IL-23-P19 VACCINES

Abstract

Disclosed is a vaccine, preferably for use in the prevention or treatment of an interleukin 23 (IL-23) related disease, comprising a peptide bound to a pharmaceutically acceptable carrier, wherein said peptide is selected from the group QPEGHH-WETQQIPSLS (SEQ ID No. 103; p8322), GHHWETQQIPSLSPSQPWQRL QPEGHHWETQ (SEQ ID No. 98; p8461), TQQIPSLSPSQ (SEQ ID No. 99; p8400), QPEGHHWETQQIPSLSPSQ (SEQ ID No. 100; p9269), QPEGHHWETQQIPSLSPS (SEQ ID No. 101; p9269-C1), and QPEGHHWETQQIPSLSP (SEQ ID No. 102; p9269-C2), especially QPEGHHWETQQIPSLS (SEQ ID No. 103; p8322) and wherein said IL-23 related disease is selected from the group psoriasis, psoriatic arthritis, rheumatoid arthritis, systemic lupus erythematosus, diabetes, preferably type 1 diabetes, atherosclerosis, inflammatory bowel disease (IBD)/M. Crohn, multiple sclerosis, Behcet disease, ankylosing spondylitis, Vogt-Koyanagi-Harada disease, chronic granulomatous disease, hidratenitis suppurtiva, anti-neutrophil cytoplasmic antibodies (ANCA-) associated vasculitides, neurodegenerative diseases, preferably M. Alzheimer or multiple sclerosis, atopic dermatitis, graft-versus-host disease, cancer, preferably Oesophagal carcinoma, colorectal carcinoma, lung adenocarcinoma, small cell carcinoma, and squamous cell carcinoma of the oral cavity, especially psoriasis, neurodegenerative diseases or IBD.

Claims

1: A composition, comprising: the peptide according to claim 14, bound to a pharmaceutically acceptable carrier.

2: The composition according to claim 1, wherein at least one cysteine residue is bound to an N- or C-terminus of the peptide.

3: The composition according to claim 1, wherein at least one cysteine residue is bound to an N-terminus of the peptide.

4: The composition according to claim 1, wherein the pharmaceutically acceptable carrier is a protein carrier.

5: The composition according to claim 4, wherein the protein carrier is selected from the group consisting of keyhole limpet haemocyanin, tetanus toxoid and diphtheria toxin.

6: The composition according to claim 1, wherein the composition is formulated with an adjuvant.

7: The method according to claim 17, the administering comprises intravenous, subcutaneous, intradermal or intramuscular administration.

8: The composition according to claim 1, wherein the peptide is contained in the composition in an amount from 0.1 ng to 10 mg.

9: The composition according to claim 1, wherein the peptide is bound to the pharmaceutically acceptable carrier by a linker.

10: The composition according to claim 9, wherein the peptide is bound to the pharmaceutically acceptable carrier by a peptide linker and the peptide linker is selected from the group consisting of Gly-Gly-Cys, Gly-Cys, Cys-Gly and Cys-Gly-Gly.

11: The composition according to claim 1, wherein the composition is a biepitopic vaccine.

12: The composition according to claim 1, wherein the composition comprises two peptides, each separately bound to a pharmaceutically acceptable carrier.

13: A vaccine kit, comprising: the composition of claim 1 and a further composition against a disease selected from the group consisting of psoriasis, psoriatic arthritis, rheumatoid arthritis, systemic lupus erythematosus, diabetes, especially type 1 diabetes; atherosclerosis, inflammatory bowel disease (IBD)/M. Crohn, multiple sclerosis, Behcet disease, ankylosing spondylitis, Vogt-Koyanagi-Harada disease, chronic granulomatous disease, hidratenitis suppurtiva, anti-neutrophil cytoplasmic antibodies (ANCA-) associated vasculitides, neurodegenerative diseases, atopic dermatitis, graft-versus-host disease, and cancer.

14: A peptide, selected from the group consisting of QPEGHHWETQ (SEQ ID No. 98; p8461), TQQIPSLSPSQ (SEQ ID No. 99; p8400), QPEGHHWETQQIPSLSPSQ (SEQ ID No. 100; p9269), QPEGHHWETQQIPSLSPS (SEQ ID No. 101; p9440), QPEGHHWETQQIPSLSP (SEQ ID No. 102; p9441), QPEGHHWETQQIPSLS (SEQ ID No. 103; p8322), QPEGHHWETQQIPS (SEQ ID No. 104; p8495), QPEGHHWETQQI P (SEQ ID No. 105; p8459-1), QPEGHHWETQQI (SEQ ID No. 106; p8460), QPEGHHWETQQ (SEQ ID No. 107; p8460-1), QPEGHHWET (SEQ ID No. 108, p8461-1), QPEGHHWE (SEQ ID No. 109; p8462), TQQIPSLSPSQPWQ (SEQ ID No. 110, p8397), TQQIPSLSPSQPW (SEQ ID No. 111, p8398), TQQIPSLSPSQP (SEQ ID No. 112, p8399), TQQIPSLSPS (SEQ ID No. 113; p8761), TQQIPSLSP (SEQ ID No. 114; p8762), and TQQIPSLS (SEQ ID No 115; p8763).

15: A peptide pair, comprising: the peptide of claim 14, wherein a first peptide of the peptide pair is selected from the group consisting of QPEGHHWETQQIPS (SEQ ID No. 104; p8495), QPEGHHWETQQI P (SEQ ID No. 105; p8459-1), QPEGHHWETQQI (SEQ ID No. 106; p8460), QPEGHHWETQQ (SEQ ID No. 107; p8460-1), QPEGHHWETQ (SEQ ID No. 98; p8461), QPEGHHWET (SEQ ID No. 108, p8461-1) and QPEGHHWE (SEQ ID No. 109; p8462) and a second peptide of the peptide pair is selected from the group consisting of TQQIPSLSPSQPWQ (SEQ ID No. 110, p8397), TQQIPSLSPSQPW (SEQ ID No. 111, p8398), TQQIPSLSPSQP (SEQ ID No. 112, p8399), TQQIPSLSPSQ (SEQ ID No. 99; p8400), TQQIPSLSPS (SEQ ID No. 113; p8761), TQQIPSLSP (SEQ ID No. 114; p8762) and TQQIPSLS (SEQ ID No 115; p8763).

16: The composition according to claim 1, wherein the peptide is selected from the group consisting of QPEGHHWETQQIPSLS (SEQ ID No. 103; p8322), QPEGHHWETQ (SEQ ID No. 98; p8461), TQQIPSLSPSQ (SEQ ID No. 99; p8400), QPEGHHWETQQIPSLSPSQ (SEQ ID No. 100; p9269), QPEGHHWETQQIPSLSPS (SEQ ID No. 101; p9440), and QPEGHHWETQQIPSLSP (SEQ ID No. 102; p9441).

17: A method of treating an interleukin 23 related disease, comprising: administering the composition of claim 1 to a subject in need thereof, wherein the interleukin 23 related disease is selected from the group consisting of psoriasis, psoriatic arthritis, rheumatoid arthritis, systemic lupus erythematosus, diabetes, especially type 1 diabetes; atherosclerosis, inflammatory bowel disease/M. Crohn, multiple sclerosis, Behcet disease, ankylosing spondylitis, Vogt-Koyanagi-Harada disease, chronic granulomatous disease, hidratenitis suppurtiva, anti-neutrophil cytoplasmic antibodies associated vasculitides, neurodegenerative diseases, atopic dermatitis, graft-versus-host disease, and cancer.

