Login| Sign Up| Help| Contact|

Patent Searching and Data


Title:
MODULATION OF TH1 AND TH2 IMMUNE RESPONSE BY LACTIC ACID WHEY
Document Type and Number:
WIPO Patent Application WO/2012/057636
Kind Code:
A1
Abstract:
The present invention relates to the use of lactic acid whey or a derivative thereof for modulating the Th-1 and Th-2 immune responses in a subject in need thereof. The present invention relates to a method of modulating an immune response in the treatment or prevention of diseases or conditions including those caused by intracellular or extracellular pathogens, and allergens.

Inventors:
MCCONNELL MICHELLE AVRIL (NZ)
HAGGARTY NEILL WARD (NZ)
BUCHAN KERRY THERESE (NZ)
BUCHAN GLENN STUART
Application Number:
PCT/NZ2011/000228
Publication Date:
May 03, 2012
Filing Date:
October 27, 2011
Export Citation:
Click for automatic bibliography generation   Help
Assignee:
FONTERRA CORPORATE RES AND DEV LTD (NZ)
FONTERRA LTD (NZ)
AUCKLAND UNISERVICES LTD (NZ)
MCCONNELL MICHELLE AVRIL (NZ)
HAGGARTY NEILL WARD (NZ)
BUCHAN KERRY THERESE (NZ)
International Classes:
A61K38/20; A23C21/02; C07K14/52
Domestic Patent References:
WO2009151331A12009-12-17
WO2006096073A12006-09-14
WO2003053158A22003-07-03
Foreign References:
EP2165713A12010-03-24
Other References:
NAKANISHI. K ET AL., ANNUAL REVIEW OF IMMUNOLOGY, vol. 19, 2001, pages 423 - 474
MCILROY. A ET AL.: "Histamine and prostaglandin E2 up-regulate the production of Th2-attracting chemokines (CCL 17 and CCL22) and down-regulate IFN-y-induced CXCL 10 production by immature human dendritic cells.", IMMUNOLOGY, vol. 117, 2006, pages 507 - 516
Attorney, Agent or Firm:
MANSELL, John B. et al. (P.O.Box 949, 6140 Wellington, NZ)
Download PDF:
Claims:
What we claim is:

1. A method of modulating an immune response in a subject in need thereof, the method

comprising administering lactic acid whey or a derivative thereof to the subject.

2. The method of claim 1 wherein the immune response is a Thl immune response.

3. The method of claim 2 wherein the modulation of the immune response is an upregulation of the Thl response.

4. The method of claim 2 wherein modulation of the immune response is a downregulation of the Thl response.

5. The method of claim 2 wherein modulation of the immune response is an upregulation of at least one Thl cytokine, such as at least one Thl -specific cytokine.

6. The method of claim 2 wherein modulation of the immune response is a downregulation of at least one Thl cytokine, such as at least one Thl-specific cytokine.

7. The method of claim 5 or claim 6 wherein the at least one Thl-specific cytokine is selected from the group comprising IL- b, IL-2, IL-3, IL-6, IL-12, IFN-γ, TNF-oc or TNF-β.

8. The method of claim 3, wherein upregulation of a Thl response is characterised by

a. increased IgG2a, or

b. increased macrophage activation or proliferation, or

c. increased CD 8+ cytotoxic T-cell activation or proliferation, or

d. any combination of two or more of (a) to (c) above.

9. The method of claim 1 wherein the immune response is a Th2 immune response.

10. The method of claim 9 wherein the modulation of the immune response is a downregulation of the Th2 response.

11. The method of claim 9 wherein the modulation of the immune response is an upregulation of the Th2 response.

12. The method of claim 9 wherein the modulation of the immune response is a downregulation of at least one Th2 cytokine, such as at least one Th2-specific cytokine.

13. The method of claim 9 wherein the modulation of the immune response is an upregulation of at least one Th2 cytokine, such as at least one Th2-specific cytokine.

14. The method of claim 12 or 13 wherein the at least one Th2-specific cytokine is selected from the group comprising IL-4, IL-5, IL-6, IL- 0 or IL-13.

15. The method of claim 9, wherein upregulation of a Th2 response is characterised by

a. increased IgGl, or

b. an upregulation of B-cells, including an activation or proliferation of B-cells, or

production of IgA isotype, or

c. an increase in the production of neutralizing lolling antibodies, or

d. any combination of two or more of (a) to (c) above.

16. The method of claim 1, wherein the modulation of the immune response is activation of

regulatory T cells.

17. The method of claim 1, wherein the modulation of the immune response is upregulation of IL-17 cytokine.

18. A method of modulating a combined Thl artd Th2 immune response in a subject in need

thereof, the method comprising administering lactic acid whey or a derivative thereof to the subject.

19. The method of claim 18, wherein the modulation of the combined Thl and Th2 immune

response is an upregulation of the Thl immune response and a downregulation of the Th2 immune response.

20. The method of claim 18, wherein the modulation of the combined Thl and Th2 immune

response is an upregulation of the Th2 immune response and a downregulation of the Thl response.

21. The method of claim 8, wherein the modulation of the combined Thl and Th2 immune

response is a downregulation of both the Thl and Th2 immune responses.

22. The method of claim 18, wherein the modulation of the combined Thl and Th2 immune

response is characterised by

a. an increase in me production of at least one Thl -specific cytokine, or

b. an increase in the production of at least one Th2-specific cytokine, or

c. a decrease in the production of at least one Thl -specific cytokine, or

d. a decrease in the production of at least one Th2-specific cytokine, or

e. any combination of two of (a) to (d) above.

23. The method of claim 22, wherein the at least one Th -specific cytokine is selected from the group comprising IL-lb, IL-2, IL-3, IL-12, IFN-γ, TNF-a or TNF-β.

24. The method of claim 22, wherein the at least one Th2-specific cytokine is selected from the

group comprising IL-4, IL-5, IL-6, IL-10 or IL-13.

25. The method of any one of claims 1 to 24 wherein the modulation of the immune response is characterised by a cytokine profile associated with a Th-1 response.

26. The method of claim 25 wherein the cytokine profile associated with a Th-1 response is

characterised by an increase in the production of IL-2 and IFN-γ.

27. The method of any one of claims 1 to 24 wherein the modulation of the immune response is characterised by a cytokine profile associated with a Th-2 response.

28. The method of claim 27 wherein the cytokine profile associated with a Th-2 response is

characterised by an increase in the production of IL-4 and IL-10.

29. The method of claim 1 or claim 18 wherein the immune response is a Thl immune response without significant Th2 immune response.

30. The method of claim 29 wherein the immune response is a Thl immune response in the absence of a significant IgA response, or in the absence of a significant IgE response, or in the absence of a significant IgG response, including the absence of a significant IgGl response, or the absence of a significant IgG2 response, or in the absence of a significant IgM response.

31. The method of claim 1 or claim 18 wherein the immune response is a Th2 response without significant Thl response.

32. A method of modulating an immune response of a subject to a pathogen, the method comprising . administering lactic acid whey or. a derivative thereof to a subject in need thereof.

33. The method of claim 32 wherein the pathogen is a bacterial pathogen.

34. The method of claim 33 wherein the bacterial pathogen is selected from the group comprising Bacteroides (e.g. B. fragilis), Brucella (e.g. B. melitensis, B. ovis, B. abortus, B. suis, B. pinnipediae and B. neotomae), Chlamydia (e.g. C. trachomatis), Chlamydophila (e.g. C. pneumoniae, C.

psittaci and C. abortus), Legionella (e.g. L. pneumophila), Listeria (e.g. L. monocytogene), Plasmodium (e.g. P. falciparum), Rickettsia (e.g. R. rickettsii, R. akari, R. conorii, R. siberica, R. australis, R. japonica, R. africae, R. prowazekii and R. typhi) , Salmonella, Streptococcus (e.g. S. pneumonia and S. pyogenes), Bordetella (e.g. B. pertussis, B. parapertussis and B. bronchiseptica), Borrelia (e.g. B. burgdorferi, B. afzelii, B. garinii and B. valaisiana), Campylobacter (e.g. C. jejuni and C. coli), Clostridium (e.g. C. botulinum, C. difficile, C. perfringens and C. tetani) , Corynebacterium (e.g. C. di theriae), Enterococcus (e.g. E. avium, E. durans, E. faecalis, E. faecium and E. solitaries), Escherichia (e.g. E. coli), Haemophilus, Helicobacter (e.g. H. pylori), Leptospira (e.g. L. interrogans, L. kirschneri, L. noguchii, L. alexanderi, L. weilii, L.

genomospecies 1, L. borgpetersenii, L. santarosai and L. kmetyi), Mycobacterium (e.g. M.

tuberculosis, M. bovis, M. africanum, M. microti and M. leprae) , Mycoplasma (e.g. M. pnemoniae and M. genitalium), Pseudomonas, Shigella, Staphylococcus, Trepnema (e.g. T. pallidum), Vibrio (e.g. V. cholerae and V. parahaemolyticus and V. vulnificus ), Yersinia (e.g. Y. enterocolitica, Y. pestis and Y. pseudotuberculosis) Ehrlichia (e.g. E. chaffeensis, Ehrlichia phagocytophila geno group, Ehrlichia sp.), Coxiella burnetii, Leishmania sp., Toxpolasma gondii, Trypanosoma cruzi, Histoplasma sp. or Francisella tukrensis.

35. The method of claim 32 wherein the pathogen is a viral pathogen.

36. The method of claim 35 wherein the viral pathogen is selected from the group comprising

Hepatitis C, Adenoviruses, Picornaviruses including coxsackievirus, hepatitis A virus, poliovirus, Herpesviruses including epstein-barr virus, herpes simplex type 1 , herpes simplex type 2, human cytomegalovirus, human herpesvirus type 8, varicella-zoster virus, Hepadnaviruses including hepatitis B virus, Flaviviruses including hepatitis C virus, yellow fever virus, dengue virus, West Nile virus, Retroviruses including human immunodeficiency virus (HIV), Orthomyxoviruses including influenza virus, Paramyxoviruses including measles virus, mumps virus, parainfluenza virus, respiratory syncytial virus, Papillomaviruses including papillomavirus, Rhabdoviruses including rabies virus, Togaviruses including Rubella virus, and other viruses including vaccinia, avipox, adeno-associated virus, modified Vaccinia Strain Ankara, Semliki Forest virus, poxvirus, and coronaviruses.

37. The method of claim 32 wherein the pathogen is a fungal pathogen.

38. The method of claim 37 wherein the fungal pathogen is selected from the group comprising Cryptococcus (C. neoformans, C. gattii), Pneumocystisi (P. carinii, P. jiroveci), Candida

(C.alb cans), Aspergillus (A. fumigates, A. deflectus, A. flavus, A. flavipes), Acremonium, Penicillium, Paecilomyces, Coccidioides, Histoplasma capsulatum, Blastomyces dermatitidis, Microsporum (M. canis), Trichophyton (Γ. verrucosum, T. mentagrophytes), Epidermophyton or Malassezia (M. pachydermatis).

39. A method of modulating an immune response in the treatment or prevention of an allergic

condition in a subject in need thereof, or of modulating an immune response in a subject in need thereof to an allergen, the method comprising administering lactic acid whey or a derivative thereof to the subject.

40. The method of claim 39 wherein the condition is an atopic condition.

41. The method of claim 39 wherein the condition is an eosinophilia.

42. The method of claim 39 wherein the condition is a Th2 mediated condition.

43. The method of claim 39 wherein the condition is selected from allergic rhinitis, hay fever, atopic rhinoconjunctivitis, urticaria, asthma and atopic eczema.

44. The method of claim 43 wherein the condition is asthma.

45. The method of claim 39 wherein the condition is selected from contact dermatitis, eczema, hives (urticaria), allergic conjunctivitis, hay fever, allergic rhinitis, airborne allergies including tree (e.g. birch pollen), weed (e.g. ragweed), and grass pollen allergies, latex allergies, food allergies (e.g. peanut, shellfish, milk protein), drug allergies (e.g. to penicillin), insect sting allergies (e.g.

honeybee allergies, wasp allergies, hornet allergies, yellow jacket allergies, fire ant allergies), mold allergies (e.g. to alternaria, cladosporium, aspergillus, penicillium, helminthosporium, epicoccum, fusarium, mucor, rhizopus, and aureobasidium), dust mite allergies, animal allergies (e.g.

household pets such as cats and dogs), allergic bronchopulmonary aspergillosis, occupational asthma, and episodic angioedema with eosinophilia.

46. The method of claim 39 wherein the allergic condition is selected from airway, lung, blood or skin eosinophilia.

47. The method of claim 41 wherein the eosinophilia is selected from eosinophilic ascites,

eosinophilic cellulitis, eosinophilic fasciitis, eosinophilic gastroenteritis, coeliac disease, allergic colitis, eosinophilic esophagitis, eosinophilic pancreatitis, eosinophilic pneumonias,

bronchiectasis, eosinophilic synovitis, nasal eosinophilia, tropical pulmonary eosinophilia, Churg Strauss syndrome, pulmonary eosinophilia, idiopathic hyper-eosinophilic syndrome,

inflammatory bowel disease, eosinophilic cholangitis, eosinophilic leukaemia and other eosinophilic cancers, familial (hereditary eosinophilia), eosinophilic granuloma, eosinophilia- myalgia syndrome, cystic fibrosis, nasal polyposis, eosinophil meningitis, Wegener's

granulomatosis, polyarteritis nodosa, pemphigus vulgaris, bullous pemphigoid, dermatitis herpetiformis, erythema multiforme, eosinophilic cellulites, parasitic infections (Ascaris Toxocara canis, Filariasis, Ankylostomiasis, Trichinosis, Strongyloidiasis, FascioHasis, Schistosomiasis).

