CZ20013081A3 - Epitopes and a extratopes obtained from C-epsilon-2 region or C-epsilon-4 IgE region, their antagonists a their therapeutic use - Google Patents

Abstract

The present invention relates to the provision of novel medicaments for the treatment, prevention or amelioration of allergic disease. In particular, the novel medicaments are epitopes or mimotopes derived from the C epsilon 3 or C epsilon 4 domains of IgE. These novel regions may be the target for both passive and active immunoprophylaxis or immunotherapy. The invention further relates to methods for production of the medicaments, pharmaceutical compositions containing them and their use in medicine. Also forming an aspect of the present invention are ligands, especially monoclonal antibodies, which are capable of binding the IgE regions of the present invention, and their use in medicine as passive immunotherapy or immunoprophylaxis.

Description

Epitopes and mimotopes derived from the C-epsilon-2 or Cepsilon-4 IgE region, their antagonists and their therapeutic use

Technical field

The invention relates to providing a novel medicament for the treatment, prevention or amelioration of an allergic disease. In particular, the novel drugs are epitopes or mimotopes derived from the Ce3 and Ce4 IgE regions. These new regions may be targets for passive and active immunoprophylaxis or immunotherapy. The invention further provides methods of producing medicaments, pharmaceutical compositions containing them and their use in medicine. The invention further provides ligands, particularly monoclonal antibodies, that are capable of binding to the IgE regions of the invention and their use in medicine as passive immunotherapy or immunoprophylaxis.

BACKGROUND OF THE INVENTION

With allergy histamine, an allergic response, symptoms commonly associated with the release of allergic mediators such as cells into surrounding tissues and vascular from immune structures.

Histamine is in cells and basophils, normally stored in masts when released for up to a time, due to interaction with allergen specific IgE. The role of IgE in mediating allergic reactions such as asthma, food allergy, atopic dermatitis, allergic rhinitis is well known.

as

B-cells, when pollen or mite allergens, react with allergen-specific IgE synthesis. The allergen specific IgE then binds to the FcεRI receptor (high affinity IgE receptor) found on basophils and mast cells. Any subsequent encounter with the allergen results in the release of histamine from mast cells or basophils by chaining

surrounding IgE / FcεRI complexes (Sutton and

Gould, Nature, 1993, 366: 421-428, EP 0 477 231 B1).

Like all immunoglobulins, IgE contains two heavy and two light chains. The heavy chain ε contains five regions: one variable region (VH) and four constant regions (Cel to Cs4). The molecular weight of IgE is approximately 190,000. The heavy chain comprises approximately 550 amino acids. The structure of IgE is described in Padlan and Davis, Mol. Immunol., 23, 1063-75, 1986) and Heim et al., (The 2IgE structure model was deposited 2/10/90 in a PDB (Protein Data Bank, Research Laboratory for Structural Bioinformatics, http: / pdb-browsers.ebi). ac.uk)). Each of the IgE domains contains a dented barrel of seven antiparallel chains of extended segments (β ~) of the polypeptide labeled on f and divided into two β-sheets. The four β-chains (a, b, d and e) form one sheet opposite the other sheet of the three chains (c, f and g) (shown in Figure 8). The shape of each β-sheet is maintained by lateral packing of the side chain formed by amino acid residues from the side antiparallel chains in each sheet (and is further stabilized by the hydrogen bond between these chains). A loop of residues forming the non-elongated arrangement connects the antiparallel β-sheets, either in the sheet or in the space between the opposing sheets. The connection from string a to string b is designated as loop A-B, and so on. Loops A-B and d-e belong topologically to the four chain leaf and loop f-g belongs to the three chain leaf. Interference between a pair of opposing leaves provides a hydrophobic inner portion of the globular region. This water-free hydrophobic core results from the tight packing of the side chains, which lie opposite each other from opposite β-sheets.

A number of passive or active immunotherapeutic approaches have been investigated in the past and have been identified as interfering with the histamine-mediated mechanism of mediated release

IgE. These approaches involve interfering with IgE or binding allergen / IgE complexes to FcεRI or FcεRII (a low affinity IgE receptor) with either passive administration of antibodies or passive administration of IgE-derived peptides to competitively bind to receptors. In addition, some authors describe the use of specific IgE-derived peptides in active immunization with stimulated release of histamine, which inhibits immune responses.

In the course of their investigation, a number of problems have been considered to be considered when designing new anti-allergic therapies. One of the most serious problems that have arisen in the development of IgE crosslinking in the histamine releasing signal. The most common case is that the production of anti-IgE antibodies during active vaccination is capable of allowing histamine release by itself through the network of surrounding complexes

IgE-receptor in the absence of allergen.

This phenomenon is called anaphylactogenicity.

Of course, a number of commonly available monoclonal antibodies against

IgEs that are normally used in detection assays

IgE are anaphylactogenic and potentially unsafe for use when administered to a patient.

Whether or not the antibodies are anaphylactogenic depends on the location of the target epitope on the IgE molecule. However, based on current knowledge in this field and despite the great interest and efforts of researchers, it is difficult to predict what characteristics any antibody or epitope has and whether or not it has a positive clinical effect in a patient.

Therefore, in order to achieve the safety or efficacy of a passively administered or vaccine-induced antibody, it must bind to an IgE region that is able to interfere with a pathway that promotes histamine release without being anaphylactic in itself. The invention describes all of these objectives and provides drugs that are capable of generating non-anaphylactic antibodies that inhibit histamine release. These drugs may form the basis of an active vaccine, or may be used to produce the right antibodies in the case of passive immunotherapy, or may be passively administered alone for therapeutic effect.

Much work has been done by researchers to identify specific anti-IgE antibodies that have some beneficial effects against an IgE-mediated allergic reaction (WO

90/15878,

WO

89/04834, WO 93/05810). Much effort has also been devoted to identifying epitopes recognized by these useful antibodies in order to generate peptide mimotopes of such epitopes and to be used as immunogens to produce anti-IgE antibodies.

WO 97/31948 describes work of this type and further describes IgE peptides from Cs3 and Cs4 regions conjugated to carrier molecules for active vaccination purposes. These immunogens can be used in vaccine studies and are reported to be capable of generating antibodies that subsequently inhibit histamine release in vivo. Also described herein are monoclonal antibodies (BSW17) which are said to be capable of binding to IgE peptides contained in the Οε3 region, which are useful for active vaccination purposes.

EP 0 477 231 B1 discloses immunogens derived from the Ce4 IgE region (residues 497 to 506, also known as Stanworth decapeptide) conjugated to hemocyanin of the gastric gastropod (KLH) used in active vaccine immunoprophylaxis. WO 96/14333 is a continuation of the work described in EP 0 477 231 B1.

Other approaches are based on the identification of peptides derived from the 0ε3 and 0ε4 regions, which themselves compete for the binding of IgE to receptors on basophils or mast cells with a low level of basophils or mast cells. high affinity

WO 98/24808, EP 0303

SUMMARY OF THE INVENTION (described in WO 93/04173,

625 B1, EP 0 341 290).

The invention discloses are used in therapy. allergies.

IgE sequences that immunoprophylaxis or associated with treatment that incorporate from the continuing portions exposed to the surface and identified epitopes.

IgE, infra

These or identify new active or passive

These sequences were not before

The invention describes peptides themselves, specific isolated epitopes which have been found to provide mimotopes of these novel peptides or mimotopes can be used in the treatment of allergy can be used as a vaccine to induce IgE auto-antibodies during active immunoprophylaxis or allergy, reduce or eliminate immunotherapy in reducing allergic symptoms in vaccinated subjects.

Surprisingly, the anti-IgE antibodies induced by the peptides of the invention are non-anaphylactic and are capable of blocking IgE-mediated histamine release from mast cells and basophils.

Regions of human IgE which are peptides of the invention and which may serve to provide a basis for peptide modification are:

Table 1:

Peptide < sequence the position of the IgE sequence and region SEO ID NO. P5 rasgkpvnhstrkeekqrngtl Cs3 1 P6 GTRDWIEGE Ct3 2 P7 PHLPRALMRSTTKTSGPRA Cs3 / Cs4 3 P8 PEWPGSRDKRT Cs4 (Pro451 - Thr461) 4 P9 EQKDE Ce4 5 P200 LSRPSPFDLFIRKSPTITC Ct3 6 P210 ³ WLHNEVQLPDARHSTTQPRKT Ce4 7 1-90N LFIRKS Cs3 81 2-90N PSKGTVN Ct3 82 3-90N LHNEVQLPDARHSTTQPRKTKGS Ce4 83 4-90N SVNPGK Cs4 ' 84

.............

Peptides that incorporate these epitopes form a preferred aspect of the invention. Also disclosed are mimotopes having the same characteristics as these epitopes, and immunogens comprising such mimotopes that generate an immune response that cross-reacts with an IgE epitope in accordance with an IgE molecule.

The invention further encompasses isolated peptides comprising these IgE epitopes alone and any mimotopes thereof. The term "mimotope" is defined as an entity that is sufficiently similar to the natural IgE epitope to be recognized by antibodies that recognize the natural IgE epitope (Gheysen, M. et al., 1986, Synthetic peptides as Wiley, Chichester, Ciba Foundation Symposium 119, p130-149, Gheysen, M. M., 1986, Molecular Immunology, 23, 7, 709-715), or is capable of generating antibodies when coupled to a suitable carrier. These antibodies cross-react with the natural IgE epitope.

The mimotopes of the invention may be peptide or non-peptide in nature. The peptide mimotope on the surface of exposed IgE epitopes identified above may also have exactly the same sequence as the natural epitope. Such a molecule is described as an epitope mimotope because, although the two molecules share the same sequence, the mimotope will not be present in accordance with the structure of the entire IgE region and as such may have a slightly different spatial arrangement compared to the natural IgE epitope. The skilled artisan will appreciate that the linear sequences identified above (P1 to P7), when lying adjacent to other sequences in the tertiary structure of IgE, are remote from the primary IgE sequence. As such, for example, the P1 mimotope may be fused or interrupted and may contain or mimic P1 segments and segments prepared from these distant amino acid residues.

The mimotopes of the invention mimic the surface exposed area of the IgE structure, but in these areas the surface exposed areas that correlate with the loop structure are considered to be dominant. IgE domain structure is described in "Introduction to Protein Structure (page 304, ^2nd Edition, Branden and Tooze, Garland Publishing, New York, ISBN 0 8153 2305-0) and has the form of beta-barrel, which originated from two opposed parallel beta-sheets (shown in Figure 8). The mimotopes may therefore comprise a loop with an extension of the N- and C-termini, which may be natural amino acid residues from surrounding leaves. As an example, the P100 region that contains the AB Cs3 loop, P8 contains the AB Cs4 loop, P5 includes the CD Cs3 loop, and P110 contains the CD Ce4 loop. Similarly, mimotopes of these loops form an aspect of the invention. Particularly preferred loops are CD Cs3 or Cs4 loops and AB Cs4 loop.