18: The composition according to claim 11, wherein the biepitopic vaccine comprises a peptide selected from the group consisting of QPEGHHWETQQIPSLSPSQ (SEQ ID No. 100; p9269), QPEGHHWETQQIPSLSPS (SEQ ID No. 101; p9440), QPEGHHWETQQIPSLSP (SEQ ID No. 102; p9441), and QPEGHHWETQQI PSLS (SEQ ID No. 103; p8322).

19: The composition according to claim 12, wherein a first peptide is selected from the group consisting of QPEGHHWETQQI PS (SEQ ID No. 104; p8495), QPEGHHWETQQI P (SEQ ID No. 105; p8459-1), QPEGHHWETQQI (SEQ ID No. 106; p8460), QPEGHHWETQQ (SEQ ID No. 107; p8460-1), QPEGHHWETQ (SEQ ID No. 98; p8461), QPEGHHWET (SEQ ID No. 108, p8461-1) and QPEGHHWE (SEQ ID No. 109; p8462) and a second peptide is selected from the group consisting of TQQIPSLSPSQPWQ (SEQ ID No. 110, p8397), TQQIPSLSPSQPW (SEQ ID No. 111, p8398), TQQIPSLSPSQP (SEQ ID No. 112, p8399), TQQIPSLSPSQ (SEQ ID No. 99; p8400), TQQIPSLSPS (SEQ ID No. 113; p8761), TQQIPSLSP (SEQ ID No. 114; p8762) and TQQIPSLS (SEQ ID No 115; p8763).

Description

[0052] FIG. 1: Inhibition of IL-23 function by sera induced with peptides from p19 as measured with STAT3-assay (A) and splenocyte assay (B) and p40, as measured with STAT3-assay (C): Peptides from p19 and p40 (grey columns) elicit murine sera that inhibit IL-23 function. Serum induced with irrelevant peptide (p4994, black column) was used as negative control. Peptides used for the elicitation of immune sera are denoted on the abscissa, below are the domains in which the respective peptides are situated. Pooled sera from five animals each have been used. The same sera were used for both assays. (D): Lack of inhibition of IL-23 function by sera induced by other p19-derived peptides as measured in the STAT3-assay. Ineffective p19-peptide-induced sera (grey columns) are compared with p6063 serum (white column). Pooled sera from five animals each have been used.

[0053] FIG. 2: The p6063-region contains two discrete epitopes. Binding of p6063-specific serum antibodies to a series of overlapping, glass-attached 12-mer peptides on a microarray, sliding by one amino acid, was measured after detection with fluorescence-labelled secondary antibody in a fluorescence-reader. A: Position of the peptides relative to the p19-sequence (upper and lower margin, the sequence of p6063 is printed in bold letters). Data are oD680 and correspond to the amount of antibody bound by a given peptide, darker shades indicate higher values. B: Each of the serial peptides overlaps at least partly with p6063 and is represented by a column. Data are expressed as percentage relative to the strongest binder. The affiliation of a peptide to a certain epitope is indicated by the colour of the column, the sequences of the individual peptides detected by the serum are displayed on the abscissa.

[0054] FIG. 3: Competition of the IL-23-inhibitory power of p6063 specific-serum as measured in the splenocyte assay. Peptides were added to the splenocyte assay to compete with IL-23 for binding of serum-antibodies. Open columns: Function controls: stimulation of IL-17-production by IL-23 and anti-p6063-serum-caused inhibition thereof. Black columns: Negative controls with irrelevant sera. Light grey columns: Competition of anti-p6063-serum with single peptides in the indicated concentrations. Dark grey: Competition of anti-p6063-serum with combined peptides in the indicated concentrations. The N-terminal competitor was p8464 (p19.sub.140-147) and the C-terminal p7434 (p19.sub.144-158). Data are given as percentage of IL-17 expression as compared to splenocytes stimulated in the presence of irrelevant serum (p4994, derived from human C5a). The irrelevant serum was generated with this same peptide. p6063 serum was taken from one single animal with a strong immune response. Control sera are pools from five animals. Serum concentration was 2.5%.

[0055] FIG. 4: Search for the minimal immunologically relevant sequences. Truncations were performed in the N-terminal (A) and the C-terminal epitope (B) of domain 3. The panels on the left illustrate the position of the peptides in relation to the sequence of domain 3 (bold letters: p6063); the panels on the right illustrate functional inhibition of IL-17 expression by the respective sera. IL-17A expression as measured by splenocyte assay: cells stimulated with 1 ng/ml IL-23 in the presence of sera raised against the denoted peptides. Data are given as percentage of IL-17 expression as compared to splenocytes stimulated in the presence of irrelevant serum (p4994, black columns). Serum pools from 5 animals are used.

[0056] FIG. 5: Biepitopic and bivalent vaccines. A: Comparison of serum elicited against the biepitopic peptide p9269 with sera against the monoepitopic peptides p8461 and p8400. IL-17A expression as measured by splenocyte assay: cells stimulated with 1 ng/ml IL-23 in the presence of sera raised against the denoted peptides. Data are given as percentage of IL-17 expression as compared to splenocytes stimulated in the presence of irrelevant serum (p4994, black columns) B: Potential proteasomal cleavage sites in p9269. The size of the black bars between letters indicates the probability of cleavage. The area shaded in gray highlights a sequence predicted to be a strong MHC I-binder (See Tab 2). C: Homologies of p9269-derived hexapeptides with unrelated proteins. The size of the bars correlates to the number as indicated on the y-axis of proteins with homology to a given hexamer, indicated on the x-axis. D: Comparison of IL-23 inhibitory capacity of p9269 (left pair of columns), p8322 (middle pair of columns) and p8461 concomitantly injected with p8400 (right pair of columns)triggered sera. Results are compiled from multiple independent experiments and shown as mean valuesS.E.M. Details as in (A). E: Potential proteasomal cleavage sites in p8322. Details as in (B). F: Homologies of p8322-derived hexapeptides with unrelated proteins. Details as in (C).

[0057] FIG. 6: Bivalent vaccines. A-D: Potential epitopes with SYFPEITHI-cores >0 within peptides p9269 (p19.sub.136-154) (A), p8322 (p19.sub.136-151) (B), p8461 (p19.sub.136-145) (C) and p8400 (p19.sub.144-154) (D). Numerals on the abscissae represent the SYFPEITHI-score of a given peptide, those on the ordinates the number of peptides. Column size represents the number of peptides with the respective SYFPEITHI-Score. E, F: Bivalent vaccinations with peptides from different regions of the same subunit (E) or different subunits (F). 30 g of each single peptide were injected individually (white columns). When peptides were concomitantly injected (grey column), 15 g of each peptide was used. Data are given as relative STAT3-phosphorylation as compared to STAT3-phosphorylation of T cells stimulated in the presence of irrelevant serum (anti p4994-serum, black column). Data represent mean of two independent experiments S.E.M. Sera are pools of five animals each. Both peptides were coupled to KLH.