48. The method of claim 39 wherein the condition is selected from Th2-mediated asthma, allergies, eczema, microbial or parasite infection, and autoimmune diseases including Crohn's disease.

49. The method of claim 39 wherein the subject is susceptible to anaphylaxis.

50. The method of claim 39 wherein the administration of the lactic acid whey or a derivative thereof is prophylactic administration.

51. The method of claim 39 wherein the subject is undergoing an allergy de-sensitisation regime.

52. The method of claim 39 wherein the condition is selected from Th2 mediated asthma, allergies, eczema, microbial or parasite infection, and autoimmune diseases including ulcerative colitis.

53. A method of modulating an immune response in a subject in need thereof, the method

comprising co-administration of lactic acid whey or a derivative thereof and one or more therapeutically active agent to the subject.

54. The method of claim 53 wherein the therapeutically active ingredient is an antigen.

55. The method of claim 54 wherein the antigen is capable of modulating or eliciting a Thl immune response, or is capable of modulating or eliciting a Th2 immune response.

56. The method of claim 54 wherein the antigen is derived from a bacterial pathogen, a viral

pathogen, or a fungal pathogen.

57. The method of claim 56 wherein the bacterial antigen is an antigen derived from a bacterial

pathogen selected from the group comprising Bacteroides (e.g. B. fragilis), Brucella (e.g. B.

melitensis, B. ovis, B. abortus, B. suis, B. pinnipediae and B. neotomae), Chlamydia (e.g. C.

trachomatis), Chlamydophila (e.g. C. pneumoniae, C. psittaci and C. abortus), Legionella (e.g. L. pneumophila), Listeria (e.g. L. monocytogene), Plasmodium (e.g. P. falciparum), Rickettsia (e.g. R. rickettsii, R. akari, R. conorii, R. siberica, R. australis, R. japonica, R. africae, R. prowazekii and R. typhi) , Salmonella, Streptococcus (e.g. S. pneumonia and S. pyogenes), Bordetella (e.g. B. pertussis, B. parapertussis and B. bronchiseptica), Borrelia (e.g. B. burgdorferi, B. afzelii, B. garinii and B. valaisiana), Campylobacter (e.g. C. jejuni and C. coli), Clostridium (e.g. C. botulinum, C. difficile, C. perfringens and C. tetani) , Corynebacterium (e.g. C. diptheriae), Enterococcus (e.g. E. avium, E. durans, E. faecalis, E. faecium and E. solitaries), Escherichia (e.g. E. coli),

Haemophilus, Helicobacter (e.g. H. pylori), Leptospira (e.g. L. interrogans, L. kirschneri, L.

noguchii, L. alexanderi, L. weilii, L. genomospecies 1, L. borgpetersenii, L. santarosai and L. kmetyi), Mycobacterium (e.g. M. tuberculosis, M. bovis, M. africanum, M. microti and M. leprae) , Mycoplasma (e.g. M. pnemoniae and M. genitalium), Pseudomonas, Shigella, Staphylococcus, Trepnema (e.g. T. pallidum), Vibrio (e.g. V. cholerae and V. parahaemolyticus and V. vulnificus ), Yersinia (e.g. Y. enterocoUtica, Y. pestis and Y. pseudotuberculosis) Ehrlichia (e.g. E. chaffeensis, Ehrlichia phagocytophila geno group, Ehrlichia sp.), Coxiella burnetii, Leishmania sp.,

Toxpolasma gondii, Trypanosoma cruzi, Histoplasma sp. or Francisella tularensis.

58. The method of claim 56 wherein the viral antigen is an antigen derived from a viral pathogen selected from the group comprising Hepatitis C, Adenoviruses, Picornaviruses including coxsackievirus, hepatitis A virus, poliovirus, Herpesviruses including epstein-barr virus, herpes simplex type 1, herpes simplex type 2, human cytomegalovirus, human herpesvirus type 8, varicella-zoster virus, Hepadnaviruses including hepatitis B virus, Flaviviruses including hepatitis C virus, yellow fever virus, dengue virus, West Nile virus, Retroviruses including human immunodeficiency virus (HIV), Orthomyxoviruses including influenza virus, Paramyxoviruses including measles virus, mumps virus, parainfluenza virus, respiratory syncytial virus,

Papillomaviruses including papillomavirus, Rhabdoviruses including rabies virus, Togaviruses including Rubella virus, and other viruses including vaccinia, avipox,

59. The method of claim 56 wherein the fungal antigen is derived from a fungal pathogen selected from the group comprising Cryptococcus (C. neoformans, C. gattii), Pneumocystisi (P. carinii, P. jiroveci), Candida (C.albicans), Aspergillus (A. fumigates, A. deflectus, A. flavus, A. flavipes), Acremonium, Penicillium, Paecilomyces, Coccidioides, Histoplasma capsulatum, Blastomyces dermatitidis, Microsporum (M. canis), Trichophyton (T. verrucosum, T. mentagrophytes), Epidermophyton or Malassezia (M. pachydermatis).

60. The method of any one of claims 53 to 59 wherein the composition additionally comprises an adjuvant, or an immunostimulatory molecule.

61. The method of any one of claims 53 to 60 wherein the co-administration of the lactic acid whey or a derivative thereof and the one or more therapeutically active agent is separate, simultaneous or sequential.

62. The method of any one of claims 1 to 61 wherein the subject is an infant or neonate mammal.

63. The method of any one of claims 1 to 62 wherein the subject is infected with a bacterial

pathogen.

64. The method of any one of claims 1 to 63 wherein the subject is infected with a viral pathogen.

65. The method of any one of claims 1 to 64 wherein the subject is infected with a fungal pathogen.

66. The use of a lactic acid whey or a derivative thereof in the preparation of a medicament for use in the method or any one of claims 1 to 65.

67. Lactic acid whey or a derivative thereof for use in a method of any one of claims 1 to 65.

68. A composition comprising lactic acid whey or a derivative thereof and one or more therapeutic agents.

69. The composition of claim 68 for use in the method of any one of claims 1 to 65. A vaccine composition comprising lactic acid whey or a derivative thereof and one or more a. one or more therapeutic agents, or

b. one or more adjuvants, or

c. one or more immunos timulatory molecules, or

d. any combination of two or more of (a) to (c) above.

Description:
MODULATION OF TH1 AND TH2 IMMUNE RESPONSE BY LACTIC ACID WHEY

FIELD OF THE INVENTION

The present invention relates to the use of lactic acid whey or a derivative thereof for promoting a beneficial immune response in a subject in need thereof. In particular the present invention relates a method of modulating an immune response in the treatment or prevention of diseases or conditions including those caused by intracellular or extracellular pathogens, and allergens.

BACKGROUND

Immune responses are regulated by a wide range of molecular interactions. Immune health is the ability to keep these regulatory pathways in balance while still retaining the ability to respond appropriately. Imbalance of the immune system is a feature of several conditions e.g. arthritis, atherosclerosis, allergies, diabetes and psoriasis. A balanced immune response has an important role in providing effective immunity against pathogens as well as an appropriate reaction to allergens.

An immune response to a foreign antigen (peptide) requires the presence of an anitgen presenting cell to process the antigen and present it to either a T helper (Th) or a cytotoxic T (Tc) cell. These T cells will then become activated and secrete cytokines (immune hormones). If the foreign antigen comes from a bacteria or allergen then normally Th cells are activated. If the foreign antigen comes from a virus (or intracellular bacteria) then Tc get activated, with the help of Th cells.

In immune responses to extracellular infections (e.g. bacteria) and also to most allergens, Th cells get activated and then B cells (with antibody on their surface) subsequentiy become activated. These B cells will then 'switch' their antibody from IgM isotype to another isotype such as IgG, IgA or IgE and they will secrete large amounts of these antibodies off their surface into the blood. Those antibodies bind to the foreign antigen and act as a signal for white blood cells to engulf the bacteria or parasite and kill it. Antibodies can also signal natural killer cells and macrophages to kill viral or bacterial-infected cells. In addition this response is thought to be involved in the triggering of hypersensitivity disorders, such as allergies, asthma, eczema and hay fever.

Therefore the type of immune response elicited to pathogens can be broadly divided into a cytotoxic response or an antibody response. These responses are highly regulated by Th cells and the cytokines they secrete. Thus the cytotoxic and antobody response are termed Thl and Th2, respectively, indicating the activation status of Th cells.

Differences in the Thl and Th2 immune responses are characterised by the cytokine profiles elicited, differential production of antibodies, and the activation of specific immune cells. Cells involved in a Thl immune response have been shown to produce a high level of IFN-γ, TNF-a and IL- 2 compared to those cells involved in a Th2 response, while the latter produce greater levels of IL-4, IL-5, IL-13 and IL-10 compared to those cells involved in a Thl response. A feedback control system exists between Thl and Th2 cells to regulate the response.

In light of this, it has been proposed that a balanced Thl /Th2 immune response is essential in maintaining sufficient defence against and providing an effective and timely response to a foreign agent. Modulation of the Thl and Th2 immune responses in individuals, including those infected with or exposed to pathogens and allergens, will assist in restoring immune balance and in establishing an effective and appropriate immune response.

It is an object of the present invention to provide a method of modulating an immune response useful in the prevention or treatment of various diseases and conditions, to provide methods for eliciting a beneficial immune response in subjects in need thereof, or at least provide the public with a useful choice.

SUMMARY OF THE INVENTION

In a first aspect the present invention relates to a method of modulating an immune response in a subject in need thereof, the method comprising administering lactic acid whey or a derivative thereof to the subject.

In one embodiment, the immune response is a Thl immune response.

Accordingly, the invention provides a method of modulating a Thl immune response in a subject in need thereof, the method comprising administering lactic acid whey or a derivative thereof to the subject.

In one embodiment modulation of the immune response is an upregulation of the Thl response.

In one embodiment modulation of the immune response is an upregulation of at least one Thl cytokine, such as one Thl -specific cytokine.

In one embodiment modulation of the immune response is a downregulation of the Thl response.

In one embodiment modulation .of the immune response is a downregulation of at least one Thl cytokine, such as one Thl -specific cytokine.

In one embodiment at least one Thl -specific cytokine is selected from the gtoup comprising IL-lb, IL-2, IL-3, IL-6, IL-12, IFN-γ, TNF-α or TNF-β.

In one embodiment, upregulation of a Thl response is characterised by increased IgG2a.

In one embodiment the Thl immune response is characterised by increased macrophage activation or proliferation.

In one embodiment the Thl immune response is characterised by increased CD 8+ cytotoxic T-cell activation or proliferation.

In one embodiment, the immune response is a Th2 immune response. Accordingly, the invention provides a method of modulating a Th2 response in a subject in need thereof, the method comprising administering lactic acid whey or a derivative thereof to the subject.

In one embodiment modulation of the immune response is a downregulation of the Th2 response.

In one embodiment modulation of the immune response is a downregulation of at least one Th2 cytokine, such as one Th2-specific cytokine.

In one embodiment modulation of the immune response is an upregulation of the Th2 response.

In one embodiment modulation of the immune response is an upregulation of at least one Th2 cytokine, such as one Th2-specific cytokine.

In one embodiment the at least one Th2-specific cytokine is selected from the group comprising IL-4, IL-5, IL-6, IL-10 or IL-13.

In one embodiment, upregulation of a Th2 response is characterised by increased IgGl.

In one embodiment the Th2 immune response is characterised by an upregulation of B-cells, including an activation or proliferation of B-cells and production of IgA isotype.

In one embodiment the Th2 immune response is characterised by an increase in the production of neutralizing killing antibodies.

In one embodiment modulation of the immune response is activation of regulatory T cells.

In one embodiment modulation of the immune response is upregulation of IL-17 cytokine.

Accordingly, the invention further provides a method of modulating a combined Thl and Th2 immune response in a subject in need thereof, the method comprising administering lactic acid whey or a derivative thereof to the subject.

In one embodiment modulation of the immune response is an upregulation of the Thl immune response and a downregulation of the Th2 immune response.

In one embodiment modulation of the immune response is an upregulation of the Th2 immune response and a downregulation of the Thl response.

In another embodiment modulation of the immune response is a downregulation of both the Thl and Th2 immune responses.

In one embodiment the modulation of the immune response is characterised by an increase and/ or decrease in the production of at least one Thl -specific or Th2-specific cytokine.

In one embodiment the at least one Thl -specific cytokine is selected from the group comprising IL-lb, IL-2, IL-3, IL-12, IFN-γ, TNF-a or TNF-β.

In one embodiment the at least one Th2-specific cytokine is selected from the group comprising IL-4, IL-5, IL-6, IL-10 or IL-13. In one embodiment the modulation of the immune response is characterised by a cytokine profile associated with a Th-1 response.

In one embodiment the cytokine pro file associated with a Th-1 response is characterised by an increase in the production of IL-2 and IFN-γ.

In one embodiment the modulation of the immune response is characterised by a cytokine profile associated with a Th-2 response.

In one embodiment the cytokine profile associated with a Th-2 response is characterised by an increase in the production of IL-4 and IL-10.

In various embodiments, upregulation of a Thl response is characteristic of a cell-mediated immune response.

In various embodiments, upregulation of a Th2 response is characteristic of a humoral response.

Thus in various embodiments the modulation is a Thl immune response, or of a Th2 immune response, or of a combination of both a Thl immune response and Th2 immune response.

For example, the immune response is a Thl immune response without significant Th2 immune response. For example, the immune response is a Thl immune response, such as that indicated by an IFN-γ response, in the absence of a significant IgA response, or in the absence of a significant IgE response, or in the absence of a significant IgG response, including the absence of a significant IgGl response, or the absence of a significant IgG2 response, or in the absence of a significant IgM response.

In another example, the immune response is a Th2 response without significant Thl response.

