Live recombinant booster vaccine against tuberculosis
Embodiments of the invention comprise an improved vaccine for generating an immune response and preventing or treating mycobacterial diseases such as tuberculosis in humans and animals. Embodiments of the invention also comprise a method for using the vaccine against such mycobacterial diseases.
1. A method of generating an antibody to a Mycobacterium tuberculosis 30 kDa antigen 85B protein (SEQ ID NO: 4) comprising the steps of:
immunizing a mammal with a composition consisting essentially of Mycobacterium bovis strain Bacille Calmette-Guérin (BCG) in a primary immunization, wherein said Mycobacterium bovis strain BCG is not a recombinant strain BCG;
immunizing a mammal with a composition of matter comprising attenuated Listeria monocytogenes in a booster immunization, wherein the Listeria monocytogenes:
does not express a functional ActA protein (SEQ ID NO: 1);
expresses prfA protein having a G155S substitution mutation (SEQ ID NO: 3); and
expresses Mycobacterium tuberculosis 30 kDa antigen 85B protein (SEQ ID NO: 4);
such that an antibody to a Mycobacterium tuberculosis 30 kDa antigen 85B protein is generated.
2. The method of claim 1 , wherein the Listeria monocytogenes expresses Mycobacterium tuberculosis 32 kDa 85A protein (SEQ ID NO: 11).
3. The method of claim 1 , wherein the Listeria monocytogenes : does not express a functional InlB protein (SEQ ID NO: 2).
4. The method of claim 1 , wherein the mammal is immunized intranasally, subcutaneously, intradermally, intramuscularly or orally.
5. The method of claim 1 , wherein the mammal is a guinea pig or a mouse.
6. The method of claim 1 , wherein the mammal is a human.
7. The method of claim 1 , wherein the Listeria monocytogenes further expresses at least one protein from the following group:
Mycobacterium tuberculosis 12 kDa fragment of 16 kDa membrane protein (SEQ ID NO:5);
Mycobacterium tuberculosis 14 kDa MPT53 protein (SEQ ID NO: 6);
Mycobacterium tuberculosis 16 kDa MPT63 protein (SEQ ID NO: 7);
Mycobacterium tuberculosis 23 kDa SOD protein (SEQ ID NO: 8);
Mycobacterium tuberculosis 23.5 kDa MPT64 protein (SEQ ID NO: 9);
Mycobacterium tuberculosis 24 kDa MPT51 protein (SEQ ID NO: 10);
Mycobacterium tuberculosis 32 kDa antigen 85A protein (SEQ ID NO: 11);
Mycobacterium tuberculosis 32 kDa antigen 85C protein (SEQ ID NO: 12);
Mycobacterium tuberculosis 45 kDa MPT32 protein (SEQ ID NO: 13);
Mycobacterium tuberculosis 58 kDa glutamine synthetase protein (SEQ ID NO: 14);
Mycobacterium tuberculosis 71 kDa HSP 70 protein (SEQ ID NO: 15);
Mycobacterium tuberculosis 10.4 kDa EsxH protein (SEQ ID NO: 16);
Mycobacterium tuberculosis 14 kDa alpha crystalline homolog protein (SEQ ID NO: 17);
Mycobacterium tuberculosis 47 kDa isocytrate lysate protein (SEQ ID NO: 18);
Mycobacterium tuberculosis 7.6 kDa hypothetical protein (SEQ ID NO: 19);
Mycobacterium tuberculosis 80 kDa glcB protein (SEQ ID NO: 20);
Mycobacterium tuberculosis 110 kDa can protein (SEQ ID NO: 21); or
Mycobacterium tuberculosis 9.9 kDa ESAT-6 protein (SEQ ID NO: 22).
