IP Library › Granted Patent US 8,277,816

Granted Patent B2

US 8,277,816 · App. 11/706,576 · Granted Oct 2, 2012

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Patent No.: US 8,277,816
App. No.: 11/706,576
Granted: Oct 2, 2012
Kind: B2

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Abstract

The present invention relates to the intersection of the fields of immunology and protein engineering, and particularly to antigens and vaccines useful in prevention of infection by Bacillus anthracis . Provided are recombinant protein antigens, compositions, and methods for the production and use of such antigens and vaccine compositions.

Claims (46)

1. An isolated antigen comprising a component of Bacillus anthracis virulence factor fused to a LicKM protein;

wherein the virulence factor component comprises at least one domain having an amino acid sequence with at least 95% identity to a sequence selected from the group consisting of SEQ ID NO.: 4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1; and

wherein the LicKM protein comprises an amino acid sequence with at least 95% identity to SEQ ID NO.: 8 or SEQ ID NO.: 10.

2. An immunogenic composition comprising an antigen comprising a component of Bacillus anthracis virulence factor fused to a LicKM protein and a pharmaceutically acceptable carrier;

wherein the virulence factor component comprises at least one domain having an amino acid sequence with at least 95% identity to a sequence selected from the group consisting of SEQ ID NO.:4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1;

wherein the LicKM protein comprises an amino acid sequence with at least 95% identity to SEQ ID NO.: 8 or SEQ ID NO.: 10;

and wherein the composition is capable of eliciting an immune response upon administration to a subject.

3. An immunogenic composition comprising: at least two antigens, each of which comprises a component of Bacillus anthracis virulence factor having an amino acid sequence with at least 95% identity to a sequence selected from the group consisting of SEQ ID NO.:4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1, wherein at least one of the antigens further comprises a LicKM protein, and wherein the LicKM protein comprises an amino acid sequence with at least 95% identity to SEQ ID NO.: 8 or SEQ ID NO.: 10.

4. The immunogenic composition of claim 3 , wherein at least one of the antigens comprises a virulence factor component consisting of an amino acid sequence selected from the group consisting of about amino acid residues 1-137 of SEQ ID NO.:2, amino acid residues 251-263 of SEQ ID NO.:2, amino acid residues 138-250 of SEQ ID NO.:2, amino acid residues 264-283 of SEQ ID NO.:2, amino acid residues 429-551 of SEQ ID NO.:2, amino acid residues 306-385 of SEQ ID NO:2, and amino acid residues 552-776 of SEQ ID NO.:2.

5. The immunogenic composition of claim 3 , wherein at least one of the antigens comprises the amino acid sequence of SEQ ID NO.: 14.

6. The immunogenic composition of claim 3 , wherein at least one of the antigens comprises a virulence factor component consisting of an amino acid sequence selected from the group consisting of about amino acid residues 1-250 of SEQ ID NO.:1, amino acid residues 241-456 of SEQ ID NO.:1, amino acid residues 477-595 of SEQ ID NO.:1, and amino acid residues 608-735 of SEQ ID NO.:1.

7. The immunogenic composition of claim 3 , wherein at least one of the antigens comprises the amino acid sequence of SEQ ID NO.: 12.

8. The immunogenic composition of claim 3 , wherein the virulence factor component consists of SEQ ID NO.:15.

9. The immunogenic composition of claim 3 , wherein the virulence factor component is fused to the LicKM protein at the LicKM N-terminus or at the LicKM C-terminus, or is fused to a loop structure that is exposed on the surface of LicKM and is located far from the LicKM active domain.

10. The immunogenic composition of claim 3 , wherein the virulence factor component of at least one antigen comprises at least two amino acid sequences with at least 95% identity to a sequence selected from the group consisting of SEQ ID NO.:4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

11. The immunogenic composition of claim 10 , wherein at least one antigen comprises SEQ ID NO.:4 and at least one antigen comprises amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, or amino acid residues 552-776 of SEQ ID NO.: 2.

12. The immunogenic composition of claim 3 , wherein the antigen is produced by a plant selected from a transgenic plant and a plant which transiently expresses the antigen.

13. The immunogenic composition of claim 3 , wherein the composition comprises antigen which is purified, partially purified, or unpurified from plant cells, a plant, seeds, fruit, or an extract thereof.

14. The immunogenic composition of claim 3 , further comprising at least one vaccine adjuvant.

15. The immunogenic composition of claim 14 wherein the adjuvant is selected from the group consisting of alum, MF59, MALP2, and saponin.

16. A method for inducing an immune response in a subject, comprising administering to a subject an effective amount of an anti-anthrax immunogenic composition, wherein the administration is sufficient to stimulate production of antigen specific antibodies or stimulate a cellular immune response by the subject; thereby inducing a protective immune response;

wherein the immunogenic composition comprises an antigen comprising a component of Bacillus anthracis virulence factor fused to a LicKM protein, wherein the LicKM protein comprises an amino acid sequence with at least 95% identity to SEQ ID NO.: 8 or SEQ ID NO.: 10; and

wherein the virulence factor component comprises at least one domain having an amino acid sequence with at least 95% identity to a sequence selected from the group consisting of SEQ ID NO.:4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

17. A method for inducing an immune response in a subject, comprising administering to a subject an effective amount of an anti-anthrax immunogenic composition, wherein the immunogenic composition comprise at least two antigens, each of which comprises a component of Bacillus anthracis virulence factor having an amino acid sequence with at least 95% identity to a sequence selected from the group consisting of SEQ ID NO.:4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1, wherein at least one of the antigens further comprises a LicKM protein, wherein the LicKM protein comprises an amino acid sequence with at least 95% identity to SEQ ID NO.: 8 or SEQ ID NO.: 10, and wherein the administration is sufficient to stimulate production of antigen specific antibodies or stimulate a cellular immune response by the subject; thereby inducing a protective immune response.