Peptide mimotopes of the above identified IgE epitopes may be designed for certain purposes by the addition, deletion, or substitution of the selected amino acids. Thus, the peptides of the invention may be modified to facilitate conjugation to a protein carrier. For example, for some chemical conjugation methods, it may be necessary to include terminal cysteine in the IgE epitope. In addition, for peptide-conjugated peptides, they may need to include a hydrophobic tail remote from the conjugated tail of the peptides such that the free conjugated tail of the peptides remains attached to the surface of the carrier protein. This reduces the conformational degrees of freedom of the peptides and thus increases the likelihood that the peptide is present in the conformation closest to that of the IgE peptides as found consistent with the entire IgE molecule. For example, the peptides may be altered to have an N-terminal cysteine and a C-terminal hydrophobic amidated end. Alternatively, addition or substitution of the D-stereoisomeric form of one or more amino acids may be made to provide a preferred derivative, for example, to enhance the stability of the peptide. It will be appreciated by those skilled in the art that such modified peptides or mimotopes could be wholly or partially mimotopes that are not peptides wherein the essential residues are not necessarily limited to 20 naturally occurring amino acids. In addition, these moieties can be cyclized by methods known in the art to produce a peptide in a conformation that closely resembles a shape where the peptide sequence is consistent with the entire IgE molecule. A preferred method of cyclizing the peptide comprises adding a pair of cysteine residues to allow the formation of a disulfide bridge.

The skilled artisan will appreciate that the mimotopes or immunogens of the invention may be larger than the epitopes identified above and as such may contain the sequences described herein. Similarly, mimotopes of the invention may comprise additional N- and / or terminalminal extension of a number of other natural residues at one or both ends. The peptide mimotopes may also be retro-sequences of natural IgE sequences in which the sequence orientation is reversed, or alternatively may contain all or part of the D-stereoisomeric amino acids (inverse sequences). The peptide sequences may be retro-inverse in nature, that is, the orientation of the amino acids is reversed and the amino acids are in the stereoisomeric form. Such retro- or retro-inverse peptides have the advantage that they occur alone and as such circumvent the problems of self-tolerance of the immune system (for example P14c).

Alternatively, peptide mimotopes can be identified using antibodies that are themselves capable of binding to IgE epitopes of the invention using methods such as phage-expressing technology (EP 0 522 267 B1). This procedure generates a large number of peptide sequences that mimic the structure of natural peptides and are therefore capable

bind to antibodies against the native peptide, but cannot necessarily share substantial sequence homology with the native IgE peptide. This approach may be advantageous by allowing the identification of peptides with enhanced immunogenic properties (such as binding characteristics with higher affinity for IgE receptors or anti-IgE antibodies or capable of inducing a polyclonal immune response, wherein said antibodies bind to higher affinity IgE) or they can bypass any potential tolerance problems to the antigen itself that are associated using the native peptide sequence. In addition, this method allows the recognition of a recognition pattern for each natural peptide based on its shared chemical properties among the recognized mimotope sequences.

Examples of such mimotopes are:

Peptide- sequence description ' SEO ID NO. Pil CRASGKPVNHSTRKEEKQRNGLL P5 mimotope » 8 Saw (Ac) GKPVNHSTGGC P5 mimotop 9 PI lb (Ac) GKPVNHSTRKEEKQRNGC P5 mimotop 10 Lung CGKPVNHSTRKEEKQRNGLL (NH,) P5 mimotope ' 11 PI Id (Ac) RASGKPVNHSTGGC P5 mimotop 12 P12 CGTRDWIEGLL P6 mimotop 13 P12a CGTRDWIEGETL (NH) P6 mimotope: 14 P12b (Ac) GTRDWIEGETGC P6 mimotop 15 Dec P13 CHPHLPRALMLL P7 mimotop » 16 P13a CGTHPHLPRALM (NH) P7 mimotop 17 P13b (Ac) THPHLPRALMRSC P7 mimotope · 18 P13c (Ac) GPHLPRALMRSSSC P7 mimotop 19 Dec P14 APEWPGSRDKRTC PS mimotop 20 May P14a (Ac) APEWPGSRDKRTLAGGC P8 mimotop 21 PI4b CGGATPEWPGSRDKRTL (NH) P8 mimotop 22nd P14c CTRKDRSGPWEPA _(NH2) P8 retro 23 P14d * (cycle) (Ac) APCWPGSRDCRTLAG P8 mimotop. 24 P14d (cycle) (Ac) ACPEWPGSRDRCTLAG P8 mimotop 25 C-1C14 CATPEWPGSRDKRTLCG P8 mimotop 26 C-1Cl3 CATPEWPGSRDKRTCG P8 mimotop 27 Mar:

«·

| C3C12 TPCWPGSRDKRCG P8 off top 28 'P9a CGAEWEQKDEL (NH) P9 mimotop 29 P9b (Ac) AEWEQKDEFIC P9 mimotop 30 P9b * (Ac) GEQDEFIC P9 mimotop 31 P9a * CAEGEQKDEL (NH) P9 mimotop. 32 Čarl] CPEWPGCRDKRTG P8 mimotop 85 Carl2 TPEWPGCRDKRCG P8 mimotop 86

Alternatively, peptide mimotopes can be generated to enhance the immunogenicity of a peptide by increasing its affinity for a polyclonal antibody against an IgE peptide, the effect of which can be measured by methods known in the art, such as Biocore experiments. To achieve this peptide sequence, the following general rules can be selectively changed:

• maintain structural constraints, that is, proline and glycine should not be replaced • other positions may be substituted with an amino acid that has similar physicochemical properties.

That is, the amino acid residue may be replaced by an amino acid that is most similar to the amino acid. For example, A can be replaced by V, L or I as described in the following table.

the original rest examples of substitutions preferred substitution AND V, L, I IN R K, Q, N TO N Q, Η, K, R Q D E E C WITH WITH Q N N E D D G AND AND H N, Q, K, R N AND L, V, M, A, F. L L I, V, M, A, F. AND TO R, Q, N R | M L, F, I 1 L

F L, V, I, A, Y, W W P AND AND with T T T WITH WITH w Y, F Y Y W, F, T, S F in I, L, M, F, A. L

Particularly preferred IgE peptides are P8 and variants thereof (such as P14 or P14a). These peptides, when coupled to a carrier, are potent in inducing an immune response against IgE. This immune response is capable of inhibiting histamine release from human basophils. P8 variants or mimotopes are described primarily as any peptide-based immunogen that is capable of inducing an immune response that is able to recognize P8. Without being limited by the scope of the invention, some P8 variants may be described by the general formula in which certain amino acids may be replaced by their closest complement. Using this procedure, peptide mimotopes of P8 can be described by the general formula:

P, Xlø X ₂ , Ρ, Χβζ X ₄ of ΧδΛ% 6r ΧδΛ or

Ρ, X _1f X ₂ , P, G, X ₄ , R where , D, X ₅ , Xs symbol Xi Yippee amino acid selected of E, D, N or symbol X ₂ Yippee amino acid selected of W, Y or F symbol X3 Yippee amino acid selected of G or A, symbol X ₄ Yippee amino acid selected of S, T or M. symbol X ₅ Yippee amino acid selected of R or K a symbol X ₆ Yippee amino acid selected of D or E.

P8 mimotopes can also be identified using antibodies that are capable of binding to P8 themselves, further using techniques such as phage-expressing technology (described in EP 0 552 267 B1). Monoclonal antibodies such as P14 / 23, P14 / 31, P14 / 33 are particularly suitable in this regard.

The invention therefore describes novel epitopes and mimotopes thereof and their use in the manufacture of pharmaceutical compositions for the prophylaxis or therapy of allergy. Further disclosed are immunogens comprising at least one of the epitopes or mimotopes of the invention and carrier molecules that are suitable for use in vaccines for immunoprophylaxis or allergy therapy. Similarly, the epitopes, mimotopes or immunogens of the invention are suitable for use in medicine and in the treatment or prophylaxis of an allergic disease. Preferred immunogens and vaccines of the invention comprise a P8 IgE epitope or mimotopes thereof that include P14.

Various methods by which an epitope or mimotope are presented have been shown to have substantial effects on binding to monoclonal antibodies and immune response following vaccination. For example, when cyclic peptides are used, the length and phase of the loop are altered, this may have significant effects on the binding activity of cyclic mimotopes to monoclonal antibodies P14 (P14 / 23, P14 / 31 or P14 / 33). A new system has been developed which selects cyclization sites, thereby increasing the likelihood that the cyclic peptides maintain the correct loop structure containing the correct amino acid residues. In this way, the protein is likely to be encapsulated in a conformation that most resembles that conformation that peptides would normally conform to when they were consistent with the entire IgE region. Therefore, cyclic mimotopes without limitation by the invention that meet these new rules constitute a preferred aspect of the invention.

Putative mimotope sequences that do not meet these rules can still produce a useful antiserum (e.g., P14 and Pil), such the following examples being only subset of mimotope types of the invention.

Examples of preferred peptides that meet these newly defined structural rules are:

Table 3

peptide sequence Mim otop SEO ID NO. CSRPSPFDLFIRKSPTITC A-BsmvčkaCs3 33 CSRPSPFDLFIRKSPTC A-BsmyčkaCs3 35 CPSPFDLFIRKSPTITC A-BsmyčkaCs3 41 CPSPFDLFIRKSPC A-BsmyčkaCs3 43 CTWSRASGKPVNHSTC CD ^sm yčkaCs3 58 CTWSRASGKPVNHC C-DsmyčkaCs3 60 CSRASGKPVNHSTC C-Dsmyčkac _s 3 66 CSRASGKPVNHC C-DsmyčkaCs3 68 CYAFATPEWPGSRDKRTLAC A-BsmyčkaCs4 45 CYAFATPEWPGSRDKRTC A-BsmyčkaCs4 47 CFATPEWPGSRDKRTLAC A-Bs loopCe4 53 CFATPEWPGSRDKRTC A-BsmyčkaCs4 55 CQWLHNEVQLPDARHC Č-DsmyčkaCs4 70 CQWLHNEVQLPDAC C-D loopCe4 72 CLHNEVQLPDARHC C-DsmyčkaCs4 78 CLHNEVQLPDAC C-DsmyčkaCs4 80

The mimotopes of the invention are believed to be small, mimicking a region selected from the entire IgE region in which a natural epitope is detected.

less than 100 contain less than 75

The peptide mimotopes would have amino acids, amino acids other than 50 amino acids, and up to 25 amino acids.

contain less contain preferably more preferably and most preferably Specific should therefore be examples of preferred peptide mimotopes are P14 and PH, which contain 13 and 23 amino acids. The non-peptide mimotopes are believed to be the same size based on the volume of the molecule as compared to their peptide supplements.

It is known in the art which methods can be used to confirm the status of a specific construct as mimotope, which is within the scope of the invention. Such methods include, but are not limited to, the following. In a putative mimotope, it is possible to test the immunogenicity of an antisera-forming construct where the putative mimotope cross-reacts with the natural IgE molecule, and are also useful in blocking the release of allergic mediators from allergic effector cells. The specificity of this response can be confirmed by competition experiments by blocking the activity of the antisera with mimotope alone or natural IgE and / or specific monoclonal antibodies known to bind to an epitope in IgE. Specific examples of such monoclonal antibodies suitable for use in competition assays include P14 / 23, P14 / 31 or P14 / 33, which confirm the status of a putative mimotope as P8 mimotope. · '

In one embodiment of the invention, at least one IgE epitope or mimotope is linked to carrier molecules to produce immunogens suitable for vaccination protocols. It is preferred that the carrier molecules are not related to the native IgE molecule. The mimotopes may be linked by chemical covalent conjugation or by expression of genetically manipulated fusion partners using a linker sequence. In one embodiment, the peptides of the invention are expressed in a fusion partner fusion molecule, wherein the peptide sequence is detected in the primary fusion partner sequence.