It will be appreciated that the focus of the invention is to modulate an immune response so as to be effective in the treatment or prevention of the diseases or conditions described herein. It will similarly be appreciated that, given the nature of the immune response, modulating a Thl immune response may also modulate a Th2 immune response, such that the subject's response to the methods of the invention may in fact be a combination of both a Thl immune response and a Th2 immune response.

In a further aspect the present invention relates to a method of modulating an immune response of a subject to a pathogen, the method comprising administering lactic acid whey or a derivative thereof to a subject in need thereof.

In one embodiment the pathogen is a bacterial pathogen.

In one embodiment, the bacterial pathogen is selected from the group comprising Bacteroides (e.g. B.fragilis), Brucella (e.g. B. melitensis, B. ovis, B. abortus, B. suis, B. pinnipediae and B. neotomae), Chlamydia (e.g. C. trachomatis), Chlamydophila (e.g. C. pneumoniae, C. psittaci and C. abortus), Legionella (e.g. L. pneumophila), Listeria (e.g. L. monocytogene), Plasmodium (e.g. P. falciparum), Rickettsia (e.g. R rickettsii, R, akari, R conorii, R siberica, R australis, R japonica, R africae, R prowa^ekii and R y tphi) , Salmonella, Streptococcus (e.g. S. pneumonia and S. pyogenes), Bordetella (e.g. B. pertussis, B. parapertussis and B.

bronchiseptica), Bomlia (e.g. B. burgdorferi, B. B. garinii and B. valaisiand), Campylobacter (e.g. C. jejuni and C. coli), Clostridium (e.g. C. botulinum, C. difficile, C perfringens and C. tetani) , Corynebacterium (e.g. C. diptheriae), Enterococcus (e.g. E. avium, E. durans, E. faecalis, E.faecium and E. solitaries), Escherichia (e.g. E. coli), Haemophilus, Helicobacter (e.g. H. pylori), Leptospira (e.g. L, interrogans, L. kirschneri, L. noguchii, L.

alexanderi, L. weilii, L. genomospecies 1 , L. borgpetersenii, L. santarosai and L. kmetyi), Mycobacterium (e.g. M. tuberculosis, M. bovis, M. africanum, M. microti and M. leprae) , Mycoplasma (e.g. M. pnemoniae and M.

genitalium), Pseudomonas, Shigella, Staphylococcus, Trepnema (e.g. T. pallidum), Vibrio (e.g. V. cholerae and V. parahaemoyl ticus and V. vulnificus ), Yersinia (e.g. Y. enterocolitica, Y. pestis and Y. pseudotuberculosis) Ehrlichia (e.g. JE. chaffeensis, Ehrlichia phagocytophila geno group, Ehrlichia sp.), Coxiella burnetii, Leishmania sp., Toxpolasma gondii, Trypanosoma cru^i, Histoplas a sp. or Francisella tularensis.

In one embodiment the pathogen is a viral pathogen.

In one embodiment the viral pathogen is selected from the group comprising Hepatitis C, Adenoviruses, Picornaviruses including coxsackievirus, hepatitis A virus, poliovirus, Herpesviruses including epstein-barr virus, herpes simplex type 1, herpes simplex type 2, human cytomegalovirus, human herpesvirus type 8, varicella-zoster virus, Hepadnaviruses including hepatitis B virus,

Flaviviruses including hepatitis C virus, yellow fever virus, dengue virus, West Nile virus, Retroviruses including human immunodeficiency virus (HIV), Orthomyxoviruses including influenza virus,

Paramyxoviruses including measles virus, mumps virus, parainfluenza virus, respiratory syncytial virus, Papillomaviruses including papillomavirus, Rhabdoviruses including rabies virus, Togaviruses including Rubella virus, and other viruses including vaccinia, avipox, adeno-associated virus, modified Vaccinia Strain Ankara, Semliki Forest virus, poxvirus, and coronaviruses.

In one embodiment the pathogen is a fungal pathogen.

In one embodiment the fungal pathogen is selected from the group comprising Cryptococcus (C. neoformans, C. gattii), Pneumoystisi (P. carinii, P.jiroveci), Candida {C.albicans), Aspergillus (A. fumigates, A. deflectus, Aflavus, A. flampes), Acremonium, Penicillium, Paecilomyces, Coccidioides, Histoplasma capsulatum, Blastomyces dermatitidis, Microsporum (M. canis), Trichophyton (T. verrucosum, T. mentagrophytes), Epidermophyton or Malasse^ia (M. pachydermatis).

In yet a further aspect the present invention relates to a method of modulating an immune response in the treatment or prevention of an allergic condition in a subject in need thereof, the method comprising administering lactic acid whey or a derivative thereof to the subject. In yet a further aspect the present invention relates to a method of modulating an immune response in a subject in need thereof to an allergen, the method comprising administering lactic acid whey or a derivative thereof to the subject.

In one embodiment the condition is asthma. In one embodiment the condition is an atopic condition. In yet another embodiment the condition is an eosinophilia. In still another embodiment the condition is a Th2 mediated condition.

In one embodiment the condition is selected from allergic rhinitis, hay fever, atopic rhinoconjunctivitis, urticaria, asthma and atopic eczema.

In one embodiment the condition is selected from contact dermatitis, eczema, hives (urticaria), allergic conjunctivitis, hay fever, allergic rhinitis, airborne allergies including tree (e.g. birch pollen), weed (e.g. ragweed), and grass pollen allergies, latex allergies, food allergies (e.g. peanut, shellfish, milk protein), drug allergies (e.g. to penicillin), insect sting allergies (e.g. honeybee allergies, wasp allergies, hornet allergies, yellow jacket allergies, fire ant allergies), mold allergies (e.g. to alternaria, cladosporium, aspergillus, penicillium, helminthosporium, epicoccum, fusarium, mucor, rhizopus, and aureobasidium), dust mite allergies, animal allergies (e.g. household pets such as cats and dogs), allergic bronchopulmonary aspergillosis, occupational asthma, and episodic angioedema with eosinophilia.

In one embodiment the allergic condition is selected from airway, lung, blood or skin eosinophilia. In another embodiment, the eosinophilia is selected from eosmophilic ascites, eosinophilic cellulitis, eosinophilic fasciitis, eosinophilic gastroenteritis, coeliac disease, allergic colitis, eosinophilic esophagitis, eosinophilic pancreatitis, eosinophilic pneumonias, bronchiectasis, eosinophilic synovitis, nasal eosinophilia, tropical pulmonary eosmophilia, Churg Strauss syndrome, pulmonary eosinophilia, idiopathic hyper-eosinophilic syndrome, inflammatory bowel disease, eosinophilic cholangitis, eosinophilic leukaemia and other eosinophilic cancers, familial (hereditary eosinophilia), eosinophilic granuloma, eosinophilia-myalgia syndrome, cystic fibrosis, nasal polyposis, eosinophil meningitis, Wegener's granulomatosis, polyarteritis nodosa, pemphigus vulgaris, bullous pemphigoid, dermatitis herpetiformis, erythema multiforme, eosmophilic cellulites, parasitic infections (Ascaris Toxocara canis, Filariasis, Anchylostomiasis, Trichinosis, Strong loidiasis, Fascioliasis, Schistosomiasis).

In one embodiment the subject is susceptible to anaphylaxis. For example, in one embodiment the administration of the lactic acid whey or a derivative thereof is prophylactic administration.

In one embodiment the subject is undergoing an allergy de-sensitisation regime. In one embodiment the condition is selected from Th2 mediated asthma, allergies, eczema, microbial or parasite infection, and autoimmune diseases including ulcerative colitis. In yet a further aspect the present invention relates to a method of modulating an immune response in a subject in need thereof, the method comprising co-administration of lactic acid whey or a derivative thereof and a therapeutically active agent to the subject.

In one embodiment the therapeutically active ingredient is an antigen.

In one embodiment the antigen is capable of modulating or eliciting a Thl immune response.

In one embodiment the antigen is capable of modulating or eliciting a Th2 immune response.

In one embodiment the antigen is derived from a bacterial pathogen.

In one embodiment, the bacterial antigen is an antigen derived from a bacterial pathogen selected from the group comprising Bacteroides (e.g. B.fragilis), Brucella (e.g. B. melitensis, B. ovis, B. abortus,

B. suis, B. pinnipediae and B. neoto ae), Chlamydia (e.g. C. trachomatis), Chlamydophila (e.g. C. pneumoniae, C. psittaci and C. abortus), Legionella (e.g. L pneumophila), Listeria (e.g. L. monocytogene), Plasmodium (e.g. P. falciparum), Rickettsia (e.g. R. rickettsii, R. akari, R. conorii, R. siberica, R, australis, R. japonica, R. africae, R. prowa^ekii and R. typhi) , Salmonella, Streptococcus (e.g. S. pneumonia and S. pyogenes), Bordetella (e.g. B.

pertussis, B. parapertussis and B. hronchiseptica), Borrelia (e.g. B. burgdorferi, B. B. garinii and B.

valaisiana), Campylobacter (e.g. C. jejuni and C. coli), Clostridium (e.g. C. botulinum, C. difficile, C. perfringens and

C. tetani) , Corynebacterium (e.g. C. diptheriae), Enterococcus (e.g. E. avium, E. durans, E.faecalis, E.faecium and E. solitaries), Escherichia (e.g. E. coli), Haemophilus, Helicobacter (e.g. H. pylori), Leptospira (e.g. L. interrogans, L. kirschneri, L. noguchii, L. alexanderi, L. weilii, L. genomospecies 1, L botgpetersenii, L santarosai and L kmetyi), Mycobacterium (e.g. M. tuberculosis, M. bo is, M. africanum, M. microti and M. leprae) , Mycoplasma (e.g. M. pnemoniae and M. genitalium), Pseudomonas, Shigella, Staphylococcus, Trepnema (e.g. T. pallidum), Vibrio (e.g. V. cholerae and V. parahaemoyl ticus and V. vulnificus ), Yersinia (e.g. Y. enterocolitica, Y. pestis and Y.

pseudotuberculosis) Ehrlichia (e.g. E. chaffeensis, Ehrlichia phagocytophila geno group, Ehrlichia sp.), Coxiella burnetii, Leishmania sp., Toxpolasma gondii, Trypanosoma cruri, Histoplasma sp. or Francisella tularensis.

In one embodiment, the antigen is derived from a viral pathogen.

In one embodiment, the viral antigen is an antigen derived from a viral pathogen selected from the group comprising Hepatitis C, Adenoviruses, Picornaviruses including coxsackievirus, hepatitis A virus, poliovirus, Herpesviruses including epstein-barr virus, herpes simplex type 1, herpes simplex type 2, human cytomegalovirus, human herpesvirus type 8, varicella-zoster virus,

Hepadnaviruses including hepatitis B virus, Flaviviruses including hepatitis C virus, yellow fever virus, dengue virus, West Nile virus, Retroviruses including human immunodeficiency virus (HIV),

Orthomyxoviruses including influenza virus, Paramyxoviruses including measles virus, mumps virus, parainfluenza virus, respiratory syncytial virus, Papillomaviruses including papillomavirus, Rhabdoviruses including rabies virus, Togaviruses including Rubella virus, and other viruses including vaccinia, avipox,

In one embodiment the antigen is derived from a fungal pathogen.

In one embodiment the fungal antigen is derived from a fungal pathogen selected from the group comprising Cryptococcus (C. neoformans, C. gattii), Pneumocystis} (P. carinii, P.jirovect), Candida

{C. albicans), Aspergillus {A. fumigates, A deflectus, A.flavus, A. flavipes), Ammonium, Penicillium, Paecilomyces, Coccidioides, Histoplasma capsulatum, Blastomyces dermatitidis, Microspo m (M. canis), Trichophyton (T. verrucosum, T. mentagrophytes), Epidermophyton or Malasse^ia (M. pachydermatis).

In one embodiment the composition is a vaccine.

In one embodiment the composition may additionally comprise an adjuvant, or an immunostimulatory molecule.

In various embodiments the co-administration of the lactic acid whey or a derivative thereof and the one or more therapeutically active agent is separate, simultaneous or sequential.

In one embodiment the co-administration of the lactic acid whey or a derivative thereof and the one or more therapeutically active agents is capable of modulating an immune response to a bacterial pathogen.

In one embodiment the co-administration of the lactic acid whey or a derivative thereof and the one or more therapeutically active agents is capable of modulating an immune response to a viral pathogen.

In one embodiment the co-administration of the lactic acid whey or a derivative thereof and the one or more therapeutically active agents is capable of modulating an immune response to a fungal pathogen.

The use of a lactic acid whey or a derivative thereof in the preparation of a medicament for use in any of the methods described above, such as for modulating an immune response in a subject including a subject infected with a bacterial or viral pathogen, or exposed to an allergen, for example, is also contemplated.

Lactic acid whey or a derivative thereof for use in any of the methods or treatments described above is also contemplated. For example, lactic acid whey or a derivative thereof for modulating an immune response in a subject in need thereof is specifically contemplated.

Additionally, a composition comprising lactic acid whey or a derivative thereof and one or more therapeutic agents for use in any of the methods or treatments described above is also

contemplated. For example, a composition comprising lactic acid whey or a derivative thereof and one or more therapeutic agents in modulating an immune response in a subject in need thereof is also contemplated.

The following embodiments may relate to any of the above aspects. In various embodiments the subject has an elevated Thl response. In another embodiment the subject has a suppressed Th2 response.

In various embodiments the subject has an elevated Th2 response. In another embodiment the subject has a suppressed Thl response.

In various embodiments the subject is an infant or neonate mammal.

In various embodiments, the subject is infected with a bacterial pathogen.

In various embodiments the subject is infected with a viral pathogen.

In various embodiments the subject is infected with a fungal pathogen.

