Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Peptide containing doai
Reexamination Certificate
1999-01-27
2004-05-18
Nolan, Patrick J. (Department: 1644)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Peptide containing doai
C530S326000, C530S327000, C514S014800
Reexamination Certificate
active
06737406
ABSTRACT:
The present invention relates to immunologically cryptic peptides; methods for their identification in individuals and populations and their use in diagnosis and therapy of pathological conditions such as asthma and allergy, and their use in screening for therapeutic activity.
The ability of the immune system to elicit a response to a particular molecule depends critically upon its ability to recognise the presence of an antigen. Classically, the term antigen was associated with the ability of a molecule to be an antibody generator via induction of B-cells. It is now known however that T cells also possess the ability to recognise antigens. T-cell antigen recognition requires antigen presenting cells (APCs) to present antigen fragments (peptides) on their cell surface in association with molecules of the major histocompatibility complex (MHC). T cells use their antigen specific receptors (TCRs) to recognise the antigen fragments presented by the APC. Such recognition acts as a trigger to the immune system to generate a range of responses to eradicate the antigen which has been recognised.
In order to trigger an individual T cell, a critical number of TCRs must be ligated by the peptide/MHC complex presented by the APC. A peptide which reaches the surface of the APC in sufficient numbers to do this can be termed “dominant” or “sub-dominant” depending upon its ability to induce T cell activation relative to other triggering peptides. In the normal course of events i.e. physiologically, a given protein will generate more than one peptide which is capable of triggering a T cell response. The term “dominant” would then be applied to the peptide that induces the most potent or most frequent response. In addition to dominant and sub-dominant epitopes, there are potential T cell peptide epitopes within a given protein sequence (for which T cells are specific), which do not reach the APC surface in sufficient numbers to induce a response. In other words due to the mechanisms of antigen processing within the APC, certain peptides are processed and presented to T-lymphocytes efficiently and therefore stimulate T-cell responses white others are poorly processed and presented to T-lymphocytes. As these latter peptides are not present upon the APC surface in sufficient numbers to stimulate a potentially reactive T-lymphocyte, these peptides have been referred to as “cryptic peptide epitopes”.
Cryptic peptide epitopes are present both in proteins normally present in the body (self proteins) and in non-self (or foreign) proteins. In normal physiology, T cells which have the capability of reacting with a cryptic epitope cannot be detected in an in vitro primary stimulation assay (that is, T cells freshly isolated from the blood do not exhibit demonstrable proliferation when cultured with the cryptic peptide). In contrast other peptides which are efficiently processed and presented to T-lymphocytes by APC. can stimulate a proliferative response in primary culture. These are the “dominant” and “sub-dominant” epitopes.
The term “atopic allergy” is applied to a group of allergies characterised by high concentrations of immunoglobulin E (IgE). They include allergic asthma, hay fever, perennial allergic rhinitis, some forms of urticaria (hives) and eczema, allergic conjunctivitis and certain food allergies (particularly food anaphylaxis). The mechanisms of generation of the pathology of such atopic conditions involves not only the synthesis of antigen/allergen specific IgE but also the accompanying differentiation and growth of effector cells such as mast cells and eosinophils.
Allergic IgE-mediated diseases are currently treated by desensitization procedures that involve the periodic injection of allergen components or extracts. Desensitization treatments may induce an IgG response that competes with IgE for allergen, or they may induce specific suppressor T cells that block the synthesis of IgE directed against allergen. This form of treatment is not always effective and poses the risk of provoking serious side effects, particularly general anaphiylactic shock. This can be fatal unless recognised immediately and treated with adrenaline. A therapeutic treatment that would decrease or eliminate the unwanted allergic-immune response to a particular allergen, without altering the immune reactivity to other foreign antigens or triggering an allergic response itself would be of great benefit to allergic individuals.
Sometimes the normal mechanisms whereby self and non-self are immunologically distinguished may break down and an immune response may be elicited against self-antigens present in normal body tissues. This “auto-immunity” generates pathological conditions such as autoimmune thyroiditis, rheumatoid arthritis and lupus erythematosus. Therapeutic regimes are generally limited to the use of anti-inflammatory or immunosuppressive drugs which are relatively non-specific and have many undesirable side-effects.
WO 92111859 describes a method of reducing immune response to an allergen in which a non-allergen derived, non-stimulating peptide which binds to specific MHC class 11 molecules of APCs is used to inhibit T-cell response to particular allergens.
WO 91/06571 purports to disclose peptides derived from human T-cell reactive feline protein which can be used in the diagnosis, treatment or prevention of cat allergy.
WO 94/24281 relates to peptides and modified peptides of the major house dust mite allergens. The modified peptides have the intent of reducing the level of undesirable side effects associated with desensitizing therapies.
G. F. Hoyne et. al. in Immunology 83 pp 190-195 (1994) examined house dust mite allergy using peptides made from cDNA encoding the major allergen DerpI. They purport to show that peptides containing major epitopes can induce oral tolerance in mice to the whole allergen and that it is also possible to induce tolerance with other peptides. Cryptic peptides are suggested as playing a role in this process but no methods are disclosed for their identification or therapeutic use.
None of the above disclosures makes any suggestion that cryptic peptides may play a role in the pathology of atopic conditions such as asthma or other allergic diseases. The present inventors have found a method for identifying cryptic peptides and have observed that individuals with asthma or other allergy-based pathologies have T-lymphocyte populations which can be stimulated in primary culture by cryptic epitopes derived from the allergen which causes the relevant pathology. As described above, T-lymphocytes isolated from a healthy individual would not be expected to be stimulated in primary culture by a cryptic epitope. Likewise in autoimmune pathologies, a self peptide, normally cryptic to the immune system becomes recognised and elicits an immune response.
Hence there is provided according to the invention, a method of determining whether a peptide of a protein is a cryptic peptide, which method includes the steps of: i) exposing T cells with the peptide in a primary challenge; ii) measuring the reactivity of T cells with the peptide in the primary challenge of Step i; iii) exposing pre-challenged T cells with the peptide in a secondary challenge, wherein the pre-challenged T cells are obtainable by exposing the T cells to the protein; and measuring the reactivity of the pre-challenged T cells with the peptide in the secondary challenge.
The prechallenge allows expression of not only dominant and sub-dominant epitopes on the APC surface, but also of any cryptic determinants. The subsequent peptide rechallenge of these cells reveals T-cell reactivity to the dominant, sub-dominant and cryptic epitopes. Primary challenge with peptides will elicit responses from only the normally expressed dominant and sub-dominant epitopes. Thus, peptides recognised following whole antigen primary challenge followed by peptide secondary challenge, but not after peptide primary challenge alone are by definition, cryptic epitopes. The peptide is a cryptic peptide if T-cell reactivity is observable in the secondary
Kay Anthony Barrington
Larche Mark
Circassia, Ltd.
Nolan Patrick J.
Palmer & Dodge LLP
Williams Kathleen M.
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