Covalent coupling of the peptide to an immunogenic carrier can be accomplished in a manner well known in the art. For example, in the case of direct covalent coupling, carbodiimide, glutaraldehyde or (Ν- [γmaleimidobutyryloxy] succinimide ester using conventional commercially available heterobifunctional linkers such as CDAP and SPDP (using manufacturer's instructions) may be utilized. method, gel filtration and fractionation method etc ..

·· • ♦ · ·

In a preferred embodiment of the invention, it has been found that peptides, in particular cyclic peptides, can be conjugated to a carrier by preparing derivatives of an acylhydrazine peptide.

Peptide / protein carrier constructs can be produced as follows. Acylhydrazine peptide derivatives can be prepared on the solid phase as shown in Scheme 1 below:

Scheme 1:

Solid phase peptide synthesis:

Rink-resin solid phase peptide synthesis

N-AAj ... C (Trt) ...... C (Trt) ...... AA _n -Lys (Dde) -Rink-Resin Hydrazine Hydrate

Ý

N-AAi ... C (Trt) ...... C (Trt) ...... AA _n -Lys (NH ₂ ) -Rink-resin (i) succinic anhydride (ii) HBTU / HOBt / NMM / N ₂ H ₄

IN

^-ΑΑχ .-. C (Trt) ...... C (Trt) ...... AA-Lys-Rink-Resin

N HZ-

íi o

NHN 4

TFA i

AA-AAi ... C (-SH) ...... C (-SH)

.AA _n -Lys-CONH ₂ 1 N HZ- 0 z = 0

oxidation of H ₂ O ₂ t

X-AAi ... C ... C ... AA _n -Lys-CONH ₂ I

N HZThese peptide derivatives can be well known in the art using the well known Fmoc or polyethylene glycol polystyrene automated equipment process by a process that is easy to prepare using a polyamide (PEG-PS) support (methods and procedures for solid synthesis are described in Solid. Phase stage how to

Peptide

Synthesis: A

Practical Approuch

Oxford University Acid Provides

Sheppard, IRL, mediated linear, unprotected and modified peptide.

by E. Atherton and R.C.

Fission

This peptide is readily oxidized and purified to form a disulfide-bridged epitope using the methodology described in "Methods in Molecular Biology, Vol. 35: Peptide Synthesis Protocols (edited by M. W. Pennington and M. Dunn), chapter 7, pp. 91-171, D. Andreu et al.,.

The peptides thus synthesized can then be conjugated to proteinaceous carriers using the following methods:

The introduction of the arylaldehyde function utilized the active succinimide ester (BAL-Osu) prepared as shown in Scheme 2 (further details are described in WO 98/17621). Substitution of amino functions with, for example, BSA (bovine serum albumin up to 50%) supports soluble soluble protein. Greater BSA substitution results in insoluble constructs. BSA and BAL-Osu were mixed at equimolar concentration in DMSO / buffer (shown in the diagram) for 2 hours. This experimentally obtained protocol gives rise to approximately 50% substitution, as estimated by the fluorescent amine assay for free amino groups, in the following scheme 2 / 3- preparation of the modified carrier:

Scheme 2:

Scheme 3:

BSA-BAL · 444 · 4

A simple combination of engineered peptides and derivatized carrier provides peptide carrier constructions that can be easily isolated by dialysis - Scheme 4 - peptide / carrier conjugate.

Scheme 4:

I — I

X-AAj ... C .... C .... AA _n -Lys-CONH ₂

NH-Z

50% DMSO / Buffer (pH 3.5, 0.1 MNaHCO ₂ )

-8-16 h.

' ^r

NH

Lys AA ... _n ... C ..., .....

C ... AA _r X

The types of carriers used in the immunogens of the invention are well known in the art. The function of the carrier is to provide cytokine assistance to help induce an immune response against the peptides

IgE.

The list of carriers that can be used according to the invention includes:

gastric gastric hemocyanin (KLH), serum albumins such as bovine serum albumin inactivated bacterial toxins such as tetanus toxin or diphtheria (TT and DT) or recombinant fragments thereof (for example, TT fragment C fragment TT or translocation region DT) or purified protein a tuberculin derivative (PPD). Alternatively, the mimotopes or epitopes may be directly conjugated to liposome carriers, which may additionally contain immunogens capable of providing helper T-cells. It is preferred that the mimotope to carrier ratio be 1: 1 to 20: 1 and it is preferred that each carrier carries between 3 to 15 peptides.

In one embodiment of the invention, the preferred carrier is protein D from Haemophilus influenzae (EP 0 594 610 B1). Protein D is a protein binding

IgD from Haemophilus influenzae and is protected by patent WO 91/18926, EP 0 594 610 B1). In some circumstances, for example in recombinant immunogenic expression systems, it may be necessary to use fragments of protein D, for example protein D 1/3 ^Ed (comprising the N-terminal 100 to 110 amino acids of protein D (described in GB 9717953.5)).

Another preferred recombinant presentation method is in accordance with EP 0 of the invention

For example, nuclear antigen particles in particles similar to the viral IgE peptides of the fusion molecule.

421,635 B to chimeric particles are described using a hepadnavir wherein foreign peptide sequences are present. As such, the immunogens of the invention may comprise IgE peptides present in chimeric particles containing the hepatitis B nuclear antigen. In addition, the recombinant fusion proteins may comprise mimotopes of the invention and a carrier protein such as NS1 influenza virus. In the case of any recombinantly expressed protein forming part of the invention, the nucleic acid that encodes said immunogen is also an object of the invention.

The peptides of the invention can be readily synthesized by a solid phase method well known in the art. Suitable syntheses can be accomplished using "T-boc" or "Fmoc" procedures. The cyclic peptides can be synthesized by a solid-phase process using a well-known F-moc and polyamide resin process in a fully automated apparatus. Alternatively, those skilled in the art will know the necessary laboratory procedures to carry out the procedure manually. Solid phase synthesis techniques and procedures are described in "Solid Phase Peptide Synthesis: A Practical Approach" by E. Atherton and R.C. Sheppard, IRL, Oxford University Press (1989). Alternatively, the peptides can be produced by recombination methods that involve expression of nucleic acid molecules in a bacterial and mammalian cell line. Then the expressed mimotope. protein Maniatis,

Methods of recombinant coding for mimotopes are followed by peptide purification and clonning expression,

Laboratories are known

T., Fritsch, in the art and are described in U.S. Pat

E. F. and Sambrook et al., A laboratory

Press, Cold Spring Harbor, publication

Molecular manual 3 ^nd

Ed., Cold Spring

New York (1989).

contain peptides,

Harbor

The immunogens of the invention may include, including mimotopes or described above, as analogs or fragments thereof. The invention is to be cross-reactive derivatives or further described by nucleic acid portions that encode the immunogens of the invention or peptides, mimotopes or derivatives thereof.

The invention further provides the use of novel epitopes or mimotopes (as defined above) in the manufacture of pharmaceutical compositions for the prophylaxis or therapy of allergies. Immunogens comprising the mimotopes or peptides of the invention and carrier molecules are also suitable for use in vaccines for immunoprophylaxis or allergy therapy. Similarly, the mimotopes, peptides or immunogens of the invention are suitable for use in medicine and in the medical treatment or prophylaxis of an allergic disease.

The vaccines of the invention may advantageously also include an adjuvant. Suitable adjuvants for the vaccines of the invention include those adjuvants that are capable of enhancing antibody responses

* 44 against IgE peptide immunogen. Adjuvants are well known in the art (Vaccine Design - The Subunit and Adjuvans Approuch, 1995, Pharmaceutical Biotechnology, Volume 6, Eds. Powell, MF, and Newman, MJ, Plenum Press, New York and London, ISBN 0-306-44867- X). Preferred adjuvants suitable for use with the immunogens of the invention include aluminum or calcium salts (hydroxide or phosphate).

The vaccines of the invention may generally be administered as a primary or as an booster dose. Booster doses are expected to be adequately separated or administered early, at a time when the amount of circulating antibody is less than the desired value. Booster doses may contain the peptide in the absence of the parent carrier molecule. Such constructions suitable as a booster dose may include an alternative carrier or may exist even if no carrier is present.

The invention further provides an immunogen or vaccine described herein for use in medicine.

The vaccine compositions of the invention may be used to protect or treat a mammal susceptible to or suffering from an allergy by administering said vaccine by the systemic or mucosal route. These applications may include intramuscular, intraperitoneal, intradermal or subcutaneous injection or mucosal administration into the oral / digestive, respiratory, genitourinary tract. The preferred route of administration is the transdermal route, for example, through skin patches. Similarly, there is provided a method of treating allergy comprising administering to a patient suffering from or susceptible to allergy the peptides, immunogen or ligand of the invention.

The amount of protein in each vaccine dose is selected as an amount that induces immunoprotective protection without ·· · ”·: · · ··.

•. · ·; : * i; · ······· ... ...

significant adverse reactions that occur with typical vaccines. Such amount will vary depending upon the specific immunogen used and the presentation method. Generally, it is expected that each dose will contain 1 to 1000 pg of protein, preferably 1 to 500 pg, more preferably 1 to 1000 pg, with a range of 1 to 50 pg being the most preferred range. The optimum amount for a particular vaccine can be ascertained by standard studies involving observation of a suitable immune response in the subject. Following initial vaccination, subjects may be administered one or more adequately divided booster doses.

The invention further provides ligands capable of binding to the peptides of the invention. Examples of such ligands are antibodies (or Fab fragments). The use of ligands in medicine and in the manufacture of medicaments for the treatment of allergies is also described. The term "antibody" refers to a molecule that exhibits useful antigen binding specificity. It will be appreciated by those skilled in the art that the term also encompasses polypeptides that are fragments or derivatives of antibodies and which may have the same or similar function. Such antibody fragments or derivatives also fall within the term antibody.

Particularly preferred ligands are monoclonal antibodies. P14 / 23, P14 / 31 or P14 / 33, and are monoclonal antibodies that recognize P8 (which are vaccinated with P14 immunogen). Hybridomas of these antibodies were deposited under the Budapest Convention at ECACC (European Collection of Cell Cultures, Vaccine Research and Production Laboratory, Public Health Laboratory Service, Porton Down, Salisbury, Wiltshire, SP4 OJG, UK) on January 26 2000 with accession number 00012610, 00012611, 00012612. An important aspect of the invention is the use of monoclonal antibodies in the identification of new

* ·· • · ♦ • · * · · • 9 • • · • • Λ • • • t • · ♦ · • · · 9 « * ·· 9 · * · • · aa • a

IgE mimotopes for subsequent use in the treatment of allergy and use of antibodies in the manufacture of a medicament for the treatment or prophylaxis of allergy. All monoclonal antibody functions in vivo inhibit histamine release from human basophils as well as P14 / 23 and P14 / 31, and have also been shown to inhibit passive cutaneous anaphylaxis in vivo.

Therefore, IgE Cs4 mimotopes that are capable of binding to P14 / 23, P14 / 31 or P14 / 33 and immunogens comprising these mimotopes form an important aspect of the invention. Vaccines containing mimotopes that are capable of binding to p14 / 23, P14 / 31 or P14 / 33 are useful in the treatment of allergy.

In addition, antibodies elicited in one animal by vaccination with the peptides or immunogens of the invention can be purified and passively administered to another animal for the prophylaxis or therapy of allergy. The peptides of the invention can be used to generate hybridomas of monoclonal antibodies (using the novel methods described, for example, in Kohler and Milstein, Nature, 1975, 256, p495), humanized monoclonal antibodies or monoclonals incorporating CDRs, by methods known in the art. Such antibodies can be used in passive immunoprophylaxis or immunotherapy or are used to identify mimotopes of an IgE peptide.

