WO2006046613A1

WO2006046613A1 - Il-18-like peptide, fused peptide thereof, method of producing the same and use of the same

Info

Publication number: WO2006046613A1
Application number: PCT/JP2005/019709
Authority: WO
Inventors: Kazuhisa Sugimura; Kyozo Tsukamoto
Original assignee: Kagoshima University
Priority date: 2004-10-27
Filing date: 2005-10-26
Publication date: 2006-05-04
Also published as: JP2006124297A

Abstract

An IL-18-like peptide capable of regulating/controlling the interaction between IL-18, which participates in inflammation or immune abnormality diseases such as atopic dermatitis, and its receptor; a medicinal composition containing this peptide as the active ingredient; and a physiological activity inhibitor. It is intended to disclose a method of producing the above-described IL-18-like peptide which comprises screening a phage random peptide library with the use of a human anti-IL-18 antibody. This IL-18-like peptide can regulate/control the interaction between IL-18, which seemingly participates in immune abnormality diseases such as atopic dermatitis, and its receptor and is useful as an IL-18 activity inhibitor. Thus, it is expected as being applicable to various inflammatory diseases.

Description

Specification

IL-18-like peptide, its fusion peptide, production method and use

Technical field

The present invention relates to an IL-18-like peptide that can bind to an anti-IL-18 antibody that inhibits IL-18 signal, a method for producing the same, and a use thereof. This peptide is expected as a therapeutic agent for inflammation and immune abnormal diseases caused by IL-18.

Background art

[0002] Site-forced IL-18 (interleukin 18) was discovered as an interferon-gamma (IFN-γ) inducer, but subsequently became an allergic reaction mediated by IgE antibodies, and was not mediated by IgE. It is known that this is the cause of malignant dermatitis and bronchial asthma, and that this overproduction causes serious organ damage in the liver and intestinal tract. In addition to these diseases, the pathology of IL 18 has been pointed out in various diseases (see Non-Patent Document 1).

Recently, a screening method using a phage random peptide library has attracted attention as a means for identifying an interaction site in a receptor-ligand system. This is because a gene encoding a peptide molecule having a random amino acid sequence is inserted into a gene (gene 3) encoding a p3 molecule, which is a constituent protein of a fuzz, to form a p3 molecule containing the peptide molecule ( A fusion of a gene 3 protein and a peptide molecule having a random amino acid sequence (hereinafter also referred to as G3P) expressed on Ml 3 phage (phage of filamentous single-stranded DNA), and a phage random peptide library This is a method of screening the phage expressing the target peptide molecule in Nung, etc., and analyzing the base sequence of the gene 3 gene portion of the phage to determine the interaction site.

[0004] Using this method, determination of epitopes on the p53 molecule by anti-p53 monoclonal antibody (but highly homologous to p53), bEGF analog strength screened by anti-bEGF antibody ¾FGF and its receptor FGFR — Discovery of peptide sequences that bind to the binding of 1 or bind to anti-acetylylcholine receptor epitope antibody (homology to the receptor) However, it does not mention whether or not it can bind to acetylcholine).

[0005] Further, the present inventors have further developed this technique, and for CTLA-4, anti-CTLA

(4) A method for obtaining a molecule using a monoclonal antibody that mimics the three-dimensional structure of the target molecule to which the antibody binds without depending on the primary sequence of the amino acid and that can control the interaction of the receptor ligand. (Non-Patent Documents 2 and 3).

Non-Patent Document 1: J. Exp. Med. 1994 (4), 535-545, 2004

Non-Patent Document 2: Nature biotech. 16, 267-270, 1998

Non-Patent Document 3: J. Immunol. 161: 6622-6628, 1998

Disclosure of the invention

Problems to be solved by the invention

[0006] As described above, the control of IL-18 production or activity is a treatment method for such IL-18-dependent atopic dermatitis or the onset of diseases caused by excessive production of IL-18. Is extremely important as a treatment for Thl disease that induces or hate. Therefore, if an IL-18-like peptide that inhibits the physiological activity of IL-18 can be developed, it is expected to be an effective therapeutic means for many diseases involving IL-18.