18. The isolated antigen of claim 1 , wherein the virulence factor component comprises at least one domain having an amino acid sequence with at least 97% identity to a sequence selected from the group consisting of SEQ ID NO.: 4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residue residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

19. The isolated antigen of claim 1 , wherein the virulence factor component comprises at least one domain having an amino acid sequence with at least 98% identity to a sequence selected from the group consisting of SEQ ID NO.: 4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

20. The isolated antigen of claim 1 , wherein the virulence factor component consists of an amino acid sequence selected from the group consisting of SEQ ID NO.: 4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

21. The isolated antigen of claim 1 , wherein the LicKM protein has an amino acid sequence with at least 97% identity to SEQ ID NO.: 8 or SEQ ID NO.: 10.

22. The isolated antigen of claim 1 , wherein the virulence factor component is fused to the LicKM protein at the LicKM N-terminus or at the LicKM C-terminus, or is fused to a loop structure that is exposed on the surface of LicKM and is located far from the LicKM active domain.

23. The isolated antigen of claim 1 , wherein the antigen has the amino acid sequence of SEQ ID NO.: 12.

24. The isolated antigen of claim 1 , wherein the antigen has the amino acid sequence of SEQ ID NO.: 14.

25. The isolated antigen of claim 1 , wherein the antigen is produced by a plant selected from a transgenic plant and a plant which transiently expresses the antigen.

26. The isolated antigen of claim 1 , wherein the antigen is purified, partially purified, or unpurified from plant cells, a plant, seeds, fruit, or an extract thereof.

27. The immunogenic composition of claim 2 , wherein the virulence factor component comprises at least one domain having an amino acid sequence with at least 97% identity to a sequence selected from the group consisting of SEQ ID NO.: 4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

28. The immunogenic composition of claim 2 , wherein the virulence factor component comprises at least one domain having an amino acid sequence with at least 98% identity to a sequence selected from the group consisting of SEQ ID NO.: 4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

29. The immunogenic composition of claim 2 , wherein the virulence factor component consists of an amino acid sequence selected from the group consisting of SEQ ID NO.: 4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

30. The immunogenic composition of claim 2 , wherein the LicKM protein has an amino acid sequence with at least 97% identity to SEQ ID NO.: 8 or SEQ ID NO.: 10.

31. The immunogenic composition of claim 2 , wherein the virulence factor component is fused to the LicKM protein at the LicKM N-terminus or at the LicKM C-terminus, or is fused to a loop structure that is exposed on the surface of LicKM and is located far from the LicKM active domain.

32. The immunogenic composition of claim 2 , wherein the antigen has the amino acid sequence of SEQ ID NO.: 12.

33. The immunogenic composition of claim 2 , wherein the antigen has the amino acid sequence of SEQ ID NO.: 14.

34. The immunogenic composition of claim 2 , wherein the antigen is produced by a plant selected from a transgenic plant and a plant that transiently expresses the antigen.

35. The immunogenic composition of claim 2 , wherein the composition comprises antigen that is purified, partially purified, or unpurified from plant cells, a plant, seeds, fruit, or an extract thereof.

36. The immunogenic composition of claim 2 , further comprising at least one vaccine adjuvant.

37. The immunogenic composition of claim 34 , wherein the adjuvant is selected from the group consisting of alum, MF59, MALP2, and saponin.

38. The immunogenic composition of claim 3 , wherein each antigen comprises a component of Bacillus anthracis virulence factor having an amino acid sequence with at least 97% identity to a sequence selected from the group consisting of SEQ ID NO.:4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID NO.: 2, amino acid residues 264-283 of SEQ ID NO.: 2, amino acid residues 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and amino acid residues 608-735 of SEQ ID NO.: 1.

39. The immunogenic composition of claim 3 , wherein each antigen comprises a component of Bacillus anthracis virulence factor having an amino acid sequence with at least 98% identity to a sequence selected from the group consisting of SEQ ID NO.:4, SEQ ID NO: 6, SEQ ID NO: 15, amino acid residues 1-137 of SEQ ID NO.: 2, amino acid residues 251-263 of SEQ ID NO.: 2, amino acid residues 138-250 of SEQ ID residues residue 429-551 of SEQ ID NO.: 2, amino acid residues 306-385 of SEQ ID NO.: 2, amino acid residues 552-776 of SEQ ID NO.: 2, amino acid residues 1-250 of SEQ ID NO.: 1, amino acid residues 251-456 of SEQ ID NO.: 1, amino acid residues 477-595 of SEQ ID NO.: 1, and about amino acid residues 608-735 of SEQ ID NO.: 1.

Assignments (2)

ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Oct 7, 2014

From: FRAUNHOFER USA, INC.

To: IBIO, INC.

Reel/Frame 033906/0310 →

ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded May 30, 2007

From: YUSIBOV, VIDADI; METT, VADIM; MUSIYCHUCK, KONSTANTIN; SKARJINSKAIA, MARINA

To: FRAUNHOFER USA, INC.

Reel/Frame 019355/0568 →