Influenza antigens, vaccine compositions, and related methods

• US 8,124,103 B2
• Filed: 02/13/2007
• Issued: 02/28/2012
• Est. Priority Date: 02/13/2006
• Status: Active Grant

• Current Assignee: iBio, Inc.
• Original Assignee: Fraunhofer USA Inc. (Fraunhofer-Institut für Nachrichtentechnik HHI)
• Inventors: Yusibov, Vidadi, Mett, Vadim, Musiychuck, Konstantin
• Primary Examiner(s): Blumel, Benjamin P
• Application Number:
  US11/706,573
• Publication Number:
  US 20070275014A1
• Time in Patent Office: 1,841 Days
• Field of Search: None
• US Class Current: 424/210.1
• CPC Class Codes: A61K 2039/517 Plant cells
• CPC Class Codes: A61K 2039/542 oral/gastrointestinal
• CPC Class Codes: A61K 2039/543 intranasal
• CPC Class Codes: A61K 2039/55505 Inorganic adjuvants
• CPC Class Codes: A61K 2039/55566 Emulsions, e.g. Freund's adjuvant, MF59
• CPC Class Codes: A61K 2039/6031 Proteins
• CPC Class Codes: A61K 2039/70 Multivalent vaccine
• CPC Class Codes: A61K 39/00 Medicinal preparations containing antigens or antibodies materials for immunoassay G01N33/53
• CPC Class Codes: A61K 39/12 Viral antigens
• CPC Class Codes: A61K 39/145 Orthomyxoviridae, e.g. influenza virus
• CPC Class Codes: A61K 39/385 Haptens or antigens, bound to carriers
• CPC Class Codes: A61P 31/16 for influenza or rhinoviruses
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C12N 15/62 DNA sequences coding for fusion proteins
• CPC Class Codes: C12N 15/8257 for the production of primary gene products, e.g. pharmaceutical products, interferon
• CPC Class Codes: C12N 15/8258 for the production of oral vaccines (antigens) or immunoglobulins
• CPC Class Codes: C12N 2760/16122 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12N 2760/16134 Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
• CPC Class Codes: C12N 9/2402 hydrolysing O- and S- glycosyl compounds (3.2.1)
• CPC Class Codes: C12N 9/2448 : Licheninase (3.2.1.73)
• CPC Class Codes: C12Y 302/01018 : Exo-alpha-sialidase (3.2.1.18), i.e. trans-sialidase
• CPC Class Codes: C12Y 302/01073 : Licheninase (3.2.1.73)

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 influenza virus. Provided are recombinant protein antigens, compositions, and methods for the production and use of such antigens and vaccine compositions.

Claims

1. An isolated antigen comprising a component of an influenza A integral membrane protein fused to a lichenase protein;

wherein the integral membrane protein component comprises at least one immunogenic portion selected from the group consisting of an immunogenic portion of hemagglutinin (HA), an immunogenic portion of neuraminidase (NA) and an immunogenic portion of M2; and

wherein the lichenase protein is a LicKM protein having the amino acid sequence of SEQ ID NO;

30 or SEQ ID NO;

32.

2. An immunogenic composition comprising an antigen comprising a component of an influenza A integral membrane protein fused to a lichenase protein and a pharmaceutically acceptable carrier;

wherein the integral membrane protein component comprises at least one immunogenic portion selected from the group consisting of an immunogenic portion of hemagglutinin (HA), an immunogenic portion of neuraminidase (NA) and an immunogenic portion of M2;

wherein the lichenase protein is a LicKM protein having the amino acid sequence of SEQ ID NO;

30 or SEQ ID NO;

32; and

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

10. The immunogenic composition of claim 2, wherein the integral membrane protein component is fused to LicKM at the LicKM N-terminus or at the LicKM C-terminus or is fused to a surface loop of LicKM.

3. An immunogenic composition comprising at least two antigens and a pharmaceutically acceptable carrier,

wherein each of the antigens comprises a component of an influenza A integral membrane protein fused to a lichenase protein, wherein the integral membrane protein component comprises at least one immunogenic portion selected from the group consisting of an immunogenic portion of hemagglutinin (HA), an immunogenic portion of neuraminidase (NA) and an immunogenic portion of M2;

wherein the lichenase protein is a LicKM protein having the amino acid sequence of SEQ ID NO;

30 or SEQ ID NO;

32; and

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

4. The immunogenic composition of claim 3, wherein at least one of the integral membrane protein components comprises at least one immunogenic portion of hemagglutinin (HA), wherein the immunogenic portion is selected from the group consisting of SEQ ID NO:

1, SEQ ID NO;

3, SEQ ID NO;

7, SEQ ID NO;

8, SEQ ID NO;

33, SEQ ID NO;

11, SEQ ID NO;

12, SEQ ID NO;

13, SEQ ID NO;

20 and SEQ ID NO;

24.