[0058] FIG. 7: Comparison of the immunologic response against IL-23 elicited by p6063 or peptides containing the minimal epitopesp8400 and p8461(grey columns) to the responses elicited by peptides derived from sequences mentioned in other patents (open columns) (A). Data are given as relative IL-17 expression as compared to IL-17 expression by splenocytes stimulated in the presence of p6063-serum. Irrelevant serum (anti p4994-serum, black column) was included for control purposes. Data represent mean of two independent experiments S.E.M. Sera are pools of five animals each. B depicts the position of the peptides relative to the sequence of IL-23p19. The sequence covered by p6063 is shown in bold letters. Grey bars: p6063 and peptides covering minimal epitopes, white bars: peptides from sequences described in foreign patents.

EXAMPLES

Materials and Methods

Mice

[0059] BALB/cj and C57BL/6j mice were purchased from Charles River (Sulzfeld, Germany) or Janvier (St. Berthevin, France)

[0060] All animal testing was performed in accordance with actual Austrian national law (Tierversuchsgesetz 2012-TVG 2012) and with consent of the relevant authorities.

Peptides

[0061] Peptides were purchased from EMC Microcollections GmbH (Tubingen, Germany). All peptides are derived from IL-23 sequences, with the exception of the irrelevant peptide p4994 which is derived from human C5a and bears no relevant similarity to either subunit of IL-23.

Sequences

[0062] GenBank sequences AAH66268.1 (for p19) and AAD56386.1 (for p40) were chosen as templates for the sequences and numeration of the vaccination peptides. They represent the complete sequences of the cytokine subunits including the putative leader sequences. These sequences are representative, since they have 100% sequence homology to the vast majority of the sequences of the complete IL-23 subunit proteins retrievable by GeneBank and SwissProt. Peptide sequences were tested with blastp (http://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastp&PAGE TYP E=BlastSearch&LINK LOC=blasthome) for IL-23 specificity.

Coupling of Peptides

[0063] KLH (SIGMA-ALDRICH, St. Louis, Mo. or biosyn Arzneimittel GmbH, Fellbach, Germany) is activated by incubation with N-gamma-maleimidobutyryl-oxysuccinimide ester (GMBS; AppliChem, Darmstadt, Germany) for 30 min at RT at a weight ratio of 1:2 and then dialyzed against Na-phosphate buffer (pH 6.7). 1 mg/ml peptide with a N- or C-terminally added Cysteine in 10% DMSO/20 mM Na-phosphate buffer (pH 6.8) is added to an equal volume of activated KLH and incubated for two hours at RT. Coupling efficiency is tested in an Ellmann assay or HPLC.

[0064] CRM197 (Pfenex Inc., San Diego, Calif.) is activated by incubation with N-gamma-maleimidobutyryl-oxysuccinimide ester (GMBS; AppliChem, Darmstadt, Germany) for 30 min at RT and then dialyzed against Na-phosphate buffer (pH 6.7). 1 mg/ml peptide with a N- or C-terminally added Cysteine in 10% DMSO/10 mM Na-phosphate buffer (pH 6.8) is added to an equal volume of activated CRM and incubated for two hours at RT. Coupling efficiency is tested by HPLC.

Preparation of the Vaccines

[0065] Carrier-coupled peptide is diluted in 1PBS and water to a final net peptide-concentration of 150 g/ml and sterile-filtered through a 0.22 m-mesh. Then 1/10 Vol 10 mg/ml Alum (Brenntag Biosector A/S, Frederiksund, DK) is added. The vaccine is aliquoted, incubated for 1h at RT and stored at 4 C. until use.

Immunization of Mice

[0066] Vaccines are vortexed and applied subcutaneously in the flank of mice (30 g net AFFITOPE in 200 l) with an insulin syringe with a G30-gauge (Omnican50, B.Braun Melsungen AG, Melsungen, Germany). Vaccination is repeated four times, on days 0, 14, 28 and 42. Mice are monitored one hour after injection for symptoms of distress. Typically, one peptide is tested in a group of five mice. When binary vaccines were tested, 15 g of each AFFITOPE were successively applied in separate injections in both flanks of the same animal.

Collection of Sera

[0067] Pre-plasma as well as the control plasma after the second and third vaccinations are taken by tail-clipping. 20 l of blood are taken from the tail vein with an EDTA-coated capillary (Hirschmann Laborgerate, Eberstadt, Germany) and blown out of the capillary into an Eppendorf-tube, containing 180 l PBS. The tube is centrifuged at 13000g for 10 min. at 4 C., then the supernatant is transferred to a new tube and frozen at 80 C. until further analysis.

[0068] For the collection of the final serum, mice are deeply anaesthesized. Blood is quantitativelytypically 600 lcollected from the animals in a serum tube (BD Microtainer, Becton Dickinson, Heidelberg, Germany). The tubes are left for 30 min. at RT, then centrifuged at RT for 3 min at 2000g. The supernatant is collected and transferred to an Eppendorf-tube and frozen at 80 C. until further analysis.

[0069] After blood collection, mice are sacrificed by cervical dislocation.

Testing of Sera

Response to Injected Peptide

[0070] BSA-coupled peptide (1 M, 50 l) is bound to 96-well plates.

[0071] Plates are incubated o/n at 4 C., then blocked after removal of unbound peptide for one hour at 37 C. with blocking buffer (PBS/1% BSA). After removal of blocking buffer, 50 l of serum is added to each well in serial 1:2 dilution steps with dilution buffer (PBS/0.1% BSA, 0.1% Tween20) starting with a dilution of 1:100. After one hour at 37 C., the supernatant is removed and the plates are washed three times (PBS, 0.1% Tween20). 50 l of biotinylated anti-mouse IgG (Southern Biotech, Birmingham, Ala., USA) at a concentration of 0.25 g/ml are added and incubated for one hour at 37 C. Unbound antibody is removed by three subsequent washing steps. 50 l/well (0.25 U/ml) of Streptavidin-horseradish peroxidase (Roche, Mannheim, Germany) are added and plates are incubated for 30 minutes at 37 C. After three washes, 50 l of ABTS substrate solution (0.068 M ABTS (AppliChem) in 0.1M Citric acid, pH 4.3) with H.sub.2O.sub.2 (1:1000) are added to each well and incubated at RT for 40 minutes. The reaction is stopped by addition of 50 l stop solution (1% SDS in water). Serum antibody concentrations are then measured on a Microwell reader (BioTek Power Wave 340 (BioTek, Winooski, Vt., USA) or Tecan Sunrise (Tecan Group Ltd., Mannedorf, Switzerland)) at 405 nm.

Crossreactivity to IL-23

[0072] rhu IL-23 (HumanZyme, Chicago, Ill., USA) at a concentration of 0.5 g/ml is deployed in 50 l aliquots in the wells of a 96-well plate. Plates are incubated o/n at 4 C. After removal of unbound coating agent, prediluted serum is added to each well at ascending dilution rates. After one hour at 37 C., the supernatant is removed and the plates are washed three times (PBS, 0.1% Tween20). 50 l of biotinylated anti-mouse IgG at a concentration of 0.25 g/ml are added and incubated for one hour at 37 C. Unbound antibody is removed by three subsequent washing steps. 50 l/well of streptavidin-horseradish peroxidase (0.25 U/ml) are added and plates are incubated for 30 minutes at 37 C. After three washes, 50 l of ABTS substrate solution (0.068 M ABTS in 0.1M Citric acid, pH 4.3) with H.sub.2O.sub.2 (1:1000) are added to each well and incubated at RT for 40 minutes. The reaction is stopped by addition of 50 l stop solution (1% SDS in water). Serum antibody concentrations are then measured on a Microwell reader at 405 nm.