8. The method of claim 1 , wherein the Listeria monocytogenes further expresses at least one protein from group A:
Mycobacterium tuberculosis 12 kDa fragment of 16 kDa membrane protein (SEQ ID NO:5);
Mycobacterium tuberculosis 14 kDa MPT53 protein (SEQ ID NO: 6);
Mycobacterium tuberculosis 16 kDa MPT63 protein (SEQ ID NO: 7);
Mycobacterium tuberculosis 23 kDa SOD protein (SEQ ID NO: 8);
Mycobacterium tuberculosis 23.5 kDa MPT64 protein (SEQ ID NO: 9);
Mycobacterium tuberculosis 24 kDa MPT51 protein (SEQ ID NO: 10);
Mycobacterium tuberculosis 32 kDa antigen 85A protein (SEQ ID NO: 11);
Mycobacterium tuberculosis 32 kDa antigen 85C protein (SEQ ID NO: 12);
Mycobacterium tuberculosis 45 kDa MPT32 protein (SEQ ID NO: 13);
Mycobacterium tuberculosis 58 kDa glutamine synthetase protein (SEQ ID NO: 14);
Mycobacterium tuberculosis 71 kDa HSP 70 protein (SEQ ID NO: 15);
Mycobacterium tuberculosis 10.4 kDa EsxH protein (SEQ ID NO: 16);
Mycobacterium tuberculosis 80 kDa glcB protein (SEQ ID NO: 20);
Mycobacterium tuberculosis 110 kDa can protein (SEQ ID NO: 21); or
Mycobacterium tuberculosis 9.9 kDa ESAT-6 protein (SEQ ID NO: 22);
at least one protein from group B:
Mycobacterium tuberculosis 14 kDa alpha crystalline homolog protein (SEQ ID NO: 17);
Mycobacterium tuberculosis 47 kDa isocytrate lysate protein (SEQ ID NO: 18); or
Mycobacterium tuberculosis 7.6 kDa hypothetical protein (SEQ ID NO: 19).
9. The method of claim 1 , wherein the Mycobacterium tuberculosis 30 kDa antigen 85B protein is coupled to a heterologous protein sequence comprising the N-terminal 100 amino acids of the ActA protein.
10. The method of claim 9 , wherein the expression of the Mycobacterium tuberculosis 30 kDa antigen 85B protein is controlled by an ActA promoter.
11. A method of generating an immune response to a Mycobacterium tuberculosis 30 kDa antigen 85B protein (SEQ ID NO: 4) comprising the steps of: immunizing a mammal with a composition consisting essentially of Mycobacterium bovis strain Bacille Calmette-Guérin (BCG) in a primary immunization, wherein said Mycobacterium bovis strain BCG is not a recombinant strain BCG;
immunizing a mammal with a composition of matter comprising attenuated Listeria monocytogenes in a booster immunization, wherein the Listeria monocytogenes:
does not express a functional ActA protein (SEQ ID NO: 1);
expresses prfA protein having a G155S substitution mutation (SEQ ID NO: 3); and
expresses Mycobacterium tuberculosis 30 kDa antigen 85B protein (SEQ ID NO: 4);
such that an immune response to a Mycobacterium tuberculosis 30 kDa antigen 85B protein is generated.
12. The method of claim 11 , wherein the Listeria monocytogenes expresses Mycobacterium tuberculosis 10.4 kDa EsxH protein (SEQ ID NO: 16).
13. The method of claim 11 , wherein the Listeria monocytogenes expresses Mycobacterium tuberculosis 32 kDa 85A protein (SEQ ID NO: 11).
14. The method of claim 11 , wherein the Listeria monocytogenes expresses Mycobacterium tuberculosis 23.5 kDa MPT64 protein (SEQ ID NO: 9).
15. The method of claim 11 , wherein the Listeria monocytogenes expresses Mycobacterium tuberculosis 9.9 kDa ESAT-6 protein (SEQ ID NO: 22).
16. The method of claim 11 , wherein the Listeria monocytogenes expresses Mycobacterium tuberculosis 14 kDa alpha crystalline homolog protein (SEQ ID NO: 17).
17. The method of claim 11 , wherein the Listeria monocytogenes expresses Mycobacterium tuberculosis 47 kDa isocitrate lysate protein (SEQ ID NO: 18).
18. The method of claim 11 , wherein the Listeria monocytogenes expresses Mycobacterium tuberculosis 7.6 kDa hypothetical protein (SEQ ID NO: 19).
19. The method of claim 11 , wherein the Mycobacterium tuberculosis 30 kDa antigen 85B protein is coupled to a heterologous protein sequence comprising the N-terminal 100 amino acids of the ActA protein.
20. The method of claim 19 , wherein the expression of the Mycobacterium tuberculosis 30 kDa antigen 85B protein is controlled by an ActA promoter.