They can be used as ligands of the invention in the prophylaxis or treatment of allergy, providing pharmaceutical compositions comprising the ligands of the invention. Preferred pharmaceutical compositions suitable for the treatment or prophylaxis of allergy include the monoclonal antibodies P14 / 23, P14 / 31 or P14 / 33.

The invention may further describe the use of diagnostic tests. For example, a set of ligands that recognize the various peptides of the invention can be used in titer testing

• ·· • · « • ·· • ·· ·· • • ·· ♦ · · • · • • • • • 4 · • • ·· ···· ··· • ··· • ·· • ··

anti-IgE antibodies in the serum collected from the patient. However, the peptides can themselves be used to type circulating anti-IgE. In some circumstances, it is appropriate to test the amount of circulating anti-IgE, for example in atopic patients, and as such, the peptides and poly / monoclonal antibodies of the invention may be used to diagnose atopy. In addition, the peptides can be used to affinity remove circulating anti-IgE from patient blood prior to re-introduction of blood to the patient by infusion.

The invention further provides a method of identifying peptide immunogens for immunoprophylaxis or allergy therapy, which comprises using a computer model of the IgE structure and identifying those IgE peptides that are exposed on the surface. These regions can then be introduced into immunogens and used in medicine. Similarly, the use of P14 / 23, P14 / 31 or P14 / 33 in identifying peptides for use in immunoprophylaxis or allergy therapy forms part of the invention.

Vaccine preparation is generally described in New Trends and Developments in Vaccines, ed. Volier et al., University Park Press, Baltimore, Maryland, U.S.A. The conjugation of proteins to macromolecules is described in Likhite, U.S. Patent No. 4,372,945 and Armor et al., U.S. Patent No. 4,474,757.

Overview of the drawings

Figure 1 shows the surface exposure of Cs3 and Cs4 of human IgE as calculated from the model of Padel and Davis 1986.

Figure 2 shows inhibition of histamine release and anaphylactogenicity of P14 antiserum. Monoclonal antibodies PTmAb0005 and PTmAbOO11, which are used as positive controls, were added at a concentration of 1 μς / ιπΐ to serum against BSA

diluted 1/100 and 1/500 (final dilution). Antisera against P14 were added to a final dilution of 1/100 and 1/500. Cells were obtained from allergic patients who are sensitive to grass pollen. Histamine release was performed by incubation with grass pollen allergen.

Figure 3 shows inhibition of histamine release and anaphylagenicity of antiserum against P14. P14 antiserum from different mice was added in different dilutions (80x or 40x). to contain approximately 1 µg / ml anti-IgE antibodies as determined by the IgE receptor binding ELISA. Three negative controls were used: anti-BSA antisera, non-specific IgG1 and a mixture of non-specific IgG1 diluted in anti-BSA antisera. The mAb11 mAb is a monoclonal antibody known to inhibit histamine release and is used as a positive control (added at 2 μς / ml).

Figure 4 shows inhibition of histamine release and anaphylactogenicity of anti-P14 antiserum. Anti-P14 antisera from various mice were added to a final dilution of 1/50. Monoclonal antibodies were added at a concentration of 2 µg / ml either in assay buffer or in anti-BSA serum at a 1/50 dilution. The following three negative controls were used as anti-BSA antisera, non-specific IgG1 and a mixture of non-specific IgG1 diluted in anti-BSA antisera. MAb11 is a monoclonal antibody known to inhibit histamine release and was used as a positive control (added at a concentration of 2 μg / ml).

Figure 4 shows antibody response against P1 peptides. P1 peptide was loaded at 1 μς / πιΐ in carbonate buffer at 4 ° C overnight. After saturation of the plates, serially twice diluted serum was added and incubated for one hour at 37 ° C. Bound IgG was detected with anti-mouse biotin-labeled antibody, then streptavidin POD and TMB substrate. The time points are A. 14 days after vaccination 1 and 14 days after vaccination 2, B. 14 days after vaccination 3.

Figure 6 shows anti-P11 IgG titers of anti-human IgE. Human IgE was loaded at a concentration of 1 μς / ιηΐ. Twice serially diluted serum ("collected BSA is collected from control group") or PTmAb0005 (positive control monoclonal antibody) was incubated for one hour at 37 ° C. Bound IgG was detected with an anti-mouse antibody labeled with biotin.

Figure 7 shows histamine release studies with monoclonal anti-P14 antibodies with allergic basophils obtained from mite allergic patients (A10 and All) and grass pollen (G8 and G4). PT11 (PtmAbOO11) was used as a positive control and non-specific IgG2 was used as an isotype control for P14 / 23, P14 / 31 and P14 / 33.

Figure 8 shows the structure of the IgE region. (A) Each region consists of two opposing beta-sheets, one of 4 anti-parallel beta-chains (labeled 4) and the other of three anti-parallel beta-chains (labeled 3). (B) The seven strings are represented topographically as a block indicated by an arrow up to f divided between two sheets as shown. Loop string connectivity is topologically represented in the curve by arrows: blank arrows indicate loops inside a sheet, and dashed arrows indicate loops between sheets. In the structures of the IgG1 Fc region, the short chain c forms part of the C-D loop as predicted for the Fc IgE region.

Figure 9 shows A) the predicted structural alignment of the A-B loop sequences of the human IgE Cs2, 3 and 4 regions with equivalent segments from the crystallographically determined Fc structure of human IgG1 (Cy2 and Cy3 regions).

The beta chains in the IgG1 structure are underlined and designated as a and b. The amino acid residues of each chain are numbered. The vertical arrows below the sequence block indicate the expected optimum crystallization positions, indicated and connected by a dashed and continuous line, as shown in Figure 10b. (B) the predicted structural alignment of the c-d loop of Cs2.3 and 4 of human IgE as compared to the Fc of human IgG1. The beta chains in the IgG1 structure are underlined and are designated c, c'and d. The amino acid residues at the ends of each segment of the sequence are numbered. The residues in the dashed boxes form (Cy2 and Cy3) or are believed to form (Cs2, by improving the homology model and experiment, Cs3, Cs4 by modeling the homology) a protected core in the loop. It is believed that residues in the empty boxes are involved in the recognition of receptors and / or antibodies. The vertical arrows below the sequence blocks indicate the expected optimal cyclization positions, indicated and connected by a dashed and continuous line, as shown in Figure 11b.

Figure 10 (A) shows a schematic structure of the hairpin A-B in the leaf-list constant region of the Ig constant regions. Adjacent antiparallel beta-chains are represented by empty arrows and are denoted as a and b. Residues in chain a are labeled i, residues in chain b are labeled j. Residues i + n and n + m, where both a and n are 0 or equal, part of the interspace leaf-list in the area. The residues i + a and j + m, where n and m are odd numbers, form part of the surface of the solvent exposed area. Loop A-B is shown as a black arrow. Figure 10 (B) shows a schematic structure of the hairpin A-B, as in Figure 3a, with residue positions optimal for cyclization coupled with dashed or continuous lines.

Figure 11 (A) shows a schematic structure of the hairpin C-D (loop plus beta-chain support) at the edge of the inner space of the leaf-list of constant Ig regions. Opposite anti-parallel beta-chains are shown as solid arrows denoted by c and d. The residues along the c chain are denoted i, the residues along the d chain are denoted j. The residues i + n and j + m, where A is an odd number but m is an even number, form part of the leaf-list interspace in the region. The residues i + n and j + m, where n is 0 or an even number, but m is an odd number, form part of the surface area of the solvent exposed area. Loop c-d containing the short chain c'is represented by a black arrow. Figure 11 (B) shows a schematic structure of the hairpin c-d with residue positions that are optimal for cyclization linked by a dashed and continuous line.

DETAILED DESCRIPTION OF THE INVENTION

Part 1: Active vaccination studies

Example 1:

1.1: Identification of peptide

The peptides were identified as follows.

Model structures of human IgE are described in Padlan and Davies Mol. Immunol., 23, 1063-75 (1986).

Peptides were identified which were both continuous and exposed in solvent. This was achieved using molecular simulation software (MSI) to calculate the accessibility of each amino acid of IgE. Zprůměrizoval accessible surface via the transparent window of five residues, and thereby determine the IgE peptide region having a diameter greater than said pětiméru 80 A ^second

The test results are shown in Figure 1.

• ·

Results

In Figure 1, there are natural peptide numbers that can be used as immunogens for the production of anti-IgE antibodies.

Table no. 4: Natural exposed surface and continuous IgE peptides using the model described in Padlan and Davies et al., 1986

Peptide sequence the sequence position and the IgE region SEQ ID NO. P5 RASGKPVNHSTRKEEKQRNGTL Ce3 1 P6 GTRDWIEGE Ce3 2 P7 PHLPRALMRSTTKTSGPRA Ce3 / Ce4 3 PS PEWPGSRDKRT Ce4 (Pro451-Thr462) 4 P9 EQKDE Ce4 5 P200 LSRPSPFDLFIRKSPTITC Ce3 6 P210 WLHNEVQLPDARHSTTQPRKT Ce4 7

In addition to these peptides identified above using the same selection criteria with the IgE model described by Helm et al. the following peptides were identified (the 2IgE structural model deposited 2/10/90 in PDB (Protein Data Bank, Research Collaboratory for Structural Bioinformatics, http: / pdb-browsers.ebi.ac.uk)).

Table 5: Peptides identified using the

Helma at al 1990

name sequence position' SEQ ID NO. 1-90N LFIRKS Ce3 81 2-90N PSKGTVN Ce3 82 3-90N LHNEVQLPDARHSTTQPRKTKGS Ce4 83 4-90N SVNPGK Ce4 84

• · ·

These peptides or mimotopes thereof were synthesized and coupled to carrier proteins for use in immunogenicity studies.

Example 1.2: Synthesis of IgE / Protein D peptide conjugates using succinimide maleimide crosslinking agent

Protein D can be coupled directly to IgE peptides to produce antigens of the invention using a succinimide maleimide crosslinking agent. This chemical procedure allows the controlled activation of the NH ₂ carrier residues by fixing the succinimide group. Maleimide groups are cysteine binding sites. Therefore, for the purposes of the following examples, IgE peptides require the addition of an N-terminal cysteine to conjugate.

The coupling agent is a selective heterobifunctional crosslinking agent wherein one end of the compound activates the amino group of the protein carrier with the succinimidyl ester and the other end attaches the sulhydryl group of the peptides to the maleimido group. The reaction scheme is as follows:

a. Activation of protein by reaction between lysine and succinimidyl ester

b. Interaction between activated protein and peptide cysteine

Example 1.3: Preparation of IgE-D protein conjugate

Protein D was dissolved in phosphate buffered saline, pH 7.2, and the concentration was 2.5 mg / ml. The interaction reagent (β-γ-maleimidobutyryloxy) succinimide ester (GMBS) was dissolved in DMSO at a concentration of 102.5 mg / ml and a protein solution was added. 1.025 mg of GMBS was used for 1 mg of protein D. The reaction solution was incubated for one hour at room temperature. The by-products were removed in the salt removal step on a sephacryl 200HR permeation gel. The eluent is phosphate buffered saline with 0.1% Tween 80 at pH 6.8. Activated proteins were collected and pooled. Peptides (as shown in Table 4 or 5 or derivatives or mimotopes thereof) were dissolved at a concentration of 4 mg / ml in 0.1 M acetic acid to prevent S-S binding. A molar ratio of 2 to 20 peptides per 1 activated protein D was used for the interaction. The peptide solution was slowly added to the protein and the mixture was incubated for 1 hour at 25 ° C. The pH was maintained at 6.6 during the interaction phase. The completion step was performed by adding cysteine (0.1 mg cysteine per milligram of activated -PD dissolved at 4 mg / ml acetic acid at 0.1 M) for 30 minutes at 25 ° C and a pH of 6.5 . To remove excess cysteine or peptide, two dialysis against NaCl was performed at a concentration of 150 mM 0.1% Tween 80.