Splenocyte Assay

[0073] IL-23-induced production of IL-17A by splenocytes and its suppression by serum antibodies is measured as published (Aggarwal et al., 2003). Briefly, spleens are excised from sacrificed C57BL/6j mice and splenocytes are singularized by mechanical disruption accompanied by DNAse I (20 g/ml)- and Collagenase D (100 Mandl Units/ml)-digest (both supplied by Roche). After removal of erythrocytes, cells are resuspended in RPMI supplemented with 10% FCS and 4 ng/ml rmuIL-2 (e-Bioscience, San Diego Calif., USA) at a concentration of 2.510.sup.6 cells/ml. Cells are stimulated with 0.1 ng/ml rhuIL-23 (R&D systems, Minneapolis, Minn., USA) and deployed in 200 l aliquots on the plates. 5 l of the sera to be tested are added to each well. The plates are incubated for three days at 37 C./5% CO.sub.2, after which the supernatants are collected and frozen at 80 C. until further analysis.

[0074] The analysis for IL-17A is performed with the IL-17A (homodimer) ELISA Ready-SET-Go! Kit (eBioscience) strictly following the instructions of the manufacturer.

Results for serum-inhibited IL-17A-expression are calculated as:

[00001]$\%.Math..Math.\mathrm{Expression}=100\frac{\mathrm{Expression}.Math..Math.\left(\mathrm{IL}-23+\mathrm{probe}\right)-\mathrm{Expression}.Math..Math.\left(\mathrm{background}\right)}{\begin{array}{c}\mathrm{Expression}.Math..Math.\left(\mathrm{IL}-23.Math.+\mathrm{irrelevant}.Math..Math.\mathrm{probe}\right)-\\ \mathrm{Expression}.Math..Math.\left(\mathrm{background}\right)\end{array}}$

STAT3 Assay

[0075] Phosphorylation of STAT3 of primary human lymphocytes is measured by flow Cytometry ((Krutzik and Nolan, 2003), modified)). Briefly, PBMC (isolated either from freshly collected blood or from Buffy coats from the Austrian Red Cross are resuspended in RPMI1640 and stimulated for three days with anti-CD3, anti-CD28 mAbs (both from Miltenyi Biotech GmbH, Bergisch Gladbach,Germany) and rhuIL-2 (eBioscience or R&D systems). After three washes in ice-cold PBS, the T cell blasts are resuspended in PBS, aliquoted to 210.sup.5 cells/200 l and stimulated with 5 ng/ml rhu IL-23 (eBioscience) with or without IL-23-inhibiting agents. After incubation for 20 minutes at 4 C., cells are fixed and permeabilized using the BD Phosflo.sup.w Kit (Becton Dickinson) following the instructions of the manufacturer. The antibody used for intracellular staining of phosphorylated STAT3 is mouse Anti-human STAT3 (pY705)-Alexa Fluor488. Mouse anti-human CD4-APC is used for counterstaining (both antibodies supplied by Becton Dickinson). Alexa Fluor488-fluorescence of the CD4.sup.+ T cell blasts is measured on a FACSCanto II Flow Cytometer (Becton Dickinson).

[0076] Serum-caused reduction of STAT3-phosphorylation is calculated as:

[00002]$\%.Math..Math.\mathrm{Expression}=100\frac{\mathrm{Expression}.Math..Math.\left(\mathrm{IL}-23+\mathrm{probe}\right)-\mathrm{Expression}.Math..Math.\left(\mathrm{unstimulated}\right)}{\begin{array}{c}\mathrm{Expression}.Math..Math.\left(\mathrm{IL}-23.Math.+\mathrm{irrelevant}.Math..Math.\mathrm{probe}\right)-\\ \mathrm{Expression}.Math..Math.\left(\mathrm{unstimulated}\right)\end{array}}$

Fine Epitope Mapping

[0077] Microarray-based fine epitope mapping was performed by PEPperPRINT GmbH (Heidelberg, GE) using the PEPperMAP technology. Briefly, the sequence of the domain 3 and its immediate surroundings was split into overlapping 12mer peptides, sliding by one amino acid. The resulting peptides were spotted in triplicate on a glass slide. These treated slides were incubated with diluted murine anti-p6063-serum. Secondary goat anti-mouse IgG (H+L) DyLight680 antibody was used to detect serum antibodies. Fluorescence intensity was measured with a LI-COR Odyssey Imaging System (LI-COR Biosciences, NE, US) and quantified.

In Silico Analyses

[0078] SYFPEITHI (http://syfpeithi.de/) was used to predict MHC class I and II affinities of fragments contained in vaccination peptides of different length. PAProC (http://www.paproc.de/) was used to predict proteasomal cleavage sites.

Sequences

[0079]

TABLE-US-00001 Name SeqID Position Sequence IL-23p19 1 p19.sub.1-189 GenBankAAH66268.1 IL-23p40 2 p40.sub.1-328 GenBankAAD56386.1 p6058 3 p19.sub.20-33 RAVPGGSSPAWTQC p6059 4 p19.sub.42-59 C-TLAWSAHPLVGHMDLREE p6294 5 p19.sub.46-59 C-SAHPLVGHMDLREE p7457 6 p19.sub.51-72 C-VGHMDLREEGDEETTNDVPHIQ p7458 7 p19.sub.51-72 VGHMDLREEGDEETTNDVPHIQ-C p7459 8 p19.sub.90-110 C-LQRIHQGLIFYEKLLGSDIFT p7460 9 p19.sub.90-110 LQRIHQGLIFYEKLLGSDIFT-C p6061 10 p19.sub.100-119 C-YEKLLGSDIFTGEPSLLPDS p6060 11 p19.sub.100-129 C-YEKLLGSDIFTGEPSLLPDSPVGQLHASLL p6291 12 p19.sub.105-121 C-GSDIFTGEPSLLPDSPV p6062 13 p19.sub.121-135 C-VGQLHASLLGLSQLL p7461 14 p19.sub.130-149 C-GLSQLLQPEGHHWETQQIPS p7462 15 p19.sub.130-149 GLSQLLQPEGHHWETQQIPS-C p6063 16 p19.sub.139-159 C-GHHWETQQIPSLSPSQPWQRL p7463 17 p19.sub.167-188 C-RSLQAFVAVAARVFAHGAATLS p7464 18 p19.sub.167-188 RSLQAFVAVAARVFAHGAATLS-C p7434 19 p19.sub.144-158 C-TQQIPSLSPSQPWQR p8464 20 p19.sub.140-147 C-HHWETQQI p7432 21 P19.sub.138-158 C-EGHHWETQQIPSLSPSQPWQR p8320 22 p19.sub.132-151 C-SQLLQPEGHHWETQQIPSLS p8321 23 p19.sub.134-151 C-LLQPEGHHWETQQIPSLS p8322 24 p19.sub.136-151 C-QPEGHHWETQQIPSLS p8323 25 p19.sub.138-151 C-EGHHWETQQIPSLS p8459 26 p19.sub.136-149 C-QPEGHHWETQQIPS p8460 27 p19.sub.136-147 C-QPEGHHWETQQI p8461 28 p19.sub.136-145 C-QPEGHHWETQ p8462 29 p19.sub.136-143 C-QPEGHHWE p8397 30 p19.sub.144-157 C-TQQIPSLSPSQPWQ p8398 31 p19.sub.144-156 C-TQQIPSLSPSQPW p8399 32 p19.sub.144-155 C-TQQIPSLSPSQP p8400 33 p19.sub.144-154 C-TQQIPSLSPSQ p8761 34 p19.sub.144-153 C-TQQIPSLSPS p8762 35 p19.sub.144-152 C-TQQIPSLSP p8763 36 p19.sub.144-151 C-TQQIPSLS p8332 37 p19.sub.137-146 C-PEGHHWETQQ p8333 38 p19.sub.155-164 C-PWQRLLLRFK p8337 39 p19.sub.127-137 C-SLLGLSQLLQP p8759 40 p19.sub.137-155 C-PEGHHWETQQIPSLSPSQP p7977 41 p19.sub.160-179 C-LLRFKILRSLQAFVAVAARV p9165 42 P19.sub.152-159 C-PSQPWQRL p9269 43 P19.sub.136-154 C-QPEGHHWETQQIPSLSPSQ p6449 44 p40.sub.35-49 C-LDWYPDAPGEMVVLT p4994 45 C5a.sub.55-74 CVVASQLRANISHKDMQLGR
List of sequences used in this study. C- followed or C preceded by the sequence indicates that the cysteine needed to attach the peptide to the carrier is not part of the original protein-sequence, while C followed preceded by the sequence indicates a naturally occurring Cysteine; peptide names (pXXXX) for the C-coupled peptide and the peptide without added C are the same due to the identical core sequence.

