Methods for improving immunological response in vaccinated animals

• US 10,441,652 B2
• Filed: 08/20/2015
• Issued: 10/15/2019
• Est. Priority Date: 08/21/2014
• Status: Active Grant

• Current Assignee: Braasch Biotech LLC
• Original Assignee: Braasch Biotech LLC
• Inventors: Haffer, Keith N.
• Primary Examiner(s): Kim, Yunsoo
• Application Number:
  US14/831,602
• Publication Number:
  US 20160051668A1
• Time in Patent Office: 1,517 Days
• Field of Search: None
• CPC Class Codes: A61K 2039/552 Veterinary vaccine
• CPC Class Codes: A61K 2039/6031 Proteins
• CPC Class Codes: A61K 2039/627 characterised by the linker
• CPC Class Codes: A61K 39/0005 Vertebrate antigens from snakes A61K39/38
• CPC Class Codes: A61K 39/385 Haptens or antigens, bound to carriers
• CPC Class Codes: C07K 14/655 Somatostatins
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C07K 2319/40 containing a tag for immunodetection, or an epitope for immunisation
• CPC Class Codes: C12N 9/1033 Chloramphenicol O-acetyltransferase (2.3.1.28)

Abstract

A method is provided for increasing an immunological response to a target antigen in an animal by administering an immunogenic amount of a vaccine comprising a polypeptide conjugate comprising the target antigen conjugated to a carrier polypeptide by means of a linker polypeptide which is rich in predicted linear B-cell epitopes.

Claims

1. A method for providing a vaccine capable of stimulating an improved immunological response to a target antigen in an animal, the method comprising

selecting a target antigen exhibiting low % predicted linear B-cell epitopes of <

10% predicted linear B-cell epitopes;

providing a polypeptide conjugate comprising the target antigen conjugated to a carrier polypeptide by means of a linker polypeptide exhibiting high % predicted linear B-cell epitopes of >

50% predicted linear B-cell epitopes; and

administering to the animal an immunogenic amount of a vaccine comprising the polypeptide conjugate comprising (a) the carrier polypeptide comprising an inactivated chloramphenicol acetyl transferase (CAT) enzyme;

(b) the linker polypeptide, and (c) the target antigen, wherein the carrier polypeptide does not stimulate a substantial T-cell response, and

wherein the improved immunological response in the animal comprises a statistically significant faster immunological response or specific effect within four days after the first administration of the vaccine compared to a control animal immunized with target antigen and/or carrier without the linker polypeptide,

wherein the % predicted linear B-cell epitopes for the target antigen is determined by

(1) generating a BepiPred 1.0 predicted Linear B-cell epitope score for each amino acid in the amino acid sequence of the target antigen;

(2) counting the amino acids for which the BepiPred 1.0 predicted Linear B-cell epitope score is above a threshold value of 0.2;

(3) dividing the number of amino acids above the threshold value by the number of amino acids in the target antigen to get a fraction; and

(4) multiplying the fraction by 100 to obtain the % predicted linear B-cell epitopes for the target antigen.

2. The method according to claim 1, wherein the target antigen exhibits low % predicted linear B-cell epitopes of <

5% predicted linear B-cell epitopes.

3. The method according to claim 1, wherein the linker polypeptide exhibits high % predicted linear B-cell epitopes of >

80% predicted linear B-cell epitopes, wherein the % predicted linear B-cell epitopes in the linker polypeptide is determined by

(1) generating a BepiPred 1.0 predicted Linear B-cell epitope score for each amino acid in the amino acid sequence of the linker polypeptide;

(2) counting the amino acids in the linker polypeptide for which the BepiPred 1.0 predicted Linear B-cell epitope score is above a threshold value;

(3) dividing the number of amino acids above the threshold value by the total number of amino acids in the linker polypeptide to get a fraction; and

(4) multiplying the fraction by 100 to get the % predicted Linear B-cell Epitopes for the linker polypeptide.

4. The method according to claim 1, wherein the carrier polypeptide comprises an inactivated chloramphenicol acetyl transferase (CAT) enzyme comprising the amino acid sequence selected from SEQ ID NO:

3, 7, 8, 26, 27, 28, or 29.

5. The method according to claim 1, wherein the linker polypeptide comprises the amino acid sequence selected from SEQ ID NO:

10, 11, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 53, 54, 55, 56 or 57.

6. The method according to claim 1, wherein the improved immunological response further comprises one or more of (1) a statistically significant increase in serum anti-target antigen IgG or serum anti-carrier IgG from the animal observed after the first or subsequent administration of the vaccine comprising the polypeptide conjugate, compared to control immunized with target antigen and/or carrier without the linker polypeptide, or (2) a statistically significant increase or decrease in a specific effect in the animal following administration of the vaccine comprising the polypeptide conjugate compared to a control.

7. The method according to claim 1, wherein the specific effect is selected from increased milk production, increased body weight, increased lean body weight, decreased percent body-fat, or decreased body weight in the animal compared to a control without immunization.

8. The method according to claim 1, wherein the immunogenicity of said target antigen in the polypeptide conjugate is increased as compared to the target antigen in the absence of the polypeptide conjugate.

9. The method according to claim 2, wherein the target antigen is a polypeptide from 5 to 40 amino acids in length.

10. The method according to claim 1, wherein the target antigen comprises the amino acid sequence consisting of SEQ ID NO:

1.

11. The method according to claim 9, wherein the target antigen comprises the amino acid sequence consisting of SEQ ID NO:

1.

12. The method according to claim 2, wherein the target antigen is not somatostatin 14.

13. The method according to claim 1, wherein the CAT enzyme has one or more 192 or 193 wild type histidine residues is independently replaced with an amino acid selected from alanine, glycine or other like amino acid.

14. The method according to claim 1, wherein the chloramphenicol acetyl transferase polypeptide is truncated at the C-terminus by 10 amino acids relative to the wild type polypeptide.

15. The method according to claim 1, wherein the vaccine is substantially endotoxin free.

16. The method according to claim 1, wherein the vaccine is at least 80% endotoxin free.

17. The method according to claim 1, wherein the BepiPred 1.0 predicted Linear B-cell epitope score threshold is 0.35.

18. The method according to claim 3, wherein the BepiPred 1.0 predicted Linear B-cell epitope score threshold is 0.2.

19. The method according to claim 1, wherein the linker polypeptide is heterologous to the target antigen.

20. The method of claim 1, wherein subsequent administration of the vaccine does not produce an anamnestic response in the animal.

21. The method according to claim 1, wherein the target antigen is from 8 to 14 amino acids in length.

22. The method according to claim 1, wherein neither the target antigen, the linker, nor the carrier polypeptide, stimulate a substantial T-cell response.