The final step is sterile filtration through 0.22 micron pore membranes. The final product is a filterable solution preserved at 4 ° C. The final peptide / PD ratio can be determined by amino acid analysis.

In an analogous manner, peptides of the invention may be conjugated to other carriers that include BSA. Pre-activated BSA can be conveniently obtained from Pierce Inc.

P8 (P14, SEQ ID NO: 20, CLEDGQVMDVDLL) and P5 (PH, SEQ ID NO: 8, CRASGKPVNHSTRKEEKQRNGLL) mimotopes were synthesized, which were coupled to protein D and BSA using the procedures described above.

Example 1.4: ELISA Methods

ELISA against peptide or peptide carrier

Immune responses against the peptide and the vehicle were examined using the ELISA procedure below. Microtiter plates (Nunc) were coated with specific antigen in PBS (at 4 ° C overnight) with either 2 gg / ml streptavidin (followed by incubation with biotin-labeled peptide (1 μΜ) for one hour at 37 ° C, Wash three times with 0.1% PBS-Tween 20. Plates were saturated with PBS-BSA 1% -Tween 20 1% buffer (Sat buffer) for 1 hour at 37 ° C. 2-step dilution (in Sat buffer), incubated for 1 hour 30 min at 37 ° C. Washing three times, adding 2 ° anti-mouse Ig (or anti-mouse isotype specific monoclonal antibodies) associated with HRP. Incubation was performed with TMB (BioRad) for 10 minutes at room temperature in the dark with a blocking reaction of 0.4 N H2SO4.

Method for Detection of Reactivity Against Human IgE in Mouse Serum (ELISA Assaying IgE Binding to Plates)

ELISA plates were coated with 1 µg / ml human chimeric IgE at pH 9.6 with carbonate / bicarbonate coating t buffer for one hour at 37 ° C or overnight at 4 ° C. Non-specific binding sites are blocked with PBS / 0.05% Tween 20 containing 5% (w / v) Marvel milk powder for 1 hour at 37 ° C. Serial dilutions of mouse serum in PBS / 0.05% Tween 20/1% (w / v) BSA / 4% serum of newly born calves were then added and incubated for 1 hour at 37 ° C. Binding to polyclonal serum was detected in goat anti-mouse IgG-biotin (1/200) followed by streptavidin-HRP (1/1000). Conjugated antibodies were detected with TMB substrate at 450 nm. A standard PTmAbOO11 curve is included on each plate so that it is possible to calculate IgE reactivity in pg / ml in serum samples.

Competition for IgE binding with mimotope peptides, soluble IgE or PTmAbOO11

Single dilution of polyclonal mouse serum was mixed with a single concentration of either mimotope peptide or human IgE in a pre-blocked 96-well polypropylene plate. The mixtures were incubated for 1 hour at 37 ° C and then added to IgE coated ELISA plates for 1 hour at 37 ° C. Binding of polyclonal serum was detected with biotin labeled goat anti-mouse IgG (1/2000) and then streptavidin-HRP (1/1000). The conjugated antibody was detected with TMB substrate at 450 nm. In the case of competition between serum and PTmAbOO11 for IgE binding, serum mixtures of biotin-labeled serum and PTmAbOO11 were added to IgE-coated ELISA plates. PTmAbOO11 binding was detected with Streptavidin-HRP (1/1000).

Example 1.5: Human basophil assay

It was determined with human basophils

One is with antibodies, and the other is caused with monoclonal antibodies.

conducted two types of tests (HBA). monoclonal anaphylactogenicity wherein the isolated is used to measure histamine release inhibition

PBMCs add antibodies

Lol PI (strong allergen) by pre-incubation

HBA

Blood was obtained from allergic blood vessels in tubes containing heparin and donors and collected to clean non-erythrocytes. Cells were washed once per second and resuspended in HBH / HSA at a cell density of one milliliter. In the wells, HBH / HSA cells were coated with 2.0 x 10 ⁶ cells of 96-well V-bottom plates containing

100 μΐ of diluted test sample, 100 μΐ of cell suspension or monoclonal antibodies was added. Each test sample was tested at each dilution. The contents in the dilution range with the cells were mixed at 37 ° C for wells for minutes and using a mixer.

For each serum dilution, 10 µΐ HBH / HSA, anaphylactogenicity, was added to the three wells.

mixer and use additional minutes.

for minutes.

were incubated with the wells added

1/1 make up

The contents of the wells were then incubated again at the temperature. Incubation was terminated by centrifugation. The histamine assay was used to determine the EIA of histamine using control wells performed and shaken up to 3 ° C for 500 g with them.

Supernatants were separated to use a commercially available kit (Immunotech). Routine in a test containing cells without a test sample to make a sample to spontaneous and triggered release was determined. Cell samples were lysed by double freezing / thawing and assayed for the total amount of histamine contained in the cells.

The results were expressed as follows:

Anaphylactogenicity test:

histamine release caused by test samples =% histamine release from test sample treated cells -% histamine released spontaneously

Block test:

The degree of inhibition of histamine release can be calculated using the formula:

% inhibition =

1- (histamine released from cells treated with test sample *) (histamine release from antigen-stimulated cells *) x 100

The values are adjusted in case of spontaneous relaxation.

Example 2: Immunization of mice with P14 conjugates (P14-BSA, P14-BSA) induce the production of antibodies against human IgE

The conjugates containing the P14 mimotope (25 µg protein / dose) described in Example 1 were administered to groups of 10 BalbC mice using an oil-water emulsion containing QS21 and 3D-MPL described in WO 95/17210 as adjuvant. Booster dose was performed on days 14, 24 and 72, sera harvested 14 days after each immunization.

Immune responses against the peptide and against IgE bound to IgE were monitored using the ELISA methods described in Example 1. Anaphylactogenicity and functional were then tested in the antiserum and functional. The activity to inhibit the release of histamine from human allergic basophils (methods as described in Example 1).

Results of immunogenicity

Both PD-P14 and BSA-P14 conjugates were able to induce an immune response against P14 and against IgE. Results for anti-peptide and anti-IgE responses induced by BSAP14 conjugates, as measured on day 14 after the third and fourth vaccination, are shown in Table 6. PTmAbOO11 is a monoclonal antibody known to bind the Cs2 region of IgE and it was used to quantify IgE responses and expressed in μς / πτί.

Table 6: Immunogenicity results for BSAP14 conjugates

responses against peptides (14 days after 3) mean titer anti-IgE response (14 days po3) ^ g / ml (PTmAbOO11)) IgE responses (14 days after 4) (μς / πιΐ (PTmAbOOll)) AV SD GM AV SD GM AV SD GM 2 974 22 667 15 492 9, 9 2.18 0.7 22.9 33.5 4.8

notes: AV (mean), SD (mean deviation), GM (geometric mean)

Mice vaccinated with BSA alone as controls do not produce any detectable responses to the peptide or IgE.

Results of functional activity

It has been found that the antiserum resulting from P14 vaccination is functional in that it is effective in inhibiting histamine release from allergic human basophils upon allergen induction (see Figures 2,3 and 4). However, the serum was not found to be anaphylactogenic (shown in Figures 2, 3 and 4).

Summary

P14 (mimotope P8) has been shown to be able to generate high titers of anti-p14 and anti-IgE antibodies in mice. Subsequently, these antibodies have been shown to be functional by inhibiting histamine release from allergic human basophils and are not anaphylactogenic. Therefore, P14 and P8 can be used in the treatment or prophylaxis of allergy.

Example 3: Immunization of mice with Pil conjugates (P11-BSA, P11-BSA) induces production of antibodies against human IgE

The epitope of P1 peptides of human IgE was linked to maleimide-activated BSA (Pierce) (BSA-CRASGKPVNHSTRKEEKQRNGLL). 25 µg of conjugate formed in SBAS2 was injected IM with eight BALB / c female mice on days 0, 14 and 28. One control group of mice was injected with BSA / SBAS2. Blood samples were taken 14 days after each injection (four samples were taken on day 24 after 3 to increase serum availability). Peptide and IgE vaccine-derived antibodies were measured by ELISA as described in Example 1.

Results

A homogeneous anti-Pil IgG response could be detected after the first injection, but increased further after the second and third injections (Figures 5a and 5b). All mice showed an anti-IgE response (which is in the range of 28 to 244 µg / ml as expressed in mAb005 equivalents) after the third injection (Figure 6).

Part 2: Functional activity of epitope-specific monoclonal antibodies

Example 4:

Functional activity of monoclonal antibodies raised against P14

Monoclonal antibodies have been generated that specifically recognize P8 and its mimotopes using techniques known in the art. The P14-BSA conjugate described in Part 1 of these examples was injected into groups of adjuvanted Balb / C mice (w / w) containing QS21 and 3D-MPL. Spleen cells were harvested and fused to the B cell line SP2 / O and the supernatants tested for reactivity against P14 and IgE peptides. Several cell lines, P14 / 23, P14 / 31 and P14 / 33, were deposited under the Budapest Convention at ECACC 26/1/00 with accession numbers 00012610, 00012611, 00012612. It was confirmed that all three monoclonal antibodies bind to IgE. Especially using ELISA binding assays, it was shown that P14 specifically binds to IgE and further used P14 competitive assays against an IgE binding monoclonal antibody.

The functional activity of these monoclonal antibodies was tested by inhibiting histamine release from human basophils as described in Example 1.

Results

All P14 monoclonal antibodies were tested on basophils taken from four different allergic patients (A patients were allergic to mite antigen, G patients were allergic to grass pollen) · PT11 (PTmAbOO11) includes antibodies known to inhibit release as positive control histamine in vitro. All three P14 monoclonal antibodies (23, 31 and 33) are effective in inhibiting histamine release from allergic basophils (shown in Figure 7).

Example 5:

Antibodies against

IgE induced after immunization with conjugate are able to block local allergic response in monkey skin anaphylaxis model

P14 / 23 and P14 / 31 were also tested for in vivo activity. The local skin mast cells of African Green Monkeys were scraped and their sensitivity was increased by intradermal administration of 100 ng of anti-NP IgE (human anti-nitrophenylacetyl (NP) IgE, obtained from Serotech). After 24 hours, the dose range of the monoclonal antibodies was injected into the same injection site as human IgE into one arm. Phosphate buffered saline (PBS) or non-specific human IgE (specific for human cytomegalovirus (CMV.) Or human human immunodeficiency virus (HIV)) was applied to control sites on the opposite arm of the same animals. After 5 hours, 10 mg of BSA-NP conjugate (obtained from Biosearch Laboratories) was injected intravenously. After 15 to 30 minutes, control animals from anaphylaxis developed an easily observable near-circular edema, which can be measured in millimeters. The results are expressed either as mean mean swelling values in a group of three monkeys or as a percentage of inhibition compared to PBS controls. PTmAbOO11 is a monoclonal antibody used as a positive control. SbmAb0006 were used as a negative control.