TABLE-US-00002 Name SeqID Position Sequence p6063 16 p19.sub.139-159 C- GHHWETQQIPSLSPSQPWQRL p8322 24 P19.sub.136-151 C-QPEGHHWETQQIPSLS p8461 28 P19.sub.136-145 C-QPEGHHWETQ p8400 33 p19.sub.144-154 C-TQQIPSLSPSQ p9269 43 p19.sub.136-154 C-QPEGHHWETQQIPSLSPSQ -p9440 46 P19.sub.136-153 C-QPEGHHWETQQIPSLSPS -p9441 47 p19.sub.136-152 C-QPEGHHWETQQIPSLSP
List of claimed sequences. C- followed or C preceded by the sequence indicates that the cysteine needed to attach the peptide to the carrier is not part of the original protein-sequence, while C followed preceded by the sequence indicates a naturally occurring Cysteine

Results

[0080] Definition of Region of Interest (Domain 3/p6063)

[0081] To be qualified as a potential vaccine, a peptide is required to elicit sera which fulfil three conditions: The serum must a) react with the immunizing peptide, b) crossreact with the original target, i.e.: IL-23 and c) interfere with IL-23 function. Every peptide used for immunization in this study was assayed for these conditions.

Screening for Epitopes that Induce IL-23 Binding Sera

[0082] We used 16 overlapping peptides to screen the p19 subunit for immunogenic regions. The peptides were N- or C-terminally linked to the carrier and cover approximately 90% of the sequence. While all elicited sera were able to bind the immunizing peptides (data not shown), we found that 14/16 sera contained antibodies that crossreacted with rhuIL-23 (Tab. 1). Likewise, the target region of Ustekinumab was demonstrated to contain immunogenic regions to obtain data which would allow us comparison with peptides from a known immunogenic region (Tab. 1).

Screening for Functionally Relevant Epitopes

[0083] To determine whether the peptide-specific sera were able to interfere with IL-23 function, we employed two assays: Firstly the splenocyte assay, where we used rhu-IL-23 to stimulate IL-17A production in murine cells, and secondly the STAT3-assay that uses primary human cells as a read-out for rhuIL-23 function via STAT3-phosphorylation upon binding of the human IL-23 receptor.

[0084] We found three regions in IL-23p19 containing immunogenic epitopes that repeatedly induced functionally relevant antibodies (FIGS. 1A and B). These regions were dubbed domain 1, 2 and 3 respectively. p6059 (p19.sub.42-57) is situated in domain 1. In crystal structure models of the cytokine (http://www.ncbi.nlm.nih.gov/Structure/mmdb/mmdbsrv.cgi?uid=66205, http://www.ncbi.nlm.nih.gov/Structure/mmdb/mmdbsrv.cgi?uid=66470), this domain was found to be situated on the interface between the two subunits. p6061 (p19.sub.100-119) and p6063 (p19.sub.139-159) are situated in domains 2 and 3, respectively. These domains are situated facing outward of the complex according to the models and thus are likely to be accessible for antibodies. They therefore represent promising targets for immunologic intervention against IL-23. The peptide from the Ustekinumab target region, p6449 (p40.sub.35-49) also inhibits IL-23 function (FIG. 1C). It is situated on a loop on the far end of the large subunit facing outward.

[0085] Data from p-19-peptide-induced sera not inhibiting IL-23 function are depicted in FIG. 1D.

[0086] Of the tested peptides, p6063 (p19.sub.139-159) repeatedly elicited the most potent IL-23-inhibiting sera. Thus, we chose to focus on domain 3 for the development of an anti-IL-23 vaccinating agent. The location of domain 3 suggests that functional inhibition of IL-23 by domain-specific sera occurs via an inhibition of IL-23/IL-23R interaction.

Fine Epitope Analysis of Domain 3

[0087] Exact epitope mapping surprisingly revealed the presence of two close yet discrete epitopes in the p6063 region. Intriguingly, antibodies from anti-p6063 serum recognize two different regions, as demonstrated by microarray-based binding studies (FIG. 2).

[0088] In order to confirm the epitope-specificity of the functionally relevant antibodies contained within p6063 serum, we added p6063 peptide to the splenocyte assay to provide a specific competitor to IL-23 for the serum antibodies. Indeed, addition of this peptide abolished IL-23 inhibition completely, whereas addition of irrelevant peptide (p4994) did not interfere with inhibition, corroborating that it is an effect caused by specific antibodies and not by unspecific interference. (FIG. 3).

[0089] To confirm these findings and to generate supplementary information about the exact location of the two discrete epitopes, we used truncated forms of the p6063 peptide, namely p8464 (p19.sub.140-147) from the N-terminus and p7434 (p19.sub.144-158) from the C-terminus as competitors. Both peptides were able to abolish IL-23-inhibition only partly, corroborating that a) the p6063 region contains more than one relevant epitope and b) these peptides indeed competed with different epitopes for the serum antibodies (FIG. 3). Consequently, combined addition of p8464 and p7434 to the assay leads to blocking of inhibition equal to the one obtained with p6063. Combining either truncated peptide with the irrelevant peptide did not result in synergistic effects (FIG. 3).

Definition of Minimal Epitopes

[0090] Aiming at the definition of the minimal core sequences, we vaccinated mice with successively truncated forms of peptides contained within domain 3.