In various embodiments, the viral pathogen or bacterial pathogen is selected from the group comprising Bacteroides (e.g. B.fragilis), Brucella (e.g. B. melitensis, B. ovis, B. abortus, B. suis, B. pinnipediae and

B. neotomae), Chlamydia (e.g. C. trachomatis), Chlamydophila (e.g. C. pneumoniae, C. psittaci and C. abortus), Legionella (e.g. E pneumophila), Listeria (e.g. E monocytogene), Plasmodium (e.g. P. falciparum), Rickettsia (e.g. R. rickettsii, R, akari, R. conorii, R, siberica, R. australis, R. japonica, R. africae, R. prowa^ekii and R. typhi) , Salmonella, Streptococcus (e.g. S. pneumonia and S. pyogenes , Bordetella (e.g. B. pertussis, B. parapertussis and B. bronchiseptica), Borrelia (e.g. B. burgdorferi, B. B. garinii and B. valaisiand), Campylobacter (e.g. C. jejuni and C. coli), Clostridium (e.g. C botulinum, C. difficile, C. perfringens and C. tetani) , Corynebacterium (e.g. C. diptheriae), Enterococcus (e.g. E. avium, E. durans, E.faecalis, E.faecium and E. solitaries), Escherichia (e.g. E. coli), Haemophilus, Helicobacter (e.g. H. pylori), Leptospira (e.g. E interrogans, E kirschneri, E. noguchii, E alexanderi, L. weilii, L. genomospecies 1 , E borgpetersenii, E. santarosai and E kmetyi), Mycobacterium (e.g. M. tuberculosis, M. bovis, M. africanum, M. microti and M. leprae) , Mycoplasma (e.g. M. pnemoniae and M.

genitalium), Pseudomonas, Shigella, Staphylococcus, Trepnema (e.g. T. pallidum), Vibrio (e.g. V. cholerae and V. parahaemoyl ticus and V. vulnificus ), Yersinia (e.g. Y. enterocolitica, Y. pestis and Y . pseudotuberculosis) Ehrlichia (e.g. E. chaffeensis, Ehrlichia phagocytophila geno group, Ehrlichia sp.), Coxiella burnetii, Eeishmania sp., Toxpolasma gondii, Trypanosoma cru^i, Histoplasma sp. or Francisella tularensis and viruses including Hepatitis

C, Adenoviruses, Picornaviruses including coxsackievirus, hepatitis A virus, poliovirus, Herpesviruses including epstein-barr virus, herpes simplex type 1 , herpes simplex type 2, human cytomegalovirus, human herpesvirus type 8, varicella- zoster virus, Hepadnaviruses including hepatitis B virus,

Flaviviruses including hepatitis C virus, yellow fever virus, dengue virus, West Nile virus, Retroviruses including human immunodeficiency virus (HIV), Orthomyxoviruses including influenza virus,

Paramyxoviruses including measles virus, mumps virus, parainfluenza virus, respiratory syncytial virus, Papillomaviruses including papillomavirus, Rhabdoviruses including rabies virus, Togaviruses including Rubella virus, and other viruses including vaccinia, avipox, adeno-associated virus, modified Vaccinia Strain Ankara, Semliki Forest virus, poxvirus, and coronaviruses.

In various embodiments the subject suffers from or is susceptible to an allergic condition. In various embodiments the allergic condition is asthma. In one embodiment the condition is an atopic condition. In yet another embodiment the condition is an eosinophilia. In still another embodiment the allergic condition is a Th2 mediated condition.

In various embodiments the condition is selected from allergic rhinitis, hay fever, atopic rhinoconjunctivitis, urticaria, asthma and atopic eczema.

In various embodiments the condition is selected from contact dermatitis, eczema, hives (urticaria), allergic conjunctivitis, hay fever, allergic rhinitis, airborne allergies including tree (e.g. birch pollen), weed (e.g. ragweed), and grass pollen allergies, latex allergies, food allergies (e.g. peanut, shellfish, milk protein), drug allergies (e.g. to penicillin), insect sting allergies (e.g. honeybee allergies, wasp allergies, hornet allergies, yellow jacket allergies, fire ant allergies), mold allergies (e.g. to alternaria, cladosporium, aspergillus, penicillium, helminthosporium, epicoccum, fusarium, mucor, rhizopus, and aureobasidium), dust mite allergies, animal allergies (e.g. household pets such as cats and dogs), allergic bronchopulmonary aspergillosis, occupational asthma, and episodic angioedema with eosinophilia.

In various embodiments the allergic condition is selected from airway, lung, blood or skin eosinophilia. In another embodiment, the eosinophilia is selected from eosinophilic ascites, eosinophilic cellulitis, eosinophilic fasciitis, eosinophilic gastroenteritis, coeliac disease, allergic colitis, eosinophilic esophagitis, eosinophilic pancreatitis, eosinophilic pneumonias, bronchiectasis, eosinophilic synovitis, nasal eosinophilia, tropical pulmonary eosinophilia, Churg Strauss syndrome, pulmonary eosinophilia, idiopathic hyper-eosinophilic syndrome, inflammatory bowel disease, eosinophilic cholangitis, eosinophilic leukaemia and other eosinophilic cancers, familial (hereditary eosinophilia), eosinophilic granuloma, sarcoidosis, eosinophilia-myalgia syndrome, cystic fibrosis, nasal polyposis, eosinophil meningitis, Wegener's granulomatosis, polyarteritis nodosa, rheumatoid arthritis, pemphigus vulgaris, bullous pemphigoid, dermatitis herpetiformis, erythema multiforme, eosinophilic cellulites, parasitic infections (Ascaris Toxocara canis, Filariasis, Anchylostomiasis, Trichinosis, Strongvloidiasis, Fascioliasis, Schistosomiasis).

In various embodiments the condition is selected from Th2 mediated asthma, allergies, eczema, microbial or parasite infection, and autoimmune diseases including Crohn's disease.

It is intended that reference to a range of numbers disclosed herein (for example, 1 to 10) also incorporates reference to all rational numbers within that range (for example, 1, 1.1, 2, 3, 3.9, 4, 5, 6, 6.5, 7, 8, 9 and 10) and also any range of rational numbers within that range (for example, 2 to 8j 1.5 to 5.5 and 3.1 to 4.7) and, therefore, all sub-ranges of all ranges expressly disclosed herein are hereby expressly disclosed. These are only examples of what is specifically intended and all possible combinations of numerical values between the lowest value and the highest value enumerated are to be considered to be expressly stated in this application in a similar manner. Further aspects and advantages of the present invention will become apparent from the ensuing description which is given by way of example only.

In this specification where reference has been made to patent specifications, other external documents, or other sources of information, this is generally for the purpose of providing a context for discussing the features of the invention. Unless specifically stated otherwise, reference to such external documents is not to be construed as an admission that such documents, or such sources of

information, in any jurisdiction, are prior art, or form part of the common general knowledge in the art.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 is a graph depicting influenza particles recovered from the lungs of animals challenged with influenza following vaccination and/ or feeding of dairy products.

Figure 2 is a graph depicting peritoneal macrophage assay for mice fed LPX2013 for seven days.

Figure 3 is a graph depicting Peritoneal macrophage assay for mice fed LPX2013 for seven days followed by a seven day wash out period where no LPX2013 was fed.

Figure 4 is a graph depicting splenocyte proliferation assay using lipoteichoic acid and splenocytes from animals aged 7, 15, 21, and 35 days.

DETAILED DESCRIPTION

The present invention relates to lactic acid whey or a derivative thereof for modulating an immune response in a subject in need thereof.

1. Definitions

The term "comprising" as used in this specification means "consisting at least in part of.

When interpreting statements in this specification which include that term, the features, prefaced by that term in each statement or claim, all need to be present but other features can also be present.

Related terms such as "comprise" and "comprised" are to be interpreted in the same manner.

The term "effective amount" is the amount required to confer therapeutic effect. The interrelationship of dosages for animals and humans (based on milligrams per meter squared of body surface) is described by Freireich, et al. (1966). Body surface area can be approximately determined from height and weight of the subject. See, e.g., Scientific Tables, Geigy Pharmaceuticals, Ardley, New York, 1970, 537. Effective doses also vary, as recognized by those skilled in the art, dependent on route of administration, excipient usage, and the like.

The term "lactic acid whey" as used herein means a product that is produced by fermentation of milk, including skim milk, with lactic acid bacteria, for example, during the manufacture of casein, caseinate or cottage and ricotta cheeses. In contrast, mineral acid whey is produced by addition of mineral acids to milk, including skim milk, for example during casein and caseinate manufacture. The basis of these two processes is to lower pH to about 4.6 to cause casein to coagulate as opposed to using the action of chymosin to cause coagulation. Therefore any acid-coagulated milk product that has not been exposed to chymosin will not contain the derivative peptide casein glycomacropeptide. Lactic acid whey and mineral acid whey do not contain casein glycomacropeptide.

The term "derivative of lactic acid whey" as used herein means a product derived from applying to lactic acid whey any of the commonly-used processes used in the dairy whey processing industry, such as a process to concentrate, fractionate or functionally enhance dairy whey. Concentration should be taken to mean any process that increases the level of solids in a feed whey stream from the native level or from that present in the feed whey stream at input, and may be accomplished by evaporation, membrane separation processes (including reverse osmosis, micro filtration and ultrafiltration), dehydration/drying processes and other processes known in the art. Fractionation processes intended are those that preferentially separate the inert or undesired elements of the whey stream from its functional or desired elements, including, for example, removal of superfluous lactose by crystallisation, removal of unwanted fat by defatting (solvent extraction or other means known in the art), or preferential retrieval of functional protein isolates by affinity processes (affinity chromatography using appropriate media for whey proteins). Functional enhancement processes material to the current invention include demineralisation, remineraHsation (ie changing the level of mineral species present in the product from their native levels and proportionate ratios, for example, by either dosing or by ion exchange), hydrolysis (of protein and/ or lactose, by enzymic or chemical means generally known) or enzymic protein cross-linking. It is intended that in various representative embodiments a derivative of lactic acid whey is accomplished by performing any of these operations singly or by combining any two or more of these operations to produce a derivative of lactic acid whey.

The term "modulating an immune response" (and different tenses of these terms) refers to the ability of an agent to stimulate the generation of antigen-specific cytolytic activity (the activity of immune cells, particularly cytotoxic T-lymphocytes) and/ or NK cell activity, improve the cellular immune response to antigens (through the activity of at least cytotoxic T-lymphocytes), improve immune protection (by at least restoring the activity of cytotoxic T-lymphocytes and/ or NK cells and enhancing cytokine production), restore immune protection (by at least restoring or stimulating the activity of cytotoxic T- lymphocytes and/or NK cell activity and enhancing cytokine production) or generate pro-inflammatory and immunoregulatory mediators (Thl and Th2 cytokines).

The term "oral administration" includes oral, buccal, enteral and intra-gastric administration.

A "subject" is an animal, preferably a mammal, more preferably a mammalian companion animal or human. Preferred companion animals include cats, dogs and horses. In one embodiment the human is an adult, a child, or an infant, or an immunocompromised adult, child, or infant. The term "treat" and its derivatives should be interpreted in their broadest possible context. The term should not be taken to imply that a subject is treated until total recovery. Accordingly, "treat" broadly includes maintaining a subject's disease progression or symptoms at a substantially static level, increasing a subject's rate of recovery, amelioration and/or prevention of the onset of the symptoms or severity of a particular condition, or extending a patient's quality of life. The term "treat" also broadly includes the maintenance of good health for sensitive individuals and building stamina for disease prevention.

2. Immune response

2.1 Thl/Th2 Immune modulation

The present inventors have found that lactic acid whey or a derivative thereof, optionally with at least one additional therapeutic agent, is able to modulate an immune response. In particular, as shown in the examples below, lactic acid whey or a derivative thereof, optionally with at least one therapeutic agent, is able to stimulate the generation of antigen-specific cytolytic activity (the activity of immune cells, particularly cytotoxic T-lymphocytes) and/ or NK cell activity, improve the cellular immune response to antigens (through the activity of at least cytotoxic T-lymphocytes), improve immune protection (by at least restoring the activity of cytotoxic T-lymphocytes and/ or NK cells and enhancing cytokine production), restore immune protection (by at least restoring or stimulating the activity of cytotoxic T-lymphocytes and/ or NK cell activity and enhancing cytokine production) and generate pro-inflammatory and immunoregulatory mediators (Thl and Th2 cytokines). It is believed that any derivative of lactic acid whey, including concentrates, isolates, hydrolysates and the like, whether or not in combination with at least one additional therapeutic agent, will exhibit similar activity as the lactic acid whey from which it derives.

As shown in the Examples below, lactic acid whey or a derivative thereof is effective in improving the generation of antigen- specific cytolytic activity and/ or NK cell activity, improving the cellular immune response to antigens, improving immune protection and restoring immune protection.

Accordingly, the present invention relates to a method of modulating the immune response of a subject comprising administration of lactic acid whey or a derivative thereof, optionally with at least one additional therapeutic agent, such as a bacterial, viral or fungal antigen, to the subject. The present invention also relates to methods of modulating the Thl and Th2 immune response in a subject. Accordingly, the invention provides for a method of upregulating or downregulating the production of cytokines associated with the Thl and Th2 immune response of a subject.

In one embodiment the Thl cytokine is selected from IL-lb, IL-2, IL-3, IL-6, IL-12, IFN-γ, TNF-a or TNF-β. In one embodiment the Th2 cytokine is selected from IL-4, IL-5, IL-6, IL-10 or IL- 13. In one embodiment the level of Thl cytokine or cytokines is increased to at least about 2, 3, 4, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750 or 800-fold its base state.

In one embodiment the level of Th2 cytokine or cytokines is decreased to at least about 2, 3, 4, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750 or 800-fold its base state.