Table 7: Results P14 / 23

Test quantity · sample (gg) center. rating edema diameter (mm) ' P14 / 23 mAbOOll mAbOOO6 20 May 0 ND 12/15 10 0 0 17/19 1 15/13 0 20/20 0.1 15/12 ND ND 0.05 15/15 ND ND 0 15/15 ND 17/17

9 9 «9 9 9« «« «t t t t t t t t t 9

Table 8: Results P14 / 31

Quantity test, sample (hb) average edema diameter (mm) P14 / 31 mAbOOU mAbOOO6 20 May 0 ND 15/15 10 0 0 15/15 1 22/25 0 20/20 0.1 22/25 ND ND 0.05 25/25 ND ND 0 20/25 ND 20/25

Although inhibition of anaphylaxis by higher doses of monoclonal antibody has been observed, these antibodies are not themselves anaphylactogenic when administered in vivo.

Example 6: Structural aspects of IgE mimotopes

The invention discloses that conformations in which the epitopes or mimotopes of the invention are important in recognizing antibodies against mimotopes as well as in the ability of peptides to produce strong immune responses against IgE. The invention describes developed structural rules that assume optimal peptide cyclization sites. Peptides that use these cyclization sites form a preferred aspect of the invention.

Since the entire IgE Fc structure has not been determined, the available models have been improved (Helm et al., Padlan and Davis, Mol. Immunol., 23, 1063-75, 1986) using the known structure of Cy2 and Cy3 IgG1 (described in Deisenhofer J., 1981, Biochemistry, 20, 23612370). In addition, models of the Cs2 region were generated by comparison with known structures of the Ig packaging unit. The inventors designed these models of IgE Fc homology thereby predicting the final and macroscopic structure of the leaf-space loops (loop AB, Figure 9A) and the loops outside the leaves of the IgE Fc regions (CD loop, shown in Figure 9B). Assumed loops have defined anticipated AB and CD IgE Fc loops with their beta-chain, mimotope-enhancing loops, Φ · ΦΦΦΦ · ΦΦΦΦ · · · · · · · · · · · · · The loops can be obtained from the primary wild-type sequence (WT) of each loop of covalent cyclization between selected specific residues along adjacent beta-chains. The cyclization is preferably released by forming a disulfide bond between the terminal cysteines, which are therefore combined to become cystine.

Based on the structural arrangements (shown in Figures 9A and 9B), easily predicted rules were derived to increase the likelihood that conformations adopted by the mimotope upon conjugation with a suitable carrier molecule are similar to those of the parental epitope.

Rule 1

The hydrophobic cysteine group should replace the WT beta-chain residues that belong to the core of the water-free Ig constant region formed by the space between the two beta-sheets.

Rule 2i

In the case of loops inside the leaf (for example, loop A-B), the cysteine group should replace the WT residues that are

from adjacent anti-parallel beta-chains (shown on 8) and which laterally pack together into the same

hand sheet.

According to rule 1, this occurs within the leaf area. Structural deviation of this rule for loops

A-B is shown schematically in Figures 10A and 10B.

Rule 2ii

In the case of loops outside the leaves (for example, C-D loop), the cysteine moiety should replace the WT residues on the anti-parallel beta-chains, one chain from each leaf. According to Rule 1, the residues forming the optimal package of pairs together with the opposing beta-sheet surfaces thus form part of the space between the leaves. The structural variation of this rule for C-D loops is shown schematically in Figures 11A and 11B. In the tables of putative mimotope sequences, in the text, there are suggestions that seem to •

• a

followed by • 4 •

Further optimally underlined, by each intermittent and uninterrupted sequence block, the splice positions selected for optimal cyclization, which are also shown in the same way in Figure 10 / Loops A-B) and Figure 11B (Loops C-D), are joined.

Under the lines

Using the sequence alignment shown in Figures 9A and 9B together with the above rules, the following list of peptides is shown in Tables 9 and 12. are shown in Figures 1ΌΒ and 11B The underlined sequences are mimotopes.

Table 9: Sequence of A-B Cs3 IgE loop

Peptide sequence (underline, optimal line, and line break) LD LD NO. 341357 CSRPSPFDLFIRKS PTITC CSRPSPFDLFIRKSPTIC CSRPSPFDLFIRKSPTC CSRPSPFDLFIRKSPC CRPSPFDLF1RKSPC CRPSPFDLFIRK3PTC CRPSPFDLFIRKS PT I _C CRPSPFDLFIRKSPTITC CPSPFDLFIRKSPTITC ČPSPFDLFIRKSPTIC CPSPFDLFIRKSPTC CPSPFDLFIRKSPC 33 34 35 36 37 38 39 40 41 42 43 44

· 4 · 4 ·

4 44 • 4 4 4 • 4 44

44 »44444

Table 10: Sequence of Ά-Β Cs4 IgE loop

Peptide sequence (underline, continuous, and interrupt, line are optimal) SQ ID NO. 446 463 CYAFATPEWPGSRDKRTC CYAFATPEWPGSRDKRTC CYAFATPEWPGSRDKRC CAFATPEWPGSRDKRC CAFATPEWPGSRDKRTC CAFATPEWPGSRDKRTLC CAFATP. EWPGSRDKRTLAC CFAT PEWPGSRDKRPRINT C F A T P E W P G S R D K R T L C CFATPEWPGSRDKRTC CFATPEWPGSRDKRC 45 46 47 48 49 50 51 52 53 54 55 56

Table 11: Cs3 IgE loop sequence

peptide sequences (optimal are underlined break, and break, by a line ÍSEQtoKo. 373 387 C TW SRASGKPVNHSTRC 57 CTWSRASGKPVNHSTC 58 CTWSRASGKPVNHSC 59 CTWSRASGKPVNHC 60 CWSRASGKPVNEC 61 CWSRASGKPVNHSC 62 CWSRASGKPVNESTC 63 CWSRASGKPVNESTRC 64 CSRASGKPVNESTRC CSRASGKPVNESTC 66 -CSRASGKPVNHSC 67 CSRASGKPVNEC 68

4 44 4

4 44 44 4 4 4 4

4 · «4

4 4 4 4 ·

4 4 4 4

444 444 44 444

4 · 4 · 44 4 4

Table 12: C-D Cs4 IgE loop mimotope sequence

peptide sequences (optimal are underlined 'break, and break, by a line SEQ ID NO. 477 491 CQWLHNEVQLPDARHSC 69 CQWLHNEVQLPDARHC 70 CQWLHNEVQLP.DARC 71 CQWLHNEVQLPDAC 72 CWLHNEVQLPDAC 73 CWLHNEVQLPDARC 74 CWLHNEVQLPDARHC 75 CWLHNEVQLPDARHSC 76 CLHNEVQLPDARHSC 77 CLHNEVQLPDARHC 78 ČLHNEVQLPDARC CLHNEVQLPDAC / 27 QA ou

Sequence Listing (2) SEQ ID NO: 1 Information:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 22 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 1:

Aru Ai and Se ^ Gly Ls Glu Lys Glu Glu Lys _T 5 '10 15

Gin Arg Asn Gly Thr Leu '20 (2) Information about SEQ ID NO: 2:

(and) SEQUENCE CHARACTERISTICS: (A) LENGTH :. 9 amino acids (B) TYPE: protein (ii) MOLECULES TYPE: human and artificial sequence (Xi) Sequence Description: SEQ ID NO: 2:

Gly Thr Arg Asp Thorn Ile Glu Gly Glu 1 5 (2) Information about SEQ ID NO: 3:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 19 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi)

Sequence Description: SEQ ID NO: 3:

His Pro Leu Pro Arg Ala Leu ⁵

For Arg Ala

Met Arg Ser Thr Thr Lys Thr

Ser Gly (2) Information about SEQ ID NO: 4:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 11 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence

• 9 - • • · ·· ·· • • ·· • • • • · • · • • • · • • • • • • ··· * ·· * · ··

(xi) Sequence description: SEQ ID NO: 4:

Pro Glu Trp Pro Gly Ser

(2) Information about SEQ ID NO: 5: (and) SEQUENCE CHARACTERISTICS: (A) LENGTH: 22 amino acids (B) TYPE: protein (ii) TYPE OF MOLECULES: human and artificial sequence (xi) Sequence Description: SEQ ID NO: 5: Glu Gin Lys Asd Glu 1 5 (2) Information about SEQ ID NO: 6: (and) SEQUENCE CHARACTERISTICS: (A) LENGTH: 19 amino acids (B) TYPE: protein (ii) TYPE OF MOLECULES: human and artificial sequence

(xi) Sequence description: SEQ ID NO: 6:

Leu Se ¹ Arg Pro Ser Pro Phe Asp Leu Phe Ile Arg Lvs Ser Pro Thr I 5 10 15

Ile Thr Cys (2) Information about SEQ ID NO: 7:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 22 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 7:

Tro Leu His Asn Glu Val Gin Le Ser Pro Ser Arg Thr Thr Thr 1 ‘5 10 .15

Gin Pro Arg Lvs Thr '

(2) SEQ ID NO: 8 information:

(and) SEQUENCE CHARACTERISTICS: (AND) LENGTH: 23 amino acids (B) TYPE: protein (ii) SPECIES MOLECULES: human and artificial sequences

• ·

(xi) Description sequence: SEQ ID NO: 8: Cys Arg Ala Ser Gly Lys Pro Val Asn His Ser Thr Arg Glu Glu 1 5 10 15 Dec Lys Gin Arg Asn Gly Leu Leu 20 May

(2) SEQ ID NO: 9 information:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 11 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 9:

Gly Lys Pro Val Asn His Ser Thr Gly Cly '1 5 10 (2) Information about SEQ ID NO: 10:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 18 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 10:

Gly Lvs Gin Glu Lys Gin Arg Asn 1'5 10 15

Gly Cys (2) Information about SEQ ID NO: 11:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 20 amino acids (B) TYPE: protein

(ii) TYPE OF MOLECULES: human and artificial sequences (xi) Sequence description SEQ ID NO: 11 Cys Gly Lvs For Val Asn His Ser Thr Arg Lys Gin Arg 1 5 • 10 15 Dec Asn Glv Leu Leu 20 May

(2) SEQ ID NO: 12 information:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 14 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 12:

Arg Ala Ser Gly Lys For Val Asn His Ser Thr Gly Gly Cys (2) Information about SEQ ID NO: 13:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 11 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 13:

Cys Gly Thr Arg Asp Trp Ile Glu Leu Leu (2) Information about SEQ ID NO: 14:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 22 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 14:

Cys Gly Thr Arg Asp Trp Ile Glu Gly Glu Thr Leu 1 5 10 (2) Information about SEQ ID NO: 15:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 12 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 15:

Gly Glu Thr Arg Asp Trp Ile Glu Glu Glu Thr Glv Cys ¹ 5 10 * (2) Information of SEQ ID NO: 16:

• · · ···:. ·. ::

• · ·· ··· ···· ...

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 12 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 16:

Cys His For His Leu For Arg Ala Leu Met Leu Leu!