[0091] To define the N-terminal epitope we started with the 14-mer p8459 (p19.sub.136-149) which contains the p8464 peptide as core with extensions both N- and C-terminally. C-terminal truncations were performed in steps of two amino acids to define the demarcation against the C-terminal epitope. The best results were obtained with the p8461 (p19.sub.136-145) 10-mer QPEGHHWETQ (FIG. 4A).

[0092] To define the minimal sequence of the C-terminal epitope, we started with the 14-mer p8397 (p19.sub.144-157). This peptide is a truncated version of p7434 and as such mimics the second epitope in domain 3. It was subsequently cropped C-terminally in steps of one amino acid. While all truncated peptides yielded functionally active sera, the best results were obtained with the 11-mer p8400 (p19.sub.144-154) TQQIPSLSPSQ (FIG. 4B).

Combination Strategies: Biepitopic Peptides and Binary Vaccines

[0093] In an attempt to further increase functional inhibition of IL-23, we injected mice with vaccines containing two immunogenic epitopes at once. The idea behind this is that immunologic attack of different sites of the same molecule/complex might not only interfere more effectively with receptor binding but is also expected to increase clearance of the organism by more polyclonal opsonisation of the target and subsequent crossactivation of phagocytic cells.

[0094] We employed two different strategies towards that aim: On one hand, we used biepitopic vaccines, on the other hand binary vaccines.

Biepitopic Vaccines

[0095] Biepitopic vaccines contain longer peptides combining two epitopes. Possible scenarios include naturally occurring stretches of epitopes in either their original or altered sequence, or the combination of epitopes from distant locations on the same or even different subunits of the complex, joined in one peptide, possibly separated by a spacer. To confirm our concept we used the peptide p9269 (p19.sub.136-154), which corresponds to the original IL-23p19-sequence spanning p8461 and p8400. Indeed, the serum elicited by p9269 inhibited IL-23-function more effectively than each of the sera against p8461 and p8400 alone (FIG. 5A).

[0096] The use of peptides as immunogens harbours two potential safety hazards: First the presence of MHC class I epitopes in the sequence of the peptide and secondly crossreactivity of the sera with other, unrelated proteins. In the first case, scenarios are conceivable, where vaccination peptides enter the MHC class I pathway, which could lead to the generation of a peptide-fragment-specific cytotoxic T cell response that might in turn be aimed against IL-23 producing cells and thus cause severe cell/tissue damage. This scenario is clearly undesirable. To address this question, we subjected p9269 to SYFPEITHI, an algorithm designed to calculate MHC class I binding peptides in a given sequence. The algorithm predicted seven potential strong MHC class I binders, consisting of four different sequences with a predilection for five different MHCI-alleles (Tab. 2). A second algorithm, PAProC, revealed that one of these sequences could be generated by proteasomal degradation (FIG. 5B).

[0097] We addressed the second question by fragmenting the sequence of p9269 into overlapping hexapeptides and performing BLAST-searches against the SWISS-Prot database. 27 homologies with unrelated proteins were found (FIG. 5C). Interestingly, the vast majority of the homologies are linked to the three C-terminal amino acids of p9269. If the BLAST-search for linear homologies is extended to a sequence elongated by one C-terminal amino acidas would be p8759one protein with a heptapeptide homology and five more proteins with hexapeptide-homologies can be found, not counting their various splice-variants.

[0098] It is conceivable that a truncation of p9269 by one, preferably two or even three C terminal amino acids should remedy both potential safety shortcomings, while still addressing both epitopes of domain 3. Omitting the three C-terminal amino acids of p9269 results in the 16-mer p8322 (p19.sub.136-151). This peptide indeed elicits sera with IL-23-inhibiting capacity similar to p9269 (FIG. 5D). While SYFPEITHI predicts six of the seven strong MHC class I binders as in the longer peptide, these fragments cannot be generated by proteasomal degradation according to PAProC (FIG. 5E). On the other hand, only one of the 27 hexapeptide homologies to unrelated proteins identified in p9269 remains in p8322 (FIG. 5F).

[0099] Taken together, these results indicate that biepitopic peptides can be more powerful in eliciting anti-IL-23 immunity than single monoepitopic peptides and can be designed to be satisfactorily safe.

Binary Vaccines

[0100] In binary vaccines, two monoepitopic peptides are concomitantly applied to the subject. Possible scenarios involve peptides from the same domain, from spacially distinct domains of from different subunits of a molecule/complex or even different targets. Peptides can be coupled to the same or to different carriers, the latter to avoid carrier-dependent epitope inhibition.

[0101] To confirm this concept, we injected mice with separate vaccines containing p8461 coupled to KLH and p8400 coupled to CRM197 applied in separate locations (i.e.: in opposing flanks). Bivalent vaccines indeed repeatedly effectuated powerful anti-IL-23 responses similar to the biepitopic peptides described above (FIG. 5D).

[0102] To address safety issues, we used the SYPPEITHI-algorithm to search for possible MHC Class I and MHC Class II binding peptides contained in the p9269 (p19.sub.136-154) 19-mer (FIG. 6A), in the p8322 (p19.sub.136-151) 16-mer (FIG. 6B) and in the peptides containing the minimal epitopes, the N-terminal p8461 (p19.sub.136-145) 10-mer (FIG. 6C) and the C-terminal p8400 (p19.sub.144-154) 11-mer (FIG. 6D). All three peptides were subjected to epitope-search for 8- to 15-mers in a totality of the 124 n-mer/MHC I/II-combinations available in SYFPEITHI. Indeed, a clear correlation between the length of the analyzed peptide and the amount of potential epitopes is apparent: p9269 contains 603 possible peptides with a SYFPEITHI-score >0, of which seven MHC class I binders display a score 20 (See Tab 2), making them potential strong MHC binders. p8322 contains 368 possible peptides with a SYFPEITHI-score >0, among them six putative strong binders as in p9269, as mentioned above. p8400 contains 142 and p8461 only 75 possible peptides with a SYFPEITHI-score >0. Neither short peptide contains potential high-binders. According to PAProC, no fragments long enough for MHC I-binding can be generated by proteasomal cleavage from p8461 or p8400 (not shown).

[0103] Further experiments demonstrate that the concept of bivalent vaccines yields also beneficial results, when the monovalent vaccines are coming from different domains of the same subunit (i.e.: p6061 (p19.sub.100-119) from domain 2 and p6063 (p19.sub.139-159) from domain 3 of the p19 subunit (FIG. 6E) or from different subunits (i.e.: p6063 from the small subunit and p6449 (p40.sub.35-49) from the large subunit) (FIG. 7F).

[0104] Taken together, these results show powerful anti-IL-23 responses elicited by bivalent vaccines and indicate that the use of shorter peptides precludes safety issues in the context of target-mediated cytotoxic responses.

Delimitation Against Previously Published Peptides

[0105] Peptides derived from sequences in or in the immediate vicinity of domain 3 described in other patents have been synthesized with a N-terminal cysteine, coupled to KLH and injected in mice. The resulting sera have been tested for anti-IL-23 activity (FIG. 7).

[0106] When tested in two independent experiments, p8461 (p19.sub.136-145) reduced IL-17 expression by splenocytes to 26.24.5% and p8400 (p19.sub.144-154) to 34.63.0%. p6063 (p19.sub.139-159) reduced IL-17 expression by splenocytes to 23.62.5% as compared to an irrelevant serum.