Means for solving the problem

[0007] The present invention relates to a target protein obtained from a phage peptide library using a monoclonal antibody that inhibits signal transduction due to binding of a receptor and a ligand that control a biological reaction and recognizes the three-dimensional structure of the molecule. Based on the inventors' knowledge that non-patent documents 2 and 3 can isolate an immunoregulatory molecule having a peptide sequence that mimics the tertiary structure of the molecule, Peptide development using M13 phage library (PhD-12 and PhD-C7, Biolabs. USA) as a smaller molecule IL-18 inhibitor was attempted to obtain several anti-IL-18 antibody-binding phage clones It was found that the peptides successfully inserted into the phage clones had a common amino acid sequence site, and based on these findings, the present invention was completed. .

That is, the present invention includes the inventions described in 1) to 9) below.

1) An IL-18-like peptide that can bind to an anti-IL-18 antibody that inhibits the IL-18 signal.

2) The IL 18-like peptide according to 1), which is homologous to the amino acid sequence of IL-18. 3) The IL-18-like peptide according to 1) or 2) above, which has a three-dimensional structure similar to IL-18, and controls or regulates the interaction between Z or IL-18 and its receptor.

[0009] 4) The IL 18-like peptide according to 1) above, which is a polypeptide having a 9-residue or 12-residue amino acid sequence, or a 7-residue amino acid sequence having cysteine at both ends.

5) The IL 18-like peptide according to 1) above, which is a polypeptide having any amino acid sequence of SEQ ID NOs: 1 to 7 in the sequence listing.

6) A fusion of the IL 18-like peptide according to any one of 1) to 5) above and the gene 3 protein.

[0010] 7) A pharmaceutical group comprising an IL 18-like peptide according to any one of 1) to 5) above or a fusion of the IL 18-like peptide according to 6) above and a gene 3 protein as an active ingredient. Adult.

8) The therapeutic agent for immune diseases comprising the IL 18-like peptide according to any one of 1) to 5) above or a fusion of the IL 18-like peptide according to 6) and the gene 3 protein as an active ingredient .

[0011] 9) Screening a phage random peptide library using an anti-IL 18 antibody, and isolating a phage clone expressing an IL-18-like peptide capable of binding to the anti-IL-18 antibody. A method for producing an IL-18-like peptide capable of binding to an anti-IL-18 antibody.

The invention's effect

[0012] The IL-18-like peptide of the present invention can control and regulate the interaction of IL-18, which is closely involved in atopic dermatitis, with a receptor, and inhibits physiological activity. It is useful as an agent. Considering the action of IL 18 in immune abnormal diseases, the IL 18-like peptide of the present invention is applicable to various inflammatory diseases.

Brief Description of Drawings

[0013]

[FIG. 1] (1) and (2) in FIG. 1 show Example 1 using IL 18 antibody (hl8-108) and an antibody with unknown antigen specificity (scFv: ml8-92) as standards. The result of having confirmed the reactivity with the phage clone obtained by (1) is shown. BEST MODE FOR CARRYING OUT THE INVENTION

[0014]

In the present invention, an IL-18-like peptide that binds to an anti-IL-18 antibody refers to an amino acid sequence of IL-18 that binds to an anti-IL-18 antibody and is very similar in structure to IL-18. Have characteristics such as homology, control and regulation of the interaction between IL-18 and its receptor. Examples include peptides having an amino acid strength of 9 or 12 amino acids.

[0015] As a specific example of the IL 18-like peptide of the present invention, SEQ ID NO: 1 (hereinafter, the peptide comprising the amino acid sequence is also referred to as 108-2 based on the derived phage clone described later), Sequence listing SEQ ID NO: 2 (hereinafter, the peptide consisting of the amino acid sequence is also referred to as 108-4 based on the phage phage clone described below), Sequence Listing SEQ ID NO: 3 (hereinafter, the peptide consisting of the amino acid sequence is described below) (Also referred to as 108-5 on the basis of the derived phage clone), SEQ ID NO: 4 (hereinafter, the peptide comprising the amino acid sequence is also referred to as 108-13 on the basis of the derived phage clone described below), SEQ ID NO: 5 (hereinafter, the peptide having the amino acid sequence ability is also referred to as 108-C7 based on the phage phage clone described below), SEQ ID NO: 6 (hereinafter referred to as the amino acid sequence). Is also referred to as 108-C25 based on the derived phage clone described later), and SEQ ID NO: 7 (hereinafter, the peptide consisting of the amino acid sequence is also referred to as 108-C27 based on the derived phage clone described later). Peptide molecules having amino acid sequence ability such as. Each symbol in each amino acid sequence is as defined above.