. 1 5 10 · (2) Information about SEQ ID NO: 17:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 13 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 17:

Cys Gly Thr His For His Leu For Arg Ala Leu Met

(10) Information about SEQ ID NO: 18:

(iii) SEQUENCE CHARACTERISTICS:

(C) LENGTH: 13 amino acids (D) TYPE: protein (iv) MOLECULAR TYPE: human and artificial sequence (xii) Sequence description: SEQ ID NO: 18:

Thr His For His Leu For Arg Ala Leu Met Arg Ser Cys

(10) Information about SEQ ID NO: 19:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 22 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 19:

Glv Pro His Leu For Arg Ala Leu Met Arg Ser Ser Ser Cys / 5 10 (2) Information about SEQ ID NO: 20:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 13 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 20:

Ala Pro Glu Tro Gly Ser Arg 'Asp Lys Arg Thr Cys! (10) Information about SEQ ID NO: 21:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 17 amino acids (B) TYPE: protein (ii) TYPE OF MOLECULES: human and artificial sequences (xi) Sequence description SEQ ID NO: 21 Ala • 1 For Glu Trp For Gly 5 Ser Arg Lys Arg Thr Leu Ala 10 Gly Gly 15 Dec Cys

(2) Information about SEQ ID NO: 22:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 17 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 22:

Cly Gly Gly Ala Thr Pro Glu Trp Pro Gly Ser Arg Lys Arg Tnr! 5 -10 15

Leu (1). Information about SEQ ID NO: 23:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 13 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 23

Cvs Thr Arg Lys Asd

5 ''10 (2) Information about SEQ ID NO: 24:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 15 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 24:

Ala Pro Cys Trp Pro Gly Ser Asp Cys Arg Thr Leu Ala Gly 1 5 10 ¹⁵ (2) Information about SEQ ID NO: 25:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 17 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 25:

Ala Cys Thr Leu Ala Gly

10 (2) Information about SEQ ID NO: 27:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 16 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 27:

Cys Ala Thr Pro Glu Trp Pro Gly Ser Lys Arg Thr Leu Cys 1 5 10 15

Gly (2) Information about SEQ ID NO: 28:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 13 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 28:

Thr--C ys Pro ys 5 5 5 * 5 10

(2) Information about SEQ ID NO: 29: (and) SEQUENCE CHARACTERISTICS: (A) LENGTH: 11 amino acids (B) TYPE: protein (ii) MOLECULES TYPE: human and artificial sequence (Xi) Sequence Description: SEQ ID NO: 29: Cys Gly Ala Glu Tro Glu Gin Lys Asp Glu Leu 1 5 * 10

(2) Information about SEQ ID NO: 30:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 11 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 30:

Ala Glu TrD Glu Gin Lys Asp Glu Phe Ile Cys 1 ‘5 10 (2) SEQ ID NO: 31 information: - (i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 9 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 31:

Gly Glu Gs Lvs Asd Glu Phe Ile Cys 1 '5 (2) Information about SEQ ID NO: 32:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 10 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 32:

Cys Ala Glu Gly

Cu Gln Lys Asp Glu Leu; 10 (2) Information about SEQ ID NO: 33:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 19 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 33:

Cys Ser Arg Pro Ser Phe Asp Leu Phe Ile Arg Lys Ser Pro Thr; ⁵ 10 15

Ile Thr Cys

SEQ

ID NO:

34:

(and)

CHARACTERISTICS

SEQUENCE:

(A) LENGTH: 18 amino acids ein (B) TYPE: prot · (ii) TYPE OF MOLECULES: human and artificial sequence (xi) Sequence description SEQ ID NO: 34 Cys 1 Ile Ser Pro Pro Ser Pro 5 Cys Phe A.sp Leu Phe: 10 ile Arg Lys SEQ ID NO Information: 35: (and) SEQUENCE CHARACTERISTICS: (A) LENGTH: 17 amino acids (B) TYPE: protein (ii) TYPE OF MOLECULES: human and artificial sequence (xi) Sequence description SEQ ID NO: 35 Cys Ser Pro Pro Ser Pro Phe Asp Leu Phe Ile Arg Lys 1 5 10

Cys

Se

Pro Thr

Ser Pro Thr (2) Information about SEQ ID NO: 36:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 16 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 36:

Cvs Ser Arg Pro Ser Pro Phe Asp Leu Phe Ile Arg Lys Ser Pro Cys' 5 10 15 (2) SEQ ID NO: 37 information:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 15 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 37:

Cvs Arg Pro Ser Pro Phe Asd Leu Phe Ile Arg Lys Se ^{1 Dt} Cvs 5 10 '*' 15 (2) SEQ ID NO: 38:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 16 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 38:

Arg Pro Ser Pro Phe Asp Leu Phe Ile Arg Lvs Ser Pro Thr Cys 5 10 15 (2) SEQ ID NO: 39 Information:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 17 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 39:

Cys Arg Pro Ser Pro Phe Asp Leu Phe Ile Arg Lys.Ser Pro Thr Ile 1 5 10 15

Cys (2) Information about SEQ ID NO: 40:

(i) SEQUENCE CHARACTERISTICS:

Pro Thr Ile (2) (2) (A) (B)

SPECIES

LENGTH: 18 amino acids

TYPE: protein

MOLECULES: human and artificial sequences

Sequence Description: SEQ ID NO: 40:

Cys

Thr

Arg Pro Ser Pro Phe Asp

Cys

Information about

SEQ ID NO: 41:

SEQUENCE CHARACTERISTICS:

(A) (B)

SPECIES

LENGTH: 17 amino acids

TYPE: protein

MOLECULES: human and artificial sequences

Information about SEQ ID NO: 42:

(and)

Cys

SEQUENCE CHARACTERISTICS:

(AND)

SPECIES

LENGTH: 16 amino acids

TYPE: protein

MOLECULES: human a

Sequence description: SEQ ID

P-O Ser Pro Phe Asp Leu Phe

Information about SEQ ID NO: 43:

artificial sequence

NO: 42:

Ile Arg Lys Ser Pro

SEQUENCE CHARACTERISTICS:

(AND)

LENGTH: 15 amino acids

TYPE: protein

SPECIES

MOLECULES: human and artificial sequences

Sequence Description: SEQ ID NO: 43:

Cys

Information about SEQ ID NO: 44:

SEQUENCE CHARACTERISTICS:

(A) LENGTH: 14 amino acids • 4 * · · (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 44:

Cys Pro Ser Pro Phe Asp Leu Phe Ile Arg Lvs Ser Pro Cys 1 5 10 (2) SEQ ID NO: 45 info:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 20 amino acids * (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 45:

Cys Tyr Ala Phe Ala Thr Pro Glu Trp Pro Gly Ser Aso Lys Ar ^{1 5} 10 · '-i ~

Thr Leu Ala Cys ⁴

Information on SEQ ID NO: 46: (and) SEQUENCE CHARACTERISTICS: (AND) LENGTH: 19 amino acids (B) TYPE: protein (ii) SPECIES MOLECULES: human and artificial sequences

(xi) Sequence description: SEQ ID NO: 46:

Lys Arg

1 5 10 15 Dec Thr Leu Cys

(2) SEQ ID NO: 47 information:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 18 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 47:

Lys Arg

Thr Cys

(2) Information on SEQ ID NO: 48 (and) SEQUENCE CHARACTERISTICS: (AND) LENGTH: 17 amino acids (B) TYPE: protein (ii) SPECIES MOLECULES: human and artificial sequences

(xi) Sequence description: SEQ ID NO: 48:

Cys Tyr Ala Phe Ala Thr Pro Glu Trp Pro Gv Ser Arg Lys Arg 1 5 10 15

Cys

> (2) Information about SEQ ID NO: 49: * (and) SEQUENCE CHARACTERISTICS: (A) LENGTH: 16 amino acids (B) TYPE: protein (ii) SPECIES MOLECULES: human and artificial sequences

(xi) Sequence description: SEQ ID NO: 49:

Cys Ma'rhe Ma Thr Pro Glu Trp Prc Gly Ser Arg Lys Arg Cys' 1 5 ¹⁰ (2) SEQ ID NO: 50 info:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 18 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 50:

Cys Ala Phe Ala Thr Pro Glu Trp Pro Gly Ser Aso Lys Arg Th Cys ^{10 15} (2) SEQ ID NO: 51 info:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 18 amino acids (B) TYPE: protein (ii)) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 51:

Cys Ala Phe Ala Thr Pro Glu Trp Pro Gly Ser

Leu Cys

(2) Information about SEQ ID NO: 52:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 19 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 52:

Cys Ala Phe Ala Thr For Glu Trp For Gly Ser Asp Asp Lys Arg Thr

1.5 10, 15

Leu Ala Cys (2) Information about SEQ ID NO: 53:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 18 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 53:

Cys Phe Ala Thr Pro Glu -Trp Pro Gly Ser Asd Lys Arg Thr Leu ¹ 5 · 10 '15

Ala Cys (2) Information about SEQ ID NO: 54:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 17 amino acids (B) TYPE: protein (ii) MOLECULES TYPE: human and artificial sequence (xi) Sequence Description: SEQ ID NO: 54:

Cys Phe Ala Thr For Glu Trp For Gly Ser L Asp Arg Thr Leu 1 · ⁵

Cys (2) Information about SEQ ID NO: 55:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 16 amino acids

(B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 55:

Cvs Phe Ala Thr Pro Glu Trp Pro Gly Ser Arg Asn Lys Arg Thr Cys 1'5 10 15 (2) SEQ ID NO: 56:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 15 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 56:

Cys Phe Ala Thr Pro Glu Trp Pro Gly Ser Arg Asd Lys Arg Cys ¹ , ⁵ 10 '15 (2) Information about SEQ ID NO: 57:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 17 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 57:

Cys Thr Trp Ser Ser Ala Ser Gly Lys Pro Val Asn His Ser Th- Arg ¹ 5 10 15

Cys (2) Information of SEQ ID NO: 58:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 16 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 58:

Cys Thr Trp Ser Arg Lys Pro Val Asn His Ser Thr Cys 1 5 -10 15.