2.2 Thl response and cell-mediated immunity

The Thl immune response leads to activation of macrophages and consequently cell-mediated immunity. Pathogenic antigens are expressed on the surface of antigen presenting cells (such as macrophages, B-lymphocytes, and dendritic cells), bound to either major histocompatibility MHC Class I or MHC Class II molecules. Presentation of pathogenic antigen coupled to MHC Class II activates a helper (CD4+) T-cell response. Upon binding of the T-cell to the antigen-MHC II complex, CD4+ T- cells proliferate, releasing cytokines, including interferon-gamma (IFN-γ) and interleukin 2 (IL-2), IL-4, IL-7, and IL-12.

Presentation of pathogenic antigens bound to MHC Class I molecules activates a cytotoxic (CD8+) T-cell response. Upon binding of the T-cell to the antigen-MHC I complex, CD8+ cells secrete perforin, resulting in pathogen cell lysis, swelling and death. Alternatively, CD8+ cells induce programmed cell death or apoptosis. Activation of CD8+ T-cells is amplified by the release of specific cytokines by CD4+ T-cells.

A cell-mediated immune response is believed to be central to the immunity against intracellular pathogens.

Methods to assess and monitor the onset or progression of a Thl immune response in a subject are well known in the art. Convenient exemplary methods include those in which the presence of or the level of one or more cytokines associated with a Thl response, such as those identified herein, is . assessed. Similarly, cell-based methods to assess or monitor the onset and progression of a Thl response are amenable to use in the present invention, and may include cell proliferation or activation assays, including assays targeted at identifying activation or expansion of one or more populations of immune cells, such as T-lymphocytes. Likewise, assays to determine upregulation of Thl -associated antibodies, such as, for example IgG2a, are well known.

In certain embodiments, methods of the invention that elicit both a Thl immune response and a Th2 are preferred.

In other embodiments, methods of the invention that elicit predominantly a Thl response are preferred. Such methods may include those that elicit a Thl immune response without a significant Th2 response, or without any detectable Th2 response. In one example, the immune response is a Thl immune response, such as that indicated by an IFN-γ response, often in the presence of a significant IgA response, or in the absence of a significant IgE response, or in the absence of a significant IgG W 201

response, including the absence of a significant IgGl response, or the absence of a significant IgG2 response.

2.3 Humoral response

The Th2 immune response leads to activation of B-cells, which produce specific antibodies that drive humoral immunity. The secreted antibodies bind to antigens presented on the surface of invading pathogens, flagging them for destruction.

It has been suggested that a combined Thl and Th2 immune response (as a consequence of an initiated Thl response) would be beneficial to achieve a more highly sensitive immune response to or enhance the level of protection against intracellular pathogens.

Again, methods to assess and monitor the onset or progression of a Th2 response are well known in the art. These include antibody binding assays, ELISA, skin-prick tests and the like.

3. Methods of assessing modulation of an immune response

Methods of assessing modulation of an immune response are well known in the art. Commonly, immunological assays are used for measuring specific immune response markers, such as, cytokines, antibodies and T-cells. Examples of assays include enzyme-linked immunosorbent assay (ELISA), liquid array multiplexing system (LAMS), ELISPOT and cell proliferation assays.

In addition to these qualitative and quantitative assays, physical and behavioural observations may also be used an indicative measures of an immune response.

4. Methods of producing lactic acid whey

4.1 Lactic acid bacteria

Lactic acid bacteria (LAB) comprise a group of bacteria with the unifying feature of converting carbohydrate into lactic acid. During fermentation of milk, LAB convert pH neutral lactose to acidic lactic acid, causing the pH of the milk to drop.

A number of different genera of lactic acid bacteria exist within the phylum Firmicutes and include Lactobacillus spp., lueuconostoc spp., Pediococcus spp., Lactococcus spp., Streptococcus spp., Aerococcus spp., Carnobactenum spp., Enterococcus spp., Oenococcus spp., Sporolactobacillus spp., Teragenococcus spp., Vagococcus spp., or Weisella spp.

Other lactic acid-producing Gram-positive bacteria belong to the phylum Actinobacteria and include the genera Aerococcus, Microbacterium and Vropionibacterium and bifidobacterium.

The use of lactic acid bacteria in industry, for example in food preparation or manufacture, has generally focussed on bacteria from the genera Lactococcus, Streptococcus, Enterococcus, Lactobacillus, Leuconostoc, Bifidobacterium and Pediococcus In the context of dairy fermentations, Streptococcus, Lactobacillus, Lactococcus, Leuconostoc and Bifidobacteria species are generally of greatest interest.

Exemplary Bifidobacteria include Bifidobacterium adolescentis, Bifidobacterium bifidum, Bifidobacterium animalis, Bifidobacterium thermophilum, Bifidobacterium breve, Bifidobacterium longum, Bifidobacterium infantis and Bifidobacterium lactis. Exemplary specific strains of Bifidobacteria include Bifidobacterium breve strain Yakult, Bifidobacterium breve R070, Bifidobacterium lactis Bbl2, Bifidobacterium longum R023, Bifidobacterium bifidum R071, Bifidobacterium infantis R033, Bifidobacterium longum BB536 and Bifidobacterium longum SBT- 2928.

Exemplary Lactobacilli include Lactobacillus acidophilus, Lactobacillus brevis, Lactobacillus bulgaricus, Lactobacillus casei, Lactobacillus cellobiosus, Lactobacillus crispatus, Lactobacillus curvatus, Lactobacillus fermentum, Lactobacillus GG (Lactobacillus rhamnosus or Lactobacillus casei subspecies rhamnosus), Lactobacillus gasseri, Lactobacillus johnsonii, Lactobacillus plantarum and Lactobacillus salivarus. Lactobacillu plantarum 299v strain originates from sour dough. Lactobacillus plantarum itself is of human origin. Other exemplary strains of Lactobacillus are Lactobacillus acidophilus BG2F04, Lactobacillus acidophilus INT-9, Lactobacillus plantarum ST31, Lactobacillus nuteri, Lactobacillus johnsonii LAI, Lactobacillus acidophilus NCFB 1748, Lactobacillus casei Shirota, Lactobacillus acidophilus NCFM, Lactobacillus acidophilus DDS-1, Lactobacillus delbrueckii subspecies delbrueckii, Lactobacillus delbrueckii subspecies bulgaricus type 2038, Lactobacillus acidophilus SBT-2062, Lactobacillus brevis, Lactobacillus salivarius UCC 118 and Lactobacillus paracasei subsp paracasei F\9.

Exemplary Lactococci include Lactococcus lactis, Lactococcus lactis subspecies cre oris {Streptococcus cremoris), Lactococcus lactis subspecies lactis NCDO 712, Lactococcus lactis subspecies lactis NIAI 527, Lactococcus lactis subspecies lactis NIAI 1061, Lactococcus lactis subspecies lactis biovar diacetylactis NIAI 8W and Lactococcus lactis subspecies lactis biovar diacetylactis ATCC 13675.

Exemplary Streptococci include Streptococcus thermophilus, and Streptococcus salivarus, including Streptococcus salivarus subspecies thermophilus type 1131.

Dairy starter cultures generally comprise lactic acid bacteria. In the present context, the expression "lactic acid bacteria" (LAB) designates a group of Gram positive, catalase negative, non- motile, non-sporulating, microaerophilic or anaerobic bacteria. These bacteria ferment sugars to produce organic acids, predominantly lactic acid. In the case of hetero-fermentative LAB, formic acid and propionic acid may also be produced.

4.2 Lactic Acid Whey

Lactic acid whey, or lactic whey is one product of natural fermentation of milk with lactic acid bacteria, and in the current context is the liquid that is separated from curds when lactic casein or a non-renneted fresh cheese (eg cottage cheese or quarg) is produced by LAB fermentation of milk or its direct derivatives. In a preferred embodiment lactic whey is derived from the manufacture of lactic casein, the manufacture of which is detailed in the Dairy Processing Handbook (1995) Publ Tetra Pak Processing Systems AB S-221 86 Lund, Sweden (p397) and will be well known to those skilled in the art. For the manufacture of lactic casein the preferred LAB are sub-species of lMctococcus lactis, notably strains of L lactis ssp cremoris and JL. lactis ssp lactis and the milk substrate is low fat or skim milk.

In another embodiment the lactic whey is derived from the manufacturer of fresh (unrenneted) cheese such as cottage cheese or quarg. Again the details for manufacture of these standard products are detailed in the Dairy Processing Handbook (1995) (p323 for cottage cheese, p 325 for quarg) and will be well known to those skilled in the art. In the manufacture of fresh cheese mixed LAB cultures typically containing strains of L lactis ssp cremons and L» lactis ssp lactis are generally used, and the milk substrate may be whole milk or fat adjusted (increased or decreased) milk.

Lactic whey may also be recovered from strained naturally fermented yoghurts such as Greek yoghurt or Labneh, also described in the Dairy Processing Handbook (1995) as "concentrated yoghurt" (p243).

Native lactic whey is predominantly water and lactose, but also contains the majority of the milk serum proteins, those milk proteins that are not precipitated at pH 4.6 (i.e. the non-casein proteins) comprising a-lactalburnin, β-lactoglobulin, bovine serum albumins (BSA), immunoglobulins, lactoferrin, lactoperoxidase, lysozyme, etc. A detailed review of the components and composition of wheys of varying origin is provided in De Witt, J N (2001): Lecturer's Handbook on Whey and Whey Products, First Edition, Publ European Whey Products Association, Brussels. Belgium. Lactic whey also contains breakdown products of the lactic acid.bacteria originally added to the milk as starter culture to facilitate production of lactic acid from lactose.

4.3 Lactic WPC

Lactic WPC (Whey Protein Concentrate) is produced by further treatment of lactic whey through membrane filtration to concentrate the protein by removing as "permeate" some of the low molecular weight solutes in the whey, particularly lactose and dissolved minerals, with some of the whey's water content. WPC commercial products are generally concentrated to a protein in dry matter level greater than 30%, and provided in dry form after spray drying. Herein, any lactic whey that has been concentrated to provide protein levels greater than the native levels in the recovered lactic whey, whether liquid or dried, shall be considered lactic WPC.

The manufacturing processes by which WPC's are produced are detailed in De Witt, J N (2001): Lecturer's Handbook on Whey and Whey Products, First Edition, Publ European Whey Products Association, Brussels. Belgium.and in the Dairy Processing Handbook (1995) in Chapter 15 Whey Processing, Protein Recovery by UF and will be familiar to those skilled in the art. 4.4 Lactic WPI

Lactic WPI (Whey Protein Isolate) is produced by further treatment of (liquid) lactic WPC through either further membrane filtration, for example microfiltration, or by chromatographic isolation. By these processes the protein of lactic whey is further concentrated to about 90% through diafiltration and ultrafiltration (membrane filtration) or affinity isolation on ion exchange resins (chromatographic isolation).

5. Other therapeutic agents

Additional therapeutic agents useful in the present invention include antigens to various bacterial, viral or fungal antigens. Various pathogens from which these antigens may be derived are listed below.

The therapeutic agents of the present invention can be co-administered with the lactic acid whey or a derivative thereof, either separately, simultaneously or sequentially. For example, the lactic acid whey or a derivative thereof can be co-administered as an adjuvant to a vaccine to provide an enhanced immune response. Alternatively, the lactic acid whey or a derivative thereof of the present invention can be sequentially administered with anti-allergy medications known to those skilled in the art for use in the treatment or prevention of the various allergic conditions as described herein.

6. Pathogens and disease states to be treated

It will be appreciated that compositions of the present invention are in part directed to the prevention or treatment of diseases caused by pathogens, including intracellular and extracellular bacteria, viruses and fungal pathogens. Representative pathogens are described in more detail below, but those skilled in the art will appreciate that the invention has application in the treatment or prevention of any disease or condition associated with a pathogen.

Intracellular bacteria include, but are not limited to Brucella, Campylobacter , Chlamydia,

Chlamydophila, Eschemhia, Haemophilus, Legionella, Listeria, Mycobacterium, Mycoplasma, Plasmodium, ickettsia, Salmonella, Shigella, Streptococcus and Treponema.

Brucella is a genus of Gram-negative non-motile, non-encapsulated coccobacilli. Brucella is the cause of brucellosis. Examples of different Brucella species include B. melitensis and B. ovis which infect ovine species, B. abortus which infects cattle, B. suis which infects swine species, B. pinnipediae isolated from marine mammals and B. neotomae. Humans typically become infected through contact with fluids from infected animals (sheep, cattle or pigs) or derived food products such as unpasteurized milk and cheese.

Campylobacter is a Gram-negative spiral bacteria with either a single or bi-polar flagella.

Campylobacter are best known for causing foodborne diseases in humans, resulting in diarrhoea, periodontistis and fever. Although over a dozen species have been implicated in human disease, the most common species include C jejuni and C. colt.

Chlamydia refers to a genus of bacteria, which includes the human pathogen Chlamydia trachomatis. Chlamydophila is a related bacterium, which includes the human pathogens Chlamydophila pneumoniae, causing pneumonia, Chlamydophila psittaci, causing respiratory psittacosis, and Chlamydophila abortus, which is associated with abortion in humans.

Escherichia is a genus of Gram-negative rod-shaped bacteria known to cause urinary tract infections and various gastrointestinal diseases. E. coli is most commonly responsible for pathogenesis in humans.

Haemophilus is a genus of Gram-negative coccobacilli bacteria that cause sepsis and bacterial meningititis.

Eegionella is a Gram-negative bacterium. The most notable species, L. pneumophila causes legionellosis or Legionnaires' disease.

Usteria species are Gram-positive bacilli. The most known pathogen in this genus is L.

monocytogenes, the causative agent of listeriosis. Usteria ivanovii is a pathogen of ruminants and is only rarely the cause of human disease.