5. The immunogenic composition of claim 3, wherein at least one of the integral membrane protein components comprises at least one immunogenic portion of neuraminidase (NA), wherein the immunogenic portion is selected from the group consisting of SEQ ID NO:

2, SEQ ID NO;

4, SEQ ID NO;

16, and SEQ ID NO;

18.

6. The immunogenic composition of claim 3, wherein at least one of the integral membrane protein components consists of full length hemagglutinin (HA) selected from SEQ ID NO:

1 and SEQ ID NO;

3.

7. The immunogenic composition of claim 3, wherein at least one of the integral membrane protein components consists of full length neuraminidase (NA) selected from SEQ ID NO:

2 and SEQ ID NO;

4.

8. The immunogenic composition of claim 3, wherein the LicKM protein comprises the amino acid sequence set forth in SEQ ID NO:

30.

9. The immunogenic composition of claim 3, wherein the LicKM protein comprises the amino acid sequence set forth in SEQ ID NO:

32.

11. The immunogenic composition of claim 3, wherein the integral membrane protein component comprises at least two immunogenic portions selected from the group consisting of SEQ ID NO:

1, SEQ ID NO;

3, SEQ ID NO;

7, SEQ ID NO;

8, SEQ ID NO;

33, SEQ ID NO;

11, SEQ ID NO;

12, SEQ ID NO;

13, SEQ ID NO;

20, SEQ ID NO;

24, SEQ ID NO;

2, SEQ ID NO;

4, SEQ ID NO;

16, and SEQ ID NO;

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

12. The immunogenic composition of claim 11, wherein the integral membrane protein component comprises an immunogenic portion of hemagglutinin (HA) and an immunogenic portion of neuraminidase (NA).

13. The immunogenic composition of claim 11, wherein the integral membrane protein component comprises full length hemagglutinin (HA) and full length neuraminidase (NA).

14. The immunogenic composition of claim 3, further comprising a third antigen, wherein the composition is capable of eliciting an immune response upon administration to an animal.

15. The immunogenic composition of claim 14, wherein the third antigen comprises a component of influenza A integral membrane protein fused to a lichenase protein;

wherein the integral membrane protein component of the first antigen comprises at least one immunogenic portion selected from the group consisting of an immunogenic portion of hemagglutinin (HA), an immunogenic portion of neuraminidase (NA) and an immunogenic portion of M2, and the integral membrane protein component of the second antigen comprises at least one immunogenic portion distinct from the first antigen selected from the group consisting of an immunogenic portion of hemagglutinin (HA), an immunogenic portion of neuraminidase (NA) and an immunogenic portion of M2; and

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

16. The immunogenic composition of claim 3 wherein the antigen is produced in a plant selected from a transgenic plant and a plant transiently expressing the antigen.

17. 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.

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

19. The immunogenic composition of claim 18 wherein the adjuvant is selected from the group consisting of alum, MF59, saponin, and MALP2.

24. The immunogenic composition of claim 18 wherein the adjuvant comprises a mixture of QS-21, and 3 de-O-acylated monophosphoryl lipid A (3D-MPL).

25. The immunogenic composition of claim 3, wherein the integral membrane protein component is fused to LicKM at the LicKM N-terminus or at the LicKM C-terminus or is fused to a surface loop of LicKM.

20. A method for inducing an immune response against influenza A infection in a subject comprising administering to a subject an effective amount of an anti-influenza A 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 an immune response;

wherein the immunogenic composition comprises an antigen comprising a component of an influenza A integral membrane protein fused to a lichenase protein, wherein the lichenase protein is a LicKM protein having the amino acid sequence of SEQ ID NO;

30 or SEQ ID NO;

32; and

wherein the integral membrane protein component comprises at least one immunogenic portion selected from the group consisting of an immunogenic portion of hemagglutinin (HA), an immunogenic portion of neuraminidase (NA) and an immunogenic portion of M2.

21. A method for producing an antigen protein comprising a component of an influenza A integral membrane protein fused to a lichenase protein, comprising:

(a) preparing a nucleic acid construct encoding an antigen comprising a component of an influenza A integral membrane protein fused to a lichenase protein;

(b) introducing the nucleic acid of step (a) into a cell; and

(c) incubating the cell under conditions favorable for expression of the antigen protein;

thereby producing the antigen protein;

wherein the integral membrane protein component comprises at least one immunogenic portion selected from the group consisting of an immunogenic portion of hemagglutinin (HA) and an immunogenic portion of neuraminidase (NA), and wherein the lichenase protein is a LicKM protein having the amino acid sequence of SEQ ID NO;

30 or SEQ ID NO;

32.

22. An isolated nucleic acid construct comprising nucleic acid sequence encoding a component of an influenza A integral membrane protein fused to a lichenase protein;

wherein the lichenase protein is a LicKM protein having the amino acid sequence of SEQ ID NO;

30 or SEQ ID NO;

32; and

wherein the integral membrane protein component comprises at least one immunogenic portion selected from the group consisting of an immunogenic portion of hemagglutinin (HA) and an immunogenic portion of neuraminidase (NA).

23. An isolated host cell comprising the nucleic acid construct of claim 22.