[0107] p8332 (p19.sub.137-146) (i.e.: PEGHHWETQQ) and p8333 (p19.sub.155-164) (i.e.: PWQRLLLRFK) are taken from EP 2 392 597 A2 [Lewis], where they are mentioned as target-sequences for bispecific antibodies against IL-23 and IL-17A. Serum elicited to p8332 inhibits IL-23 function 48% less effectively then serum against p6063, and, importantly, 33% less effectively than serum elicited against the closely overlapping p8461. p8333-serum inhibits IL-23 300% less efficiently than p6063-serum, similar to the irrelevant serum.

[0108] p8337 (p19.sub.127-137) (i.e.: SLLGLSQLLQP) was described in WO 2007/005955 A2 [Benson], where it represents a target site for engineered anti-IL-23 antibodies. The resulting serum also reduced expression of IL-17A to a degree similar to the irrelevant control serum.

[0109] p8759 (p19.sub.137-155) (i.e.: PEGHHWETQQIPSLSPSQP) was mentioned in WO 2005/108425 A1 [Bachman/Cytos] as possible peptide sequence to be used for anti-IL-23 immunization after coupling to a virus-like particle, although no immunization studies were shown using this or any other IL-23p19-derived peptide. The resulting serum reduced expression of IL-17A 92% less effectively than p6063-derived serum, 31% less effectively than p8400-derived serum and 73% less effectively than p8461-derived serum.

[0110] p7977 (p19.sub.160-179) (i.e.:LLRFKILRSLQAFVAVAARV), described in WO 03/084979 A2 [Zagury] as possible agent for anti-cytokine immunization is situated C-terminally of the p6063-region. Serum elicited with this peptide reduced IL-17A-expression 275% less effectively than p6063-derived serum serum.

[0111] p9165 (p19.sub.152-159) (i.e.: PSQPWQRL) as mentioned in WO 2007/027714 A2 [Presta] where it represents a target site for engineered anti-IL-23 antibodies, is situated at the C-terminal end of p6063. Serum elicited with this peptide reduced IL-17A-expression also only in the same range as the irrelevant serum.

[0112] These data demonstrate that peptides we had defined to contain the minimal epitopes for the N-terminal epitopei.e.: p8461 (p19.sub.136-145) and the C-terminal epitopei.e.: p8400 (p19.sub.144-154) as well as the longer p6063 elicited sera are superior in suppressing IL-23 activity to overlapping sequences earlier described.

Graphs & Tables

[0113]

TABLE-US-00003 TABLE1 PeptidesusedtoscanIL-23p19andtheUstekinumab-regionforimmunogenic regions.C- followedor-C precededbythesequenceindicatesthatthe cysteineneededtoattachthepeptidetothecarrierisnotpartofthe originalprotein-sequence,whileC precededbythesequenceindicatesa naturallyoccurringcysteine.Seraweredeemedbinding,whentheminimal dilutionfactortoattainoDmax/2wasatleast1:100. Functional Name SeqID Position Sequence Titer.sup.1 relevance.sup.2 p6058 3 p19.sub.20-33 RAVPGGSSPAWTQC 100 - p6059 4 p19.sub.42-59 C-TLAWSAHPLVGHMDLREE 3000 + p6294 5 p19.sub.46-59 C-SAHPLVGHMDLREE 1000 - p7457 6 p19.sub.51-72 C-VGHMDLREEGDEETTNDVPHIQ 300 - p7458 7 p19.sub.51-72 VGHMDLREEGDEETTNDVPHIQ-C 300 - p7459 8 p19.sub.90-110 C-LQRIHQGLIFYEKLLGSDIFT 1000 - p7460 9 p19.sub.95-110 LQRIHQGLIFYEKLLGSDIFT-C 300 - p6061 10 p19.sub.100-119 C-YEKLLGSDIFTGEPSLLPDS 3000 + p6060 11 p19.sub.100-129 C-YEKLLGSDIFTGEPSLLPDSPVGQLHASLL 1000 - p6291 12 P19.sub.105-121 C-GSDIFTGEPSLLPDSPV 1000 - p6062 13 P19.sub.121-135 C-VGQLHASLLGLSQLL 100 - p7461 14 p19.sub.130-149 C-GLSQLLQPEGHHWETQQIPS 1000 - p7462 15 p19.sub.130-149 GLSQLLQPEGHHWETQQIPS-C 1000 - p6063 16 p19.sub.138-158 C-GHHWETQQIPSLSPSQPWQRL 300 ++ p7463 17 P19.sub.167-188 C-RSLQAFVAVAARVFAHGAATLS <100 - p7464 18 p19.sub.167-188 RSLQAFVAVAARVFAHGAATLS-C <100 - p6449 44 p40.sub.35-49 C-LDWYPDAPGEMVVLT 3000 + .sup.1n is the dilution-factor in a -IL23 ELISA at which oDmax/2 is reached. .sup.2Functional relevance denotes the ability of a given serum to inhibit functional activity of rhuIL-23 as tested by splenocyte or STAT3p-assay. -: no response, +: response, ++: strong response

TABLE-US-00004 TABLE2 StrongMHCClassIbinderswithinthesequenceofp9269andp8322as definedbySYFPEITHI.Allele describestheMHCI-Alleletowhichthe respectivesequencebinds.Anemptyspaceinthiscolumnindicates thatthealleleisthesameasinthelineabove.Sequence showsthe aminoacidsequenceofthepredictedbinder.Boldlettersindicate primaryanchorpositions,underlinedlettersindicatesecondary anchors.Score representsthescorecalculatedbySYFPEITHI.Higher scoresindicateahigherprobabilitytobeastrongMHCclassI binder.Frequency denotesthefrequencyofagivenallelein differenthumangeographicaland/orethnicalpopulations.Population showsthegeographicalregionand/orethnicity,inwhichthehighest frequencyofagivenalleleoccurs.Frequency andPopulation are retrievedfromhttp://www.allelefrequencies.net/. No Allele Sequence Score Frequency Population 1 HLA-A26 ETQQIPSLS 21 0-13% Cuba 2 ETQQIPSLSP 20 3 HLA-B18* WETQQIPSL 20 0-16% Balkans 4 HLA-B37* WETQQIPSL 24 0-7% Belgium 5 HLA-B40:01* WETQQIPSL 20 0-28% HKChinese 6 HLA-B44:02* PEGHHWETQQI 20 0-25% Ireland 7 WETQQIPSL 23

[0114] From this disclosure, the following preferred embodiments are defined:

[0115] 1. Vaccine for use in the prevention or treatment of an interleukin 23 (IL-23) related disease, comprising a peptide bound to a pharmaceutically acceptable carrier, wherein said peptide is selected from the group QPEGHHWETQQIPSLS (SEQ ID No. 103; p8322), GHHWETQQIPSLSPSQPWQRL (SEQ ID No. 97; p6063), QPEGHHWETQ (SEQ ID No. 98; p8461), TQQIPSLSPSQ (SEQ ID No. 99; p8400), QPEGHHWETQQIPSLSPSQ (SEQ ID No. 100; p9269), QPEGHHWETQQIPSLSPS (SEQ ID No. 101; p9440), and QPEGHHWETQQIPSLSP (SEQ ID No. 102; p9441), especially QPEGHHWETQQIPSLS (SEQ ID No. 103; p8322) and wherein said IL-23 related disease is preferably selected from the group psoriasis, psoriatic arthritis, rheumatoid arthritis, systemic lupus erythematosus, diabetes, especially type 1 diabetes; atherosclerosis, inflammatory bowel disease (IBD)/M. Crohn, multiple sclerosis, Behcet disease, ankylosing spondylitis, Vogt-Koyanagi-Harada disease, chronic granulomatous disease, hidratenitis suppurtiva, anti-neutrophil cytoplasmic antibodies (ANCA-) associated vasculitides, neurodegenerative diseases, especially M. Alzheimer or multiple sclerosis; atopic dermatitis, graft-versus-host disease, cancer, especially Oesophagal carcinoma, colorectal carcinoma, lung adenocarcinoma, small cell carcinoma, and squamous cell carcinoma of the oral cavity; especially psoriasis, neurodegenerative diseases or IBD.