Mycobacterium is a genus of Actinobacteria. The genus includes pathogens known to cause serious diseases in mammals, including tuberculosis and leprosy. Examples of pathogen species include M. tuberculosis, M. boms, M. africanum, M. microti; M. leprae (leprosy), M. avium paratuberculosis (associated with Crohn's disease in humans and Johne's disease in sheep).

Mycoplasma is a genus of bacteria that do no exhibit a cell wall and as a result many antiboiotics are ineffective in treating disease caused by Mycoplasma. Pathogenic species include M. pnemoniae (pneumonia) and M. genitalium (pelvic inflammatory diseases).

Plasmodium is a genus of parasitic protozoa. Infection with these parasites is known to cause malaria {P. falciparum).

Rickettsia is a genus of motile, Gram-negative, non-spore forming bacteria. Rickettsia species are carried as parasites by many ticks, fleas, and lice, causing diseases such as Rocky Mountain spotted fever R. rickettsii), Rickettsialpox (R akari), Boutonneuse fever (R. conorii), Siberian tick typhus (R, sibericd), Australian tick typhus (R. australis), Oriental spotted fever (R. japonicd), African tick bite fever (K africae), Epidemic typhus (R, and Endemic typhus (R. typhi

Salmonella is a genus of rod-shaped, Gram-negative, non-spore forming, motile enterobateria that cause illnesses in humans and many animals, including typhoid fever, paratyphoid fever, and the salmonellosis. Shigella is a genus of Gram-negative, non-spore forming rod-shaped bacteria closely related to Escherichia coli and Salmonella. Shigella is the causative agent of human shigellosis (dysentery), infecting only primates but not other mammals.

Streptococcus is a genus of spherical Gram-positive bacteria known to cause a number of human diseases including meningitis, bacterial pneumonia (S. pneumoniae), endocarditis, erysipelas and necrotizing fasciitis {S. pyogenes).

Treponema is a genus of bacteria that includes the species T. pallidum, which is the causative bacterium of syphilis.

Extracellular bacteria include, but are not limited to Bordetella, Borrelia, Clostridium,

Corynebacterium, Enferococcus, Helicobacter, Leptospira, Pseudomonas, Staphylococcus, Vibrio and Yersinia.

Bordetella is a genus of Gram-negative coccobacilli known to cause pertussis or whooping cough in humans. Species of pathogen Bordetella include B. pertussis, B. parapertussis and B. bronchiseptica.

Borrelia is a spirochete bacteria known to cause Lyme disease or borreliosis, a disease transmitted by ticks. The major pathogenic species include B. burgdorferi, B. fyelii, B. garinii and B. valaisiana.

Clostridium is a genus of Gram-positive anaerobic bacteria. The genus comprises over 100 species, although four main species are pathogenic in humans - C. botulinum (botulism), C. difficile (colitis), C. perfringens (food poisoning and gas gangrene) and C. tetani (tetanus).

Corynebacterium is a genus of Gram-positive rod-shaped bacteria. Although many strains are innocuous, C. diptheriae is pathogenic and causes diphtheria in humans.

Enterococcus is a genus of Gram-positve lactic acid bacteria that aire known to cause urinary tract infections, bacteremia, bacterial endocarditis, diverticulitis and meningitis. Species include E. avium, E. durans, E.faecalis, E.faecium and E. solitarius.

Helicobacter is a genus of helical-shaped Gram-negative bacteria. The most widely known pathogenic species is H. pylori, which causes peptic ulcers, chronic gastritis, duodenitis and stomach cancer.

Leptospira are a genus of Gram-negative bacteria. Pathogenic species include L. interrogans, L kirschneri, JL noguchii, L alexanderi, L weilii, L. genomospecies 1, L borgpetersenii, L santarosai and JL kmetyi.

Pseudomonas is a genus of Gram-negative bacteria and includes strains of bacteria previously classified as Chryseomonas and Flavimonas. P. aeruginosa is known to cause sepsis and inflammation in many human organs.

Staphylococcus is a genus of Gram-positive bacteria and is a common cause of food poisoning.

Vibrio is a genus of Gram-negative bacteria known to cause a number of seafood-borne illnesses in humans, such as gastroenteritis, septicaemia and various wound infections. Pathogenic species include V. cholerae (cholera), V. parahae olyticus and V. vulnificus. Yersinia is a Gram-negative rod shaped bacteria. Specific human pathogens include Y. enterocolitica, causing Yersiniosis. Y. pestis, the causative agent of plague and the least common pathogen. Y.

pseudotuberculosis. Yersinia is implicated as one of the pathogenic auses of Reactive Arthritis.

The treatment or prevention of diseases associated with viral pathogens and fungal pathogens are also contemplated. Viral pathogens include Hepatitis C, Adenoviruses, Picornaviruses including coxsackievirus, hepatitis A virus, poUovirus, Herpesviruses including epstein-barr virus, herpes simplex type 1, herpes simplex type 2, human cytomegalovirus, human herpesvirus type 8, varicella-zoster virus, Hepadnaviruses including hepatitis B virus, Flaviviruses including hepatitis C virus, yellow fever virus, dengue virus, West Nile virus, Retroviruses including human immunodeficiency virus (HIV),

Orthomyxoviruses including influenza virus, Paramyxoviruses including measles virus, mumps virus, parainfluenza virus, respiratory syncytial virus, Papillomaviruses including papillomavirus,

Rhabdoviruses including rabies virus, Togaviruses including Rubella virus, and other viruses including vaccinia, avipox, adeno-associated virus, modified Vaccinia Strain Ankara, Semliki Forest virus, poxvirus, and coronaviruses.

Fungal pathogens include, but are not limited to Cryptococcus ( . neoformans, C. gattii), P " neumocystis! (P. carinii, P.jiroveci), Candida (C. albicans), Aspergillus {A. fumigates, A. deflectus, A.flavus, A. flavipes), Ammonium, Penicillium, Paecilomyces, Coccidioides, Histoplasma capsulatum, Blastomyces dermatitidis, Microsporum (M. canis), Trichophyton (T. verrucosum, T. mentagrophytes), Epidermophyton or Malasse^ia (M. pachydermatis).

7. Allergic conditions

It will be appreciated that compositions of the present invention are in part directed to the prevention or treatment of allergic conditions. Allergic conditions include contact dermatitis, eczema, hives (urticaria), allergic conjunctivitis, hay fever, allergic rhinitis, airborne allergies including tree (e.g. birch pollen), weed (e.g. ragweed), and grass pollen allergies, latex allergies, food allergies (e.g. peanut, shellfish, milk protein), drug allergies (e.g. to penicillin), insect sting allergies (e.g. honeybee allergies, wasp allergies, hornet allergies, yellow jacket allergies, fire ant allergies), mold allergies (e.g. to alternaria, cladosporium, aspergillus, penicillium, helminthosporium, epicoccum, fusarium, mucor, rhizopus, and aureobasidium), dust mite allergies, animal allergies (e.g. household pets such as cats and dogs), allergic bronchopulmonary aspergillosis, occupational asthma, and episodic angioedema with eosinophilia.

Some of the allergens described herein, for example, drugs, food and insect stings, may result in anaphylaxis. While it is appreciated that the methods and compositions of the present invention may not be considered a primary treatment for a patient suffering from acute anaphylaxis, the methods and compositions may be used as a prophylactic treatment to modulate an allergic reaction in a patient susceptible to anaphylaxis. For example, the compositions may be co-administered with an allergen during an allergy de-sensitisation regime. In other examples, the compositions may be administered to a patient susceptible to anaphylaxis to prevent or reduce the likelihood of a severe allergic reaction to an allergen. Such prophylactic administration of the compositions of the present invention is specifically contemplated herein.

Another aspect of the invention provides a method of treating or preventing eosinophilia. In one embodiment, the eosinophilia is selected from airway, lung, blood and skin eosinophilia. In another embodiment, the eosinophilia is selected from eosinophilic ascites, eosinophilic cellulitis, eosinophilic fasciitis, eosinophilic gastroenteritis, coeliac disease, allergic colitis, eosinophilic esophagitis, eosinophilic pancreatitis, eosinophilic pneumonias, bronchiectasis, eosinophilic synovitis, nasal eosinophilia, tropical pulmonary eosinophilia, Churg Strauss syndrome, pulmonary eosinophilia, idiopathic hyper-eosinophilic syndrome, inflammatory bowel disease, eosinophilic cholangitis, eosinophilic leukaemia and other eosinophilic cancers, familial (hereditary eosinophilia), eosinophilic granuloma, sarcoidosis, eosinophilia-myalgia syndrome, cystic fibrosis, nasal polyposis, eosinophil meningitis, Wegener's granulomatosis, polyarteritis nodosa, rheumatoid arthritis, pemphigus vulgaris, bullous pemphigoid, dermatitis herpetiformis, erythema multiforme, eosinophilic cellulites, parasitic infections (Ascaris Toxocara canis, Filariasis, Anchylostomiasis, Trichinosis, Strong loidiasis, Fascioliasis, Schistosomiasis).

8. Compositions useful according to invention

A composition useful herein may be formulated as a food, functional food, drink, food additive, drink additive, dietary supplement, nutritional product, medical food, nutraceutical, medicament or pharmaceutical. Appropriate formulations may be prepared by an art-skilled worker with regard to that skill and the teaching of this specification.

In one embodiment the present invention relates to use of lactic acid whey or a derivative thereof, in the manufacture of a food, functional food, drink, food additive, drink additive, dietary supplement, nutritional product, medical food, nutraceutical, medicament or pharmaceutical.

Preferably the composition is formulated for oral administration. Preferably the composition is for increasing the production of Thl cytokines or suppressing the production of Th2 cytokines in the systemic circulation of a subject, or other uses, as described above.

. In one embodiment the composition is in the form of a tablet, a caplet, a pill, a hard or soft capsule or a lozenge.

In one embodiment the composition is in the form of a cachet, a dispensable powder, granules, a suspension, an elixir, a liquid, a drink, or any other form that can be added to food or drink, including for example water or fruit juice. In one embodiment the composition is an enteral product, a solid enteral product or a liquid enteral product. In one embodiment the composition further comprises one or more constituents (such as antioxidants) which prevent or reduce degradation of the composition during storage or after administration.

In one embodiment, compositions useful herein include any edible consumer product which is able to carry fats, fatty acids or lipid. Examples of suitable edible consumer products include baked goods, powders, Hquids, confectionary products, reconstituted fruit products, snack bars, food bars muesli bars, spreads, sauces, dips, dairy products including ice creams, yoghurts and cheeses, drinks including dairy and non-dairy based drinks (such as milk drinks including milk shakes, and yogurt drinks), milk powders, sports supplements including dairy and non-dairy based sports supplements, food additives such as protein sprinkles and dietary supplement products including daily supplement tablets. Within this embodiment, a composition useful herein may also be an infant formula, in powder or liquid form. Suitable nutraceutical compositions useful herein may be provided in similar forms.

In certain embodiments, the composition is a nutraceutical composition suitable for infants or the elderly. For example, the composition may be administered to an infant that has a Th2 biased immunity, to restore balance in their immunity. In other examples, the composition may be

administered to an elderly subject or an immuno-suppressed patient to upregulate their Thl cytokine production, their Th2 cytokine production, or both, for example so as to restore their Thl/Th2 balance.

In one embodiment, compositions useful herein include geriatric supplements, maternal formulas, infant formulas, follow-on formulas and growing up formulas. Such products are formulated to target nutrients to the elderly, foetus, infant and child.

In preferred embodiments, the compositions useful herein are able to maintain or restore balance of immunity in a subject to whom they are administered.

For example, such compositions are able to maintain or restore balance of immunity in a foetal subject (by 'indirect' administration to a foetal subject by administration to the pregnant mother).

The term "geriatric supplement" as used in this specification means a composition for elderly subjects. The term "maternal formula" as used in this specification means a composition for pregnant woman to take during pregnancy. The term "infant formula" as used in this specification means a composition for infants aged between 0 days and 6 months old. The term "follow-on formula" as used in this specification means a composition for infants aged 6 months to 1 year. The term "growing up formula" as used in this specification means a compositions directed to infants and children aged 1 year upwards. Growing-up formula includes growing-up milk powders as will be understood by those skilled in the art.

In another embodiment, compositions useful herein include dietetic products. The term "dietetic product" means a product specially processed or formulated to satisfy particular dietary requirements which exist because of a particular physical or physiological condition and/ or specific diseases and disorders and which are presented as such.

For example, in subjects suffering from or susceptible to autoimmune conditions, compositions of the present invention may be administered to modulate the immune response. For example, to downregulate production of Thl cytokines or upregulate production of Th2 cytokines.

In general, for oral administration a dietary (a food, food additive or food supplement for example), nutraceutical or pharmaceutical composition useful herein may be formulated by a skilled worker according to known formulation techniques.

Thus, a pharmaceutical composition useful according to the invention may be formulated with an appropriate pharmaceutically acceptable carrier (including excipients, diluents, auxiliaries, and combinations thereof) selected with regard to the intended route of administration and standard pharmaceutical practice. See for example, Remington's Pharmaceutical Sciences, 16th edition, Osol, A. Ed., Mack Publishing Co., 1980.

A dosage form useful herein may be administered orally as a powder, liquid, tablet or capsule. Suitable dosage forms may contain additional agents as required, including emulsifying, antioxidant, flavouring or colouring agents, or have an enteric coating. Suitable enteric coatings are known. Enteric coatings surrounding the active ingredients and prevent the release of the active ingredients in the stomach but allow release after the dosage form has left the stomach. Dosage forms useful herein may be adapted for immediate, delayed, modified, sustained, pulsed or controlled release of the active components. Suitable formulations may contain additional agents as required, including emulsifying, antioxidant, flavouring or colouring agents.

