen receptor is altered.

7. An isolated and purified DNA fragment according to claim 1, wherein said superantigen toxin is toxic shock syndrome toxin-1 having the sequence of SEQ ID NO:

11 or a portion thereof, or an allelic portion thereof.

8. An isolated and purified DNA fragment according to claim 1, wherein said superantigen toxin is Staphylococcal enterotoxin C1 having the sequence of SEQ ID NO:

13 or a portion thereof, or an allelic portion thereof.

9. An isolated and purified DNA fragment according to claim 1, wherein said superantigen toxin is Streptococcal pyrogenic exotoxin A having the sequence of SEQ ID NO:

15 or a portion thereof, or an allelic portion thereof.

15. An isolated and purified DNA fragment according to claim 7, wherein said fragment encodes the amino acid sequence of SEQ ID NO:

12 or a portion thereof, or an allelic portion thereof.

16. An isolated and purified DNA fragment according to claim 8, wherein said fragment encodes the amino acid sequence of SEQ ID NO:

14 or a portion thereof, or an allelic portion thereof.

17. An isolated and purified DNA fragment according to claim 9, wherein said fragment encodes the amino acid sequence of SEQ ID NO:

16 or a portion thereof, or an allelic portion thereof.

18. A recombinant DNA construct comprising:

(i) a vector, and

(ii) an isolated and purified altered superantigen toxin DNA fragment according to claim 1.

24. A recombinant DNA construct according to claim 18, wherein said DNA fragment has the sequence according to SEQ ID NO:

11 or a portion thereof, or an allelic portion thereof.

25. A recombinant DNA construct according to claim 18, wherein said DNA fragment has the sequence according to SEQ ID NO:

13 or a portion thereof, or an allelic portion thereof.

26. A recombinant DNA construct according to claim 18, wherein said DNA fragment has the sequence according to SEQ ID NO:

15 or a portion thereof, or an allelic portion thereof.

32. The recombinant DNA construct according to claim 24, wherein said DNA fragment encodes the amino acids sequence specified in SEQ ID NO:

12.

33. The recombinant DNA construct according to claim 25, wherein said DNA fragment encodes the amino acids sequence specified in SEQ ID NO:

14.

34. The recombinant DNA construct according to claim 26, wherein said DNA fragment encodes the amino acids sequence specified in SEQ ID NO:

16.

40. A recombinant DNA construct according to claim 24 wherein said construct is pETTST30.

41. A recombinant DNA construct according to claim 25 wherein said construct is pETSEC45.

42. A recombinant DNA construct according to claim 26 wherein said construct is pETSPEA42.

43. A recombinant DNA construct according to claim 18, wherein said vector is an expression vector.

44. A host cell transformed with a recombinant DNA construct according to claim 18.

50. A host cell transformed with a recombinant DNA construct according to claim 32.

51. A host cell transformed with a recombinant DNA construct according to claim 33.

52. A host cell transformed with a recombinant DNA construct according to claim 34.

53. A host cell according to claim 44, wherein said cell is prokaryotic.

59. A host cell according to claim 50, wherein said cell is prokaryotic.

60. A host cell according to claim 51, wherein said cell is prokaryotic.

61. A host cell according to claim 52, wherein said cell is prokaryotic.

62. A method for producing altered superantigen toxin comprising culturing the cells according to claim 44, under conditions such that said DNA fragment is expressed and said superantigen toxin is thereby produced, and isolating said superantigen toxin.

68. A method for producing altered superantigen toxin comprising culturing the cells according to claim 50, under conditions such that said DNA fragment is expressed and said superantigen toxin is thereby produced, and isolating said superantigen toxin.

69. A method for producing altered superantigen toxin comprising culturing the cells according to claim 51, under conditions such that said DNA fragment is expressed and said superantigen toxin is thereby produced, and isolating said superantigen toxin.

70. A method for producing altered superantigen toxin comprising culturing the cells according to claim 52, under conditions such that said DNA fragment is expressed and said superantigen toxin is thereby produced, and isolating said superantigen toxin.