[0116] 2. Vaccine according to embodiment 1, wherein at least one cysteine residue is bound to the N- or C-terminus of the peptide.

[0117] 3. Vaccine according to embodiment 1 or 2, wherein at least one cysteine residue is bound to the N-terminus of the peptide.

[0118] 4. Vaccine according to any one of embodiments 1 to 3, wherein the carrier is a protein carrier.

[0119] 5. Vaccine according to embodiment 4, wherein the protein carrier is selected from the group consisting of keyhole limpet haemocyanin (KLH), tetanus toxoid (TT) or diphtheria toxin (DT).

[0120] 6. Vaccine according to any one of embodiments 1 to 5, wherein the vaccine is formulated with an adjuvant, preferably wherein the peptide bound to the carrier is adsorbed to alum.

[0121] 7. Vaccine according to any one of embodiments 1 to 6, formulated for intravenous, subcutaneous, intradermal or intramuscular administration.

[0122] 8. Vaccine according to any one of embodiments 1 to 7, wherein the peptide is contained in the vaccine in an amount from 0.1 ng to 10 mg, preferably 10 ng to 1 mg, in particular 100 ng to 100 g.

[0123] 9. Vaccine according to any one of embodiments 1 to 8, wherein the peptide is bound to the carrier by a linker, preferably a peptide linker, especially a peptide linker having from 2 to 5 amino acid residues.

[0124] 10. Vaccine according to embodiment 9, wherein the peptide linker is selected from the group Gly-Gly-Cys, Gly-Cys, Cys-Gly and Cys-Gly-Gly.

[0125] 11. Vaccine according to any one of embodiments 1 to 10, wherein the vaccine is a biepitopic vaccine, especially a vaccine comprising a peptide of the group QPEGHHWETQQIPSLSPSQ (SEQ ID No. 100; p9269), QPEGHHWETQQIPSLSPS (SEQ ID No. 101; p9440), QPEGHHWETQQIPSLSP (SEQ ID No. 102; p9441), and QPEGHHWETQQIPSLS (SEQ ID No. 103; p8322).

[0126] 12. Vaccine according to any one of embodiments 1 to 11, wherein the vaccine is a binary vaccine, especially a vaccine comprising a peptide from the group QPEGHHWETQQIPS (SEQ ID No. 104; p8495), QPEGHHWETQQIP (SEQ ID No. 105; p8459-1), QPEGHHWETQQI (SEQ ID No. 106; p8460), QPEGHHWETQQ (SEQ ID No. 107; p8460-1), QPEGHHWETQ (SEQ ID No. 98; p8461), QPEGHHWET (SEQ ID No. 108, p8461-1), QPEGHHWE (SEQ ID No. 109; p8462) and a peptide from the group TQQIPSLSPSQPWQ (SEQ ID No. 110, p8397), TQQIPSLSPSQPW (SEQ ID No. 111, p8398), TQQIPSLSPSQP (SEQ ID No. 112, p8399), TQQIPSLSPSQ (SEQ ID No. 99; p8400), TQQIPSLSPS (SEQ ID No. 113; p8761), TQQIPSLSP (SEQ ID No. 114; p8762), TQQIPSLS (SEQ ID No 115; p8763), preferably the peptide QPEGHHWETQ (SEQ ID No. 98; p8461) and the peptide TQQIPSLSPSQ (SEQ ID No. 99; p8400), each bound to a separate carrier.

[0127] 13. Vaccine kit comprising a vaccine according to any one of embodiments 1 to 12 and a further vaccine addressing the Th17/IL-23 pathway, preferably an anti-IL-23 vaccine, especially an anti-p19-IL-23 vaccine.

[0128] 14. Peptide, selected from the group GHHWETQQIPSLSPSQPWQRL (SEQ ID No. 97; p6063), QPEGHHWETQ (SEQ ID No. 98; p8461), TQQIPSLSPSQ (SEQ ID No. 99; p8400), QPEGHHWETQQIPSLSPSQ (SEQ ID No. 100; p9269), QPEGHHWETQQIPSLSPS (SEQ ID No. 101; p9440), QPEGHHWETQQIPSLSP (SEQ ID No. 102; p9441), QPEGHHWETQQIPSLS (SEQ ID No. 103; p8322), QPEGHHWETQQIPS (SEQ ID No. 104; p8495), QPEGHHWETQQIP (SEQ ID No. 105; p8459-1), QPEGHHWETQQI (SEQ ID No. 106; p8460), QPEGHHWETQQ (SEQ ID No. 107; p8460-1), QPEGHHWETQ (SEQ ID No. 98; p8461), QPEGHHWET (SEQ ID No. 108, p8461-1), QPEGHHWE (SEQ ID No. 109; p8462), TQQIPSLSPSQPWQ (SEQ ID No. 110, p8397), TQQIPSLSPSQPW (SEQ ID No. 111, p8398), TQQIPSLSPSQP (SEQ ID No. 112, p8399), TQQIPSLSPSQ (SEQ ID No. 99; p8400), TQQIPSLSPS (SEQ ID No. 113; p8761), TQQIPSLSP (SEQ ID No. 114; p8762), and TQQIPSLS (SEQ ID No 115; p8763).

[0129] 15. Peptide pair, wherein one peptide is selected from the group QPEGHHWETQQIPS (SEQ ID No. 104; p8495), QPEGHHWETQQIP (SEQ ID No. 105; p8459-1), QPEGHHWETQQI (SEQ ID No. 106; p8460), QPEGHHWETQQ (SEQ ID No. 107; p8460-1), QPEGHHWETQ (SEQ ID No. 98; p8461), QPEGHHWET (SEQ ID No. 108, p8461-1), QPEGHHWE (SEQ ID No. 109; p8462) and the second peptide is selected from the group TQQIPSLSPSQPWQ (SEQ ID No. 110, p8397), TQQIPSLSPSQPW (SEQ ID No. 111, p8398), TQQIPSLSPSQP (SEQ ID No. 112, p8399), TQQIPSLSPSQ (SEQ ID No. 99; p8400), TQQIPSLSPS (SEQ ID No. 113; p8761), TQQIPSLSP (SEQ ID No. 114; p8762), TQQIPSLS (SEQ ID No 115; p8763), preferably the peptide QPEGHHWETQ (SEQ ID No. 98; p8461) and the peptide TQQIPSLSPSQ (SEQ ID No. 99; p8400).

REFERENCES

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