Thus, the invention also is directed to doses, dosage forms, formulations, compositions and/or devices comprising lactic acid whey or a derivative thereof of the invention including those disclosed herein, useful for the therapy of diseases, disorders, and/or conditions in humans and other mammals and other disorders as disclosed herein. The use of these dosage forms, formulations, compositions and/or devices comprising one or more compositions of the invention enables effective treatment of these conditions. The invention provides, for example, dosage forms, formulations, devices and/ or compositions containing lactic acid whey or a derivative thereof of the invention. The dosage forms, formulations, devices and/ or compositions of the invention may be formulated to optimize bioavailability, immunogenicity, or to maintain plasma, blood, or tissue concentrations within the immunogenic or therapeutic range, including for extended periods. Controlled delivery preparations may also be used to optimize the antigen concentration at the site of action, for example.

The dosage forms, formulations, devices and/ or compositions of the invention may be formulated for periodic administration, for example to provide continued exposure to the one or more 00228 compositions of the invention. Strategies to elicit a beneficial immunological response, for example those that employ one or more "booster" vaccinations, are well known in the art, and such strategies may be adopted in the practise of the present invention.

Pharmaceutical compositions and dosage forms can be administered via the parenteral route, and this route will be preferred for certain embodiments of methods of eliciting an immune response, such as those described herein. Examples of parenteral dosage forms include aqueous solutions, isotonic saline or 5% glucose of the active agent, or other well-known pharmaceutically acceptable excipients. Cyclodextrins, for example, or other solubilising agents well-known to those familiar with the art, can be utilized as pharmaceutical excipients for delivery of the therapeutic agent.

Examples of dosage forms suitable for oral administration include, but are not limited to tablets, capsules, lozenges, or like forms, or any liquid forms such as syrups, aqueous solutions, emulsions and the like, capable of providing a therapeutically effective amount of a composition of the invention.

Capsules can contain any standard pharmaceutically acceptable materials such as gelatin or cellulose. Tablets can be formulated in accordance with conventional procedures by compressing mixtures of the active ingredients with a solid carrier and a lubricant. Examples of solid carriers include starch and sugar bentonite. Active ingredients can also be administered in a form of a hard shell tablet or a capsule containing a binder, e.g., lactose or mannitol, a conventional filler, and a tabletting agent.

Examples of dosage forms suitable for transmucosal delivery of the compositions and formulations of the invention include depositories solutions for enemas, pessaries, tampons, creams, gels, pastes, foams, nebulised solutions, powders and similar formulations containing in addition to the active ingredients such carriers as are known in the art to be appropriate. Specifically contemplated are dosage forms suitable for inhalation or insufflation of the compositions and formulations of the invention, including compositions comprising solutions and/ or suspensions in pharmaceutically acceptable, aqueous, or organic solvents, or mixture thereof and/or powders. Transmucosal administration of the compositions and formulations of the invention may utilize any mucosal membrane but commonly utilizes the nasal, buccal, vaginal and rectal tissues. Formulations suitable for nasal administration of the compositions and formulations of the invention may be administered in a liquid form, for example, nasal spray, nasal drops, or by aerosol adrninistration by nebulizer, including aqueous or oily solutions of the lactic acid whey or derivative thereof. Formulations for nasal adrninistration, wherein the carrier is a solid, include a coarse powder having a particle size, for example, of less than about 100 microns, preferably less, most preferably less than about 50 microns, which is administered in the manner in which snuff is taken, i.e., by rapid inhalation through the nasal passage from a container of the powder held close up to the nose. Formulations of the invention may be prepared as aqueous solutions for example in saline, solutions employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bio-availability, fluorocarbons, and/ or other solubilising or dispersing agents known in the art.

Examples of dosage forms suitable for buccal administration of the compositions and

formulations of the invention include lozenges, tablets and the like, compositions comprising solutions and/ or suspensions in pharmaceutically acceptable, aqueous, or organic solvents, or mixtures thereof and/ or powders.

Examples of formulations of compositions, including vaccines and controlled drug

formulations, useful for delivery of the compositions and formulations of the invention are found in, for example, Sweetman, S. C. (Ed.). Martindale. The Complete Drug Reference, 33rd Edition, Pharmaceutical Press, Chicago, 2002, 2483 pp.; Aulton, M. E. (Ed.) Pharmaceutics. The Science of Dosage Form Design. Churchill Livingstone, Edinburgh, 2000, 734 pp.; and, Ansel, H. C, Allen, L. V. and Popovich, N. G. Pharmaceutical Dosage Forms and Drug Delivery Systems, 7th Ed.,

Lippincott 1999, 676 pp.. Excipients employed in the manufacture of drug delivery systems are described in various publications known to those skilled in the art including, for example, Kibbe, E. H. Handbook of Pharmaceutical Excipients, 3rd Ed., American Pharmaceutical Association, Washington, 2000, 665 pp. The USP also provides examples of modified-release oral dosage forms, including those formulated as tablets or capsules. See, for example, The United States Pharmacopeia 23/National Formulary 18, The United States Pharmacopeial Convention, Inc., Rockville MD, 1995 (hereinafter "the USP"), which also describes specific tests to determine the drug release capabilities of extended- release and delayed-release tablets and capsules. The USP test for drug release for extended-release and delayed-release articles is based on drug dissolution from the dosage unit against elapsed test time. Descriptions of various test apparatus and procedures may be found in the USP. Further guidance concerning the analysis of extended release dosage forms has been provided by the F.D.A. (See Guidance for Industry. Extended release oral dosage forms: development, evaluation, and application of in vitro/in vivo correlations. Rockville, MD: Center for Drug Evaluation and Research, Food and Drug Administration, 1997).

Further examples of dosage forms of the invention include, but are not limited to modified- release (MR) dosage forms including delayed-release (DR) forms; prolonged-action (PA) forms;

controlled-release (CR) forms; extended-release (ER) forms; timed-release (TR) forms; and long-acting (LA) forms. For the most part, these terms are used to describe orally administered dosage forms, however these terms may be applicable to any of the dosage forms, formulations, compositions and/or devices described herein. These formulations effect delayed total drug release for some time after drug administration, and/ or drug release in small aliquots intermittendy after administration, and/ or drug release slowly at a controlled rate governed by the delivery system, and/ or drug release at a constant rate that does not vary, and/or drug release for a significantly longer period than usual formulations. W

The compositions useful herein may be used alone or in combination with one or more other therapeutic agents. The therapeutic agent may be a food, drink, food additive, drink additive, food component, drink component, dietary supplement, nutritional product, medical food, nutraceutical, medicament or pharmaceutical.

In one embodiment the lactic acid whey or a derivative thereof, optionally with at least one additional therapeutic agent are formulated for administration separately, simultaneously or sequentially.

In one embodiment the lactic acid whey or a derivative thereof, optionally with at least one additional therapeutic agent are formulated for co-administration.

In one embodiment the lactic acid whey or a derivative thereof, optionally with at least one additional therapeutic agent are formulated for sequential administration.

When used in combination with another therapeutic agent, the administration of a

composition useful herein and the other therapeutic agent may be simultaneous or sequential.

Simultaneous administration includes the administration of a single dosage form that comprises all components or the administration of separate dosage forms at substantially the same time. Sequential administration includes administration according to different schedules, preferably so that there is an overlap in the periods during which the composition useful herein and other therapeutic agent are provided.

Suitable agents with which the compositions of the invention can be co-administered include immunotherapeutic agents, and other suitable agents known in the art.

Additionally, it is contemplated that a composition in accordance with the invention may be formulated with additional active ingredients which may be of benefit to a subject in particular instances. For example, therapeutic agents that target the same or different facets of the disease process may be used.

It should be understood that the additional therapeutic agents listed above (both food based and pharmaceutical agents) may also be employed in a method according to the invention where they are administered separately, simultaneously or sequentially with a composition useful herein.

The efficacy of a composition useful according to the invention can be evaluated both in vitro and in vivo. Briefly, in one embodiment the composition can be tested for its ability, to for example, normalise or restore an immune response in vitro. For in vivo studies, the composition can be fed to or administered to an animal (e.g., a mouse) and its effects on modulating an immune response is then assessed. Based on the results, an appropriate dosage range and administration route can be

determined.

The amount of lactic acid whey or a derivative thereof to be aclministered is a therapeutically effective amount, which means an amount that is enough to elicit a desirable effect. A desirable effect is modulation of the immune response in a subject in need thereof. As will be appreciated, the dose of the composition administered, the period of administration, and the general administration regime may differ between subjects depending on such variables as the severity of symptoms of a subject, the type of disorder to be treated, the mode of administration chosen, and the age, sex and/ or general health of a subject and the extent to which the lactic whey product has been concentrated through removal of inert components such as water and lactose. However, by way of general example, the inventors contemplate administration of from about 50 mg to about 2000 mg per kg body weight of a composition useful herein is administered per day, preferably about 100 to about 500 mg per kg per day.

It should be appreciated that aclministration may include a single daily dose or administration of a number of discrete divided doses as may be appropriate. It should be understood that a person of ordinary skill in the art will be able without undue experimentation, having regard to that skill and this disclosure, to determine an effective dosage regime (including daily dose and timing of administration) for a given condition.

The present invention also relates to a dietary, nutraceutical or oral pharmaceutical composition comprising, consisting essentially of or consisting of lactic acid whey or a derivative thereof, optionally in combination with a therapeutically active agent.

In one embodiment a composition of the invention is a lactic acid whey fraction. In one embodiment the lactic acid whey fraction comprises at least about 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 or 99% by weight lactic acid whey or a derivative thereof, and useful ranges may be selected from any of these values (for example, from about 1 to about 99% by weight, from about 5 to about 99% by weight, from about 10 to about 99% by weight, from about 15 to about 99% by weight, from about 20 to about 99% by weight, from about 25 to about 99% by weight, from about 30 to about 99% by weight, from about 35 to about 99% by weight, from about 40 to about 99% by weight, from about 45 to about 99% by weight, from about 50 to about 99% by weight, from about 55 to about 99% by weight, from about 60 to about 99% by weight, from about 65 to about 99% by weight, from about 70 to about 99% by weight, from about 75 to about 99% by weight, from about 80 to about 99% by weight, from about 85 to about 99% by weight, from about 90 to about 99% by weight, or from about 95 to about 99% by weight).

In one embodiment the composition comprises, consists essentially of or consists of about 0.1, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 or 99% of fresh, recombined or powdered lactic acid whey or a derivative thereof and useful ranges may be selected between any of these foregoing values (for example, from about 1 to about 99%, from about 5 to about 99%, from about 10 to about 99%, from about 15 to about 99%, from about 20 to about 99%, from about 25 to about 99%, from about 30 to about 99%,. from about 35 to about 99%, from about 40 to about 99%, from about 45 to about 99%, from about 50 to about 99%, from about 55 to about 99%, from about 60 to about 99%, from about 65 to about 99%, from about 70 to about 99%, from about 75 to about 99%, from about 80 to about 99%, from about 85 to about 99%, from about 90 to about 99%, or from about 95 to about 99% ).

In one embodiment the daily dosage range (by any route) is about 10 mg to 2000 mg of lactic acid whey or a derivative thereof solids per day per kg of body weight, preferably 10 mg to 1.5 g/kg/day, preferably 10 mg to 500 mg/kg/day. A higher dose is preferred for short-term treatment and prevention and a lower dose for long-term treatment and prevention.

Various aspects of the invention will now be illustrated in non-limiting ways by reference to the following examples.

EXAMPLES

Example 1 - The effect of lactic acid whey in increasing cytokine and antibody production in mice.

This example describes the effect of various lactic acid whey compositions in increasing the production of cytokines and IgG antibody in mice.

Material and Methods

All animal experiments are approved by the University of Otago Animal Ethics Committee 1. Feeding of mice

Female C57BL/ 6 mice of different ages were fed daily fractions (5g/kg) for a total of 8 days. The six treatment groups (n= >20 per group) were as follows:

A. Animals fed 10 uL Sample A Batch 1 (0.33mg/uL in PBS)

B. Animals fed 10 uL Sample B (0.33mg/uL in PBS)

C. Animals fed 10 uL Sample C (0.33mg/uL in PBS)

D. Animals fed 10 uL Sample D (0.33mg/uL in PBS)

E. Animals fed 10 uL Sample E (0.33mg/uL in PBS)

Ctrl. PBS control (Ctrl)

Details of each sample are provided in Table 1 below: ; .

Table 1: Milk protein compositions fed to mice

Sample Active Ingredient

Sample A Commercial lactic whey protein concentrate

Sample B β-casein peptic hydrolysate

Sample C Peptic hydrolysate of a commercial whey protein isolate

Sample D Glycomacropeptide tryptic hydrolysate Sample E Whey protein derived paediatric hydrolysate

Sample F Mineral acid whey protein concentrate

Samples B and D were included as casein and casein hydrolysates have been reported as having certain immunomodulatory effects. Sample F was included as a representative example of a commonly used acid whey product.

Immunological assay

The spleens were excised aseptically from each mouse and placed in 5mL of pre-warmed DMEM medium. A single cell suspension was made by forcing the cells through a 70μιη pore nylon strainer (Falcon, Becton Dickinson) in DMEM. The spleen cells were suspended in lOmL of DMEM and centrifuged at 250 x g for 5 minutes. The supernatant was discarded and the pellet resuspended.

DMEM medium (5-lOmL) containing Foetal Calf Serum was added to each tube and centrifuged at 250 x g for 5 minutes. The cell pellets were resuspended in lOmL of DMEM, and the cell number for each counted and adjusted to 2 x 10 6 /mL. The splenocytes were stimulated with 1.25 μg /mL and 5 μg /mL ConA, 5 fxg /mL and 10 μg /mL Pokeweed mitogen or 1.25 μg /mL and 2.5 μg /mL lippolysaccharide to encourage cell proliferation, or were left untreated. Cytokine profiles were obtained from each treatment group using Bioplex plates (BioRad) according to the manufacturer's instructions.