93. A bivalent vaccine according to claim 68 wherein said altered superantigen toxins are SEA and SEB.

2-6. -6. (Cancel without prejudice or disclaimer)

10-14. -14. (Cancel without prejudice or disclaimer)

19-23. -23. (Cancel without prejudice or disclaimer)

27-31. -31. (Cancel without prejudice or disclaimer)

35-39. -39. (Cancel without prejudice or disclaimer)

45-49. -49. (Cancel without prejudice or disclaimer)

54-58. -58. (Cancel without prejudice or disclaimer)

63-67. -67 (cancel without prejudice or disclaimer)

71. An isolated and purified superantigen toxin which has been altered such that binding of the encoded altered toxin to either the MHC class II or T cell antigen receptor is altered.

72. An isolated and purified superantigen toxin according to claim 71 wherein said toxin is staphylococcal enterotoxin A.

73. An isolated and purified superantigen toxin according to claim 71 wherein said toxin is staphylococcal enterotoxin B.

74. An isolated and purified superantigen toxin according to claim 71 wherein said toxin is staphylococcal toxin shock syndrome toxin-1.

75. An isolated and purified superantigen toxin according to claim 71 wherein said toxin is staphylococcal enterotoxin C1.

76. An altered SEA superantigen toxin peptide according to claim 72 wherein position 92 has been changed to alanine.

77. An altered SEA superantigen toxin peptide according to claim 72 wherein position 70 has been changed to arginine.

78. An altered SEA superantigen toxin peptide according to claim 72 wherein position 48 has been changed to arginine.

79. An altered SEA superantigen toxin peptide according to claim 72 wherein position 64 has been mutated to alanine.

80. An altered SEB superantigen toxin peptide according to claim 73 wherein position 115 has been changed to alanine.

81. An altered SEB superantigen toxin peptide according to claim 73 wherein position 89 has been changed to alanine.

82. An altered SEB superantigen toxin peptide according to claim 73 wherein position 67 has been changed to glutamine.

83. An altered SEB superantigen toxin peptide according to claim 73 wherein position 94 has been changed to alanine.

84. An altered SEB superantigen toxin peptide according to claim 73 wherein position 61 has been changed to alanine.

94. A bivalent vaccine according to claim 71 wherein said toxin SEA is A489270 and SEB is B899445.

85. A method for the diagnosis of superantigen-associated bacterial infection comprising the steps of:

(i) contacting a sample from an individual suspected of having a superantigen-associated bacterial infection with altered superantigen toxin; and

(ii) detecting the presence or absence of a superantigen-associated bacterial infection by detecting the presence or absence of a complex formed between the altered superantigen toxin and antibodies specific therefor in the sample.

95. A multivalent vaccine against superantigen-associated bacterial infections comprising a combination of altered superantigen toxins selected from the group consisting essentially of TSST-1, SPEa, SEA, SEB, and SEC-1 or any portion or allelic form thereof, capable of eliciting protective antibodies against superantigen toxins in a pharmaceutically acceptable excipient in a pharmaceutically acceptable amount.

97. A therapeutic method for the treatment or amelioration of a superantigen-associated bacterial infection, said method comprising administering to an individual in need of such treatment an effective amount of antibodies against altered superantigen toxins in a pharmaceutically acceptable dose in a pharmaceutically acceptable excipient.

98. A therapeutic method for the treatment or amelioration of a superantigen-associated bacterial infection, said method comprising administering to an individual in need of such treatment an effective amount of altered superantigen toxins from streptococcal and staphylococcal bacteria in order to inhibit adhesion of superantigen bacterial toxin to MHC class II or T cell receptors by competitive inhibition of these interactions in a pharmaceutically acceptable dose in a pharmaceutically acceptable excipient.

99. A therapeutic method for the treatment of diseases that may not be associated directly with superantigen toxins by causing specific nonresponsiveness of T cell subsets or by expanding or stimulating specific T cell subsets, in vivo or ex vivo by use of altered superantigen toxin.

100. The DNA construct designated pETB2360210.

101. An isolated and purified superantigen toxin DNA fragment encoding Staphylococcal enterotoxin B (SEB) in which at least one amino acid of amino acid positions 40-50 of SEB and at least one amino acid selected from the group consisting of amino acid positions 18-28, 55-65, 62-72, 84-94, 86-96, 89-99, 110-120 and 205-215 of SEB have been altered such that binding of said encoded SEB to the MHC class II receptor and T cell antigen receptor is altered.

102. The DNA fragment of claim 112, which encodes SEB in which the amino acids in positions 40-50, 84-89 and 89-99 have been altered.