(2) Information about SEQ ID NO: 59:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 15 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 59:

Cvs Th ”Trp Ser Arg Ala Ser Gly Lys For Val Asn His Ser Cys 1 * 5 10 ¹⁵ (2) Information about SEQ ID NO: 60:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 14 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 60:

Cys 1r p10 (2) Information about SEQ ID NO: 61:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 14 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 61:

Cvs ^T ro Ser Arg Ala Ser Gly Lys Off Val Asn His Cys' 1 5 10 (2) Information about SEQ ID NO: 62:

and) SEQUENCE CHARACTERISTICS: (AND) LENGTH: 14 amino acids (B) TYPE: protein (ii) SPECIES MOLECULES: human and artificial sequences

(xi) Sequence description: SEQ ID NO: 62:

Cys Trp Ser Arg Ala Ser Glv Lys For Val Asn His Ser Cvs 1 5 10 (2) Information of SEQ ID NO: 63:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 15 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 63:

His Ser Thr Cys 1 * 5 10 15 (2) Information about SEQ ID NO: 64:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 16 amino acids (B) TYPE: protein

(ii) TYPE OF MOLECULES: human and artificial sequences (xi) Sequence description SEQ ID NO: 64 Cys Trp Ser Arg Ala Ser Gly Lys Pro Val Asn His Ser Thr Arg Cys 1 5 10 15 Dec

(2) Information about SEQ ID NO: 65:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 15 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 65:

Cys Ser Arg Ala Ser Gly Lvs For Val Asn His Ser Thr Arg Cys (2) SEQ ID NO: 66 info:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 14 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 66:

• ·· ♦ · · ······· Φ

Lys For War Asn His Ser Thr Cys

♦ · ·· • • ·· ♦ · • · • · · · • • · · · · • • · · · • ·· ··· ·· ··

-Cys Ser Arg Ala Ser Gly

5:

SEQ ID NO Information:

(and) SEQUENCE CHARACTERISTICS: (A) LENGTH: 13 amino acids (B) TYPE: protein (ii) MOLECULES TYPE: human and artificial sequence ‘(Xi) Sequence Description: SEQ ID NO: 67:

Cys

Ser Arg Ala Ser Gly's Lys Pro

Val Asn His Ser Cys (2) Information about SEQ ID NO: 68:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 12 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 68:

Cys Ser Arg Ala Ser Gly Lys For Val Asn His Cys ⁵ (2) Information about SEQ ID NO: 69:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 17 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 69:

Cys Gin TrD Leu His. As Glu Val Gin Leu Asp Asp Arg His Ser 1'5 10 '15

Cys (2) Information about SEQ ID NO: 70:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 16 amino acids (B) TYPE: protein • (ii)

MOLECULES TYPE: human and artificial sequence

Leu Pro Asp Arg Arg His Cvs

- - -

Sequence description:

SEQ ID NO: 70:

Cys Gin Trp Leu His

5

Asn Glu Val Gin

71:

SEQ ID NO Information:

(2)

(and) SEQUENCE CHARACTERISTICS: (A) LENGTH: 15 amino acids l (B) TYPE: protein (ii) TYPE OF MOLECULES: human and artificial (xi) Sequence description: SEQ ID NO: 71

sequence

Cys Gin Trp Leu His Asn Glu Val Gin Leu For Asp Ala Arg Cys 1 5 10 15 (2) SEQ ID NO: 72 info:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 14 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 72:

Cys Gin Thorn Leu His Asn Glu Val Gin Leu For Asd Ala Cys '5 10' (2) SEQ ID NO: 73:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 13 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 73:

Cys Trp Leu His Asn Glu Val Gin Leu For Asp Ala Cys 1 5 1 ° (2) Information about SEQ ID NO: 74:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 14 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 74:

Cys Trp Leu His Gn Val Glr. Leu Pro Asd Ala Aro Cvs ¹ 5 10 (• 2) Information about SEQ ID NO: 75:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 15 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 75:

Cys Trp Leu His Asn Glu Val Gin Leu For Asd Ala Arg H'S Cv = · ^{1 5} 10 15 (2) Information about SEQ ID NO: 76:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 16 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 76:

Cys i

Trp Leu His Asn Glu

For Asd Ala

Arg

His

Ser Cvs (2) Information about SEQ ID NO: 77:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 15 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 77:

Cys Leu His Gn Val Glr. Leu Pro Asp Ala .Arg His Ser Cys

(2) Information about SEQ ID NO: 78:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 14 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 78:

Cys Leu His Asn Glu Val Gin Leu For Asp Ala Arg His Cys 1 5 10 ´ (2) SEQ ID NO: 79 info:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 13 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 79:

Cvs Leu His Asn Glu Val Gin Leu For Asp Ala Arg Cys 1. (10) Information about SEQ ID NO: 80:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 12 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 80:

Cys Leu His Asn Glu Val Gin Leu For Asp Ala Cys 1 5 10 (2) SEQ ID NO: 81 info:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 6 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 81:

Leu Phe Ile Arg

5 (2) Information about SEQ ID NO: 82:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 7 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 82:

L'-ο Ser Lys Gly Thr Val Asn ' ⁵ (2) Information about SEQ ID NO: 83:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 23 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Sequence description: SEQ ID NO: 83:

Leu His Asn Glu Val Gin Leu For Asp Ala Arg His Ser Thr Thr Gin i_ 5 1θ l · ¹

Lys Thr Lvs Gly Ser (2) Information about SEQ ID NO: 84:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 6 amino acids (B) TYPE: protein (ii) MOLECULAR TYPE: human and artificial sequence (xi) Description of sequence: SEQ ID NO: 84:

Ser Val Asn For Gly Lvs

5 (2) Information about SEQ ID NO: 85:

(i) SEQUENCE CHARACTERISTICS:

(A) LENGTH: 13 amino acids ·· · · · · · ♦ ··· ·

φ.

(4) (B) TYPE: protein

(ii) TYPE OF MOLECULES: human and artificial sequence (xi) Sequence description SEQ ID NO: 85 Cys Pro Glu Trp For Gly Cys Aurg Asp Lys A.rg and 5 10 SEQ ID NO Information: 86: (and) SEQUENCE CHARACTERISTICS: (A) LENGTH: 13 amino acids (B) TYPE: protein (ii) TYPE OF MOLECULES: human and artificial sequence (xi) Sequence description SEQ ID NO: 86

Thr

Thr Pro Glu Trc

Pro Glv Cys Ατα ksp Lys Arg Cvs Gly

Claims (3)

A peptide comprising an isolated surface-exposed epitope region that overlaps the 0ε3 and Ce4 region of IgE, wherein the peptide is P7 (SEQ ID NO: 3) or a mimotope thereof. A peptide comprising an isolated surface-exposed epitope of the Ce4 IgE region, wherein the peptide is P8 (SEQ ID NO: 4) or a mimotope thereof. A peptide comprising an isolated surface-exposed epitope of the Cs4 IgE region, wherein the peptide is P9 (SEQ ID NO: 5) or a mimotope thereof. 4. A peptide comprising an isolated surface exposed epitope of the 0ε4 IgE region, wherein the peptide is 4-90N (SEQ ID NO: 84) or a mimotope thereof. Peptide according to Claim 2 where mimotope P8 Yippee general peptide formulas P, X _x , X _2, P X3r X ₄ , X _5, X _6, X _5, X _5, where the symbol X _x Yippee amino acid selected z E, D, N or Q, symbol X ₂ Yippee amino acid selected z W , Y or F, symbol X ₃ Yippee amino acid selected of G or A, symbol X ₄ Yippee amino acid selected z S , T or M, symbol X ₅ Yippee amino acid selected of ; R or K and symbol Xs Yippee amino acid selected from D or E. Peptide according to Claim 5 where mimotope P8 Yippee general peptide of formula P, Xj., _X2, P, G, X ₄ , R <D, X _5, X _5, where the symbol X _x Yippee amino acid selected z E, D, N or Q, symbol X ₂ Yippee amino acid selected z W , Y or F, symbol X ₄ Yippee an amino acid selected from S, an amino acid selected from R an amino acid selected from D or • ·· 44 · 4 • 4 44 44 44 4 • 4 44 4 4 • · * 4 4 • 4 4 4 4 4 4 4 4 ··· «··« 4 4 444 444 4 4 « 4 · «· T or M, symbol X ₅ is or K a symbol X ₆ is E. The mimotope of any one of claims 1 to 6, wherein the mimotope is a peptide. An immunogen for treating allergy comprising a peptide or mimotope according to any one of claims 1 to 7 and additionally comprising a carrier molecule. The immunogen of claim 8, wherein the carrier molecule is selected from protein D or hepatitis B core antigen. The immunogen of claim 8 or 9, which is a chemical conjugate of a peptide or mimotope or is expressed as a fusion protein. The immunogen of any one of claims 8 to 10, wherein the peptide or mimotope of the peptide is present in the primary carrier sequence. A vaccine composition for treating allergy comprising the peptide or immunogen of any one of claims 1 to 11 and further comprising an adjuvant. 13. A vaccine composition for the treatment of allergy comprising a peptide comprising an isolated surface exposed epitope of the Cs3 region of the IgE region, wherein the peptide is P5 (SEQ ID NO: 1) or a mimotope thereof, and an adjuvant. 14. An allergy vaccine composition comprising isolated The surface exposed epitope 0ε3 of the IgE region, where the peptide is P6 (SEQ ID NO: 1). 2) or a mimotope thereof, and an adjuvant. 15. A vaccine composition for the treatment of allergy comprising a peptide comprising an isolated surface exposed epitope of the Cs3 IgE region, wherein the peptide is P200 (SEQ ID NO: 6) or a mimotope thereof, and an adjuvant. 16. A vaccine composition for treating allergy comprising a peptide comprising an isolated surface-exposed epitope of the Ce3 IgE region, wherein the peptide is P210 (SEQ ID NO: 7) or a mimotope thereof, and an adjuvant. 17. A vaccine composition for the treatment of allergy, comprising a peptide comprising an isolated surface-exposed epitope of the Cs3 IgE region, wherein the peptide is 2-90N (SEQ ID NO: 82) or mimotope thereof, and an adjuvant. 18. A vaccine composition for the treatment of allergy comprising a peptide comprising an isolated surface-exposed epitope of the Cs4 IgE region, wherein the peptide is 3-90N (SEQ ID NO: 83) or a mimotope thereof, and an adjuvant. 19. The vaccine composition of claim 1 any of that by immunogen. claims 13 the peptide is to 18, spoj en with a molecule carriers to form 20. Vaccine means according to claim 19, marked u j i c i se that molecule carriers The core of the immunogen was selected from protein D or hepatitis antigen. B. The vaccine composition of claim 19 or 20, wherein the immunogen is a chemical conjugate of a peptide or mimotope, or wherein the immunogen is expressed as a fusion protein. 22. The vaccine composition of claims 19-21, wherein the peptide or mimotope of the peptides is present in the primary carrier sequence. 23. Ligand, who is capable 1 to 6. recognize peptides according to any of claims 24. Ligand according to claim 23, which chose from P14 / 23, P14 / 31 or P14 / 33, j enž saved according to Budapest Convention in the institution ECACC on 2 6.1 .: 2000 with access control number 00012610, 00012611, 00012612. 25. Pharmaceutical means, ..... .mu.Ci by comprising a ligand according to claim 23. 26. Pharmaceutical means, ..... .mu.Ci comprising the ligand of claim 24. A peptide according to any one of claims 1 to 6 suitable for use in medicine. A vaccine composition according to any one of claims 12 to 22, characterized in that it is suitable for use in medicine. · · · · · · · · · · · · · · · · · · · · · · · · · · · · · Immunogen according to any one of claims 8 to 11, suitable for use in medicine. Use of a peptide according to any one of claims 1 to 6 in the manufacture of a medicament for the treatment or prevention of allergy. A ligand that is capable of recognizing the peptide of any one of claims 1 to 6 and is suitable for use in medicine. Use of a ligand capable of recognizing the peptide of any one of claims 1 to 6 in the manufacture of a medicament for the treatment of allergy. Use of P14 / 23, P14 / 31 or P14 / 33 which have been deposited according to Budapest Convention at ECACC on 26.1.2000 with accession number 00012610, 00012611, 00012612, in identification of mimotope P8. 34. Peptide capable of being recognized by P14 / 23 or P14 / 33 and deposited according to Budapest at ECACC on 26.1.2000 with access P14 / 31 Convention number 00012610, 00012611, 00012612. 35. A vaccine composition comprising the peptide of claim 34. 36. A method of making a vaccine composition comprising producing an immunogen according to any one of claims 8 to 11 and preparing an immunogen with an adjuvant. 37. A method of treating a patient suffering from or prone to allergy, comprising: Administering to a patient a vaccine composition according to any one of claims 12 to 22. A method of treating a patient suffering from or susceptible to allergy, comprising administering to a patient a pharmaceutical composition according to any one of claims 25 or 26. 39. Use of any of the peptides of SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 82, 83 and 84 in the manufacture of a vaccine for the treatment of allergy.