Results

Sample A-fed mice showed a significant increase in cytokines IFN-γ, TNF-oc and IL-4 compared to the PBS control group, as shown in Table 2 below.

No significant difference in cytokine profiles was observed in Sample D- and Sample E-fed mice compared to the PBS control group.

Table 2. Effect of treatment on cytokine production

Treatment Group

Cytokine

Group mean (SEM)

(stimulant)

Control A B C D E

IL-4 15.7 217.46 311.6 229.19 15.16 188.61

Cell (1.61) (81.15) (74.48) (55.34) (8.27) (60.38)

IL-4 257.5 338.27 346.82 138.56 52.92 244.72

ConA (49.61) (88.54) (113.03) (36.33) (12.68) (58.04) '

IL-4 632.33 389.15 344.62 293.72 59.49 217.01

PWM (79.78) (119.25) (95.070 (88.53) (13.38) (45.25)

IL-4 16.36 232.66 318.24 255.01 72.84 297.96

LPS (1.83) (49.89) (77.66) (63.5) (14.65) (59.11)

IFN-Y . 14.6 2678.92 871.84 452.19 9.34 238.27

Cell (4.74) (446.27) (169.8) (126.2) (5.3) (60.16)

IFN-γ 62.57 2160.7 902.85 226.21 31.3 306.36

ConA (24.38) (395.18) (258.57) (61.930 (8.53) (80.8)

IFN-γ 127.8 3385.16 589.41 307.48 29.21 217.36

PWN (25.93) (662.03) (179.99) (97.33) (8.27) (47.89)

IFN-γ 29.53 1874.14 663.46 670.53 26.56 253.58

LPS (9.73) (497.27) (217.11) (238.27) (7.26) (57.83)

TNF-a 6.8 100.1 123.38 101.3 21.99 18.32

CeU (2.33) (14.96) (25.49) (14.27) (2.34) (10.31)

TNF-a 9.13 114.55 96.85 74.06 25.82 . 24.57

ConA (3.86) (17.07) (27.02) ' (9.91) (1.79) (9.02)

TNF-a 51.26 83.52 . 47.27 65.25 22.2 7.84

PWM (9.16) (13.35) (18.52) (13.97) (1.37) (5.44)

TNF-a 13.92 54.52 64.57 68.84 21.2 10.86

LPS (4.66) (9.52) (22.02) (11.77) (1.18) (5.44)

PWM = Pokeweed mitogen, LPS = lippolysaccharide, cell = no stimulant control

Example 2 - Efficacy of lactic acid whey fed to influenza challenged mice.

This example describes an assessment of the efficacy of various milk fraction compositions in eliciting an immune response in influenza challenged mice. Matedals and Methods

All animal experiments were approved by the University of Otago Animal Ethics Committee.

1. Immunisation of mice

Female Balb/c mice (Hercus Taieri Research Unit, New Zealand) aged 6 weeks were split into two groups— Group A. sub-cutaneously immunised (3 x 10 6 PFU in lOOuL) with influenza vaccine (heat killed influenza virus A/Puerto Rico/8/34 [HlNl] ATCC # VR-95 GenBank Accession # J02143), or Group B. non-vaccinated mice.

The vaccinated groups were immunised two weeks prior to challenge with influenza. Feeding of products, in addition to normal rodent chow (that was dairy free), to animals also started at this time.

Mice were fed fractions daily (as detailed in Table 1), by delivery into the oral cavity, in addition to dairy-free rodent chow for three weeks prior to challenge. The treatment groups (n=8 per group) were as follows:

a) Vaccinated control.

b) Vaccinated animals fed Sample A.

c) Non-vaccinated animals fed Sample A

d) Vaccinated animals fed Sample F.

e) Non- vaccinated animals fed Sample F.

f) Non-vaccinated animals fed PBS for 21 days followed by Sample A from day 1 of challenge.

g) Non-vaccinated PBS control.

All groups apart from group F were also fed from the day the other groups were vaccinated. Group F was not vaccinated and didn't begin feeding with Sample A until the day of challenge with influenza.

Other animals were fed the fractions at 5g/kg as described above either in conjunction with vaccination or alone.

2. Pathogenic challenge

21 days after the start of the feeding regime, all mice were challenged with influenza virus. To infect the mice, 15uL of virus per nostril (equivalent to a viral dose of 8 x 10 5 PFU/mouse) was administered.

3. Behavioural and physical observation

Food consumption, weight, behavioural changes and changes in general and physical appearance of each mouse was monitored and recorded daily.

4. Immunological assay The mice were anaesthetised intraperitoneally six days after the pathogenic challenge and sacrificed by cardiac puncture and cervical dislocation. Blood was collected in serum separating tubes and allowed to clot. After 30 minutes, the tubes were centrifuged at 8000 rpm for 5 minutes, and the serum frozen at -80°C until assayed. The spleen and lungs from each mouse were removed.

5. Influenza Titres from Lungs

Both lungs were removed from each animal at the time of sacrifice, suspended in 4ml of DMEM and homogenised for 20 seconds at room temperature using a T25 Basic homogeniser, (IKA

Lab technologies). The lung homogenate was centrifuged at 250g for 7 minutes to remove tissue debris and the supernatants were serially diluted 10-fold in DMEM.

25ul of the dilutions (six in total) were plated in duplicate into a 96-well plate containing confluent Madin Darby Canine Kidney (MDCK ATCC # CCL-34) cells and incubated for lh at 37°C. MDCK cells were grown in DMEM supplemented with 10% FCS and penicillin streptomycin in a humidified incubator at 37°C with 5% C0 2 .

of the DMEM medium supplemented with 2μg/ml trypsin (Difco # 215240) was added and the plates incubated for 5-6 days. At this time the virus only controls were exhibiting CPE.

The media was aspirated from each well and the cells washed with 200μ1 of PBS and the washings discarded. The remaining cells were fixed for 1 minute with ΙΟΟμΙ of methanol and then ΙΟΟμΙ of 1% crystal violet (BDH # 351884W) solution (in distilled water) was added to each well and allowed to react with the cells at RT for 5 minutes. The wells were washed and then the stain was rehydrated with 33% acetic acid.

The absorbance of each well was then measured at 595nm in a Biorad plate reader.

Titres of influenza were calculated , by the Reed-Meunch 50% endpoint titre method using the following equation:

(% infected cells above 50%) - 50 -

(% infected cells above 50%) - (% infected cells below 50%)

Results

Influenza virus was either not cultured or cultured in very low numbers from the lungs of vaccinated animals as expected. Non-vaccinated animals fed Sample A either pre or post challenge resulted in a reduction in the numbers of influenza virus able to be cultured compared to non- vaccinated animals fed PBS, see Figure 1. Sample F, a mineral acid whey protein isolate, had no effect on influenza virus numbers in the disease model. In summary, fewer influenza particles were recovered from the lungs of animals fed Sample A than control animals or those animals fed Sample F.

Discussion The results from this influenza study indicate that there is a protective effect against influenza infection mediated by Sample A when fed either pre- or post-challenge. This protective effect was not observed by feeding Sample F, which had no effect on influenza virus numbers in the disease model.

Example 3 - Efficacy of lactic acid whey fed to OVA-sensitized mice.

This example describes the efficacy of various milk fraction compositions in restoring a balance between the Thl and Th2 immune response in OVA-sensitized mice.

Materials and Methods

All animal experiments are approved by the University of Otago Animal Ethics Committee.

1. Sensitization of mice

Female Balb/c mice (Hercus Taieri Research Unit, New Zealand) aged 6 weeks are injected peritoneally with 100 μg/mL ovalbumin adsorbed to alum.

Mice are fed daily fractions (5g/kg) in addition to dairy- free rodent chow for 21 days. The six treatment groups (n=6 per group) are as follows:

a) OVA-sensitized control

b) PBS control

c) OVA sensitized animals fed Sample A Batch 1

d) OVA sensitized animals fed Sample A Batch 2

e) " OVA sensitized animals fed Sample F

f) OVA sensitized animals fed Sample C

2. Immunological assay

Feed is removed from the mice 4 hours prior to euthanasia. Two hours prior to euthanasia the mice are challenged orally with a 100 uL (40mg/ml) dose of ovalbumin. Blood is collected and the serum is analysed for OVA specific IgGl and IgG2a, IgE, mouse mast cell protease and cytokine IL-5.

Splenocytes are cultured with OVA and treated as with fractions or mitogens as described herein. Splenocyte proliferation is measured at 72 hours and cytokines are analysed at 48 hours.

Results

Results establishing that OVA-sensitized mice fed with Sample A show a reduction in the immune parameters associated with sensitisation compared to OVA-sensitized mice are supportive of the immulogical efficacy of lactic acid whey compositions as contemplated herein. Lower IgGl, IgG2a, mast cell protease and IL- 5 levels observed in Sample A-fed mice will be indicative of an immunomodulatory effect.

Example 4 - Peritoneal macrophage assay This example describes the efficacy of various milk fraction compositions in eliciting a cellular immune response in mice.

Method

Five day old C57Black 6 mice were fed Sample A (see Table 1) using the following protocol: Group 1 40 five day old mice fed PBS for seven days.

Group 2 40 five day old mice fed Sample A for seven days - 3.3mg/per pup/day

At end of seven day feeding 20 mice from each group were sacrificed and the resident macrophage numbers in the peritoneal cavity able to take up E.coti ' were measured (Figure 2). The remaining 20 mice in each group were left for a further seven days without being fed the product before being sacrificed and the resident macrophage numbers in the peritoneal cavity able to take up E.coli were measured (Figure 3).

Results

Macrophages collected from the peritoneum of mice fed Sample A were able to phagocytose more bacteria (26%) than those from PBS fed mice (17%) as shown in Figure 2. This difference was significant (p = 0.0021).

After the seven-day washout period macrophages from mice fed PBS were able to phagocytose the same number of bacteria as the mice fed Sample A (Figure 3).

Following the washout period for mice fed Sample A there was no change in the ability of peritoneal macrophages to phagocytose the bacteria (26%). However, the PBS mice were now able to phaocytose the bacteria at the same rate as the Sample A fed mice.

Example 5 - LTA assay

This example describes the analysis of various milk fraction compositions for the presence of Lipoteichoic acid (LTA), a major constituent of the cell wall of Gram-positive bacteria.

Method

Mice aged 7, 15, 21, and 35 days were sacrificed and animal weights were recorded prior to spleens being collected and weighed. Splenocytes were isolated using the Total Murine Splenocyte Preparation Method and plated at a concentration of 2 x 10 6 cells /ml in 96 well tissue culture plates (Falcon MicroTest®) prior to being stimulated with the mitogen ConcanavaUn A (T cell compartment - Sigma ensiformis Jack Bean Type IV) at 1.25 ug/ml, pokeweed mitogen (B cell compartment - Sigma Lectin from Phytolacca americana) at 5ug/ml and lipopolysaccharide (monocytes - E.coli 055:B5 Sigma) at a concentration of 0.625ug/ml and lipoteichoic acid (Sigma L215) at concentrations from 40 to 0.3125ug/mL Plates were incubated for 72 hours in a 5% C0 2 incubator. Then, Thymidine [6-3 FTJ - ( O.OCi/mmol (370 GBq/mmol) Amersham) was added to the wells and the plate incubated for a further 16 hours.

Plates were harvested onto filter membranes - glass fibre printed Filtermat A for 1450

MicroBetaTM (Perkin Elmer) using a Tomtec cell harvester. Filtermats were placed into Sample bags for Microbeta TM (Perkin Elmer) and 4 mis of Betaplate Scint fluid (Perkin Elmer) was added prior to plates being read in a Wallac 1450 MicroBeta Plus Liquid scintillation counter.

Results

Splenocytes were unresponsive to LTA even when used at concentrations of 40 g/ml (Figure 4). The response to LTA was not much above baseline, in comparison to that with mitogenic stimulation which would be expected to be an order of magnitude greater (in the order of lOOO's cpm rather than 100's cpm).

Discussion

The data presented in this example indicate that any immunological response observed on

administration of the samples tested herein is unlikely to be due to the presence of LTA.

Example 6— LPS analysis

This example describes the analysis of various milk fraction compositions of the invention for endotoxins (lipopolysaccharides, (LPS)).

Method

The samples (see Table 1) were tested by Environmental Science & Research, NZ, using the limulus amebocyte lysate assay.

Results

Endotoxin units for the various samples were as follows:

Sample A 4 EU/mg

Sample B 2 EU/mg

Sample C 4 EU/mg

Sample D 32 EU/mg

Sample E 8 EU/mg

Discussion

lOOpg of the standard endotoxin EC-5 has an activity of 1EU in this assay. Therefore, it is possible to extrapolate to estimate that Sample A has approximately 400pg of endotoxin per mg. The maximum level of endotoxin for intravenous applications of pharmaceutical and biologic product is set to 5 endotoxin units (EU) per kg of body weight per hour by all pharmacopoeias (J Pharm Pharmaceut Sci (www.cspscanada.org) 10(3):388-404, 2007, Methods of Endotoxin Removal from Biological Preparations: a Review, Perola O. Magalhaesl†, Andre M. Lopes, Priscila G. Mazzola3, Carlota Rangel- Yagui2, Thereza C. V. Penna3, Adalberto Pessoa Jr.3 ). Thus, the level of endotoxin present in Sample A at the doses administered herein is not sufficient to elicit the immune responses observed.

INDUSTRIAL APPLICATION

The present invention has utility in promoting a beneficial immune response in a subject in need thereof, including modulating a Thl immune response, or a Th2 response, including for example promoting suppression of a deleterious Th2 response in a subject. The modulation of a Thl or Th2 immune response is useful in the treatment of diseases or conditions, including those associated with or caused by pathogens or allergens, including those associated with aberrant elevation of a Th2 response.