[0016] The use form of the IL-18-like peptide of the present invention is not particularly limited as long as it retains the three-dimensional structure necessary for binding to the anti-IL-18 antibody. Can also be used in the form of a peptide alone, which can be prepared by a force incorporated in some protein or by peptide synthesis or the like. More specifically, it is an embodiment in which the aforementioned 108-2, 108-4, 108-5, 108-13, 108-C7, 108-C25, 108-C27 and these peptides are incorporated into the gene 3 protein. G3P bodies (also referred to as 108 — 2—G3P, 108—4—G3P, 108—5—G3P, 108—13—G3P, 108—C7—G3 P, 108—C25—G3P, 108—C27—G3P, respectively) Etc. The peptide May be used in a state in which the three-dimensional structure is retained based on the respective amino acid sequence information, and the amino acid sequence may be deleted, substituted, added or modified as long as the three-dimensional structure is retained.

[0017] The IL-18-like peptide of the present invention is, for example, an antibody (preferably a monoclonal antibody) that prevents binding between molecules involved in the IL-18 receptor system and recognizes the three-dimensional structure of IL-18. ) is used to present the peptide molecule having at least 9 or more § amino acids arranged in 10 ⁸ or more random, the phage random peptide library disk Lee - ing, selected phage clones, phage clones selected Can be obtained through a series of steps such as recovery of the product and purification of the product. The IL-18-like peptide of the present invention can be used not only in a purified state but also as a phage on which the peptide is presented.

[0018] The anti-IL-18 antibody used in the present invention may be an antibody of any animal species as long as it has an ability to inhibit binding between molecules involved in the IL-18 receptor system. And isotypes are not limited!

Here, the antibody recognizing conformational epitope means an antibody recognizing the three-dimensional structure of the epitope, that is, an antibody in which the antigen-binding region of the antibody is a saddle shape of the counterpart three-dimensional structure. . Specifically, it is an antibody that can bind if the antigen is in its native state, such as an immunoprecipitation reaction, but cannot bind when the antigen is in a denatured state. For example, Western blotting in a reduced state Examples include antibodies that cannot bind to antigens and cannot bind to linear peptides synthesized based on amino acid sequences of antibodies or epitopes.

[0019] Specific examples of the anti-IL-18 antibody used in the present invention include human anti-human IL-18 monoclonal antibody (hl8-108) (Japanese Patent Application No. 2003-125948).

The hl8-108 is a neutralizing antibody that inhibits the binding between IL-18 and its receptor, and connects the Fv domain of a human-derived antibody with a linker peptide! /, A single-chain Fv (scFv) antibody (Tsubakimoto Patent Application 2003-125948).

[0020] The human anti-human IL-18 antibody and the antibody fragment can be prepared by the following method according to the method described in Japanese Patent Application No. 2003-125948, for example. MRNA is extracted from peripheral blood B lymphocytes of healthy individuals, and the VH and VL chains of immunoglobulin genes are amplified by RT-PCR using primer pairs that define both ends, and H chains with various sequences Obtain the V chain population of the L chain. Next, the DNA that encodes a portion of the peptide linker, and a primer pair that defines both ends to be linked to the H chain and L chain, respectively, are amplified in combination and amplified in the V region of the H chain and L chain. Prepare diverse scFv DNA populations in random combinations. The obtained scFvDNA is incorporated into the phagemid vector pCA NTAB5E to prepare an scFv display phage library. This library is reacted with IL-18 immobilized on a plastic tube, and unreacted scFv display phage is removed by washing. Then, scFv phage clones bound to IL-18 are eluted with acid. ScFv DNA is prepared from the isolated phage clone, incorporated into an expression vector, and a host transformed with the expression vector is cultured according to a conventional method to obtain only the target scFv protein.

[0021] As an expression method of scFv DNA, for example, it can be expressed in E. coli. In the case of Escherichia coli, it can be expressed by functionally binding scFv to be expressed, such as a commonly used useful promoter and a signal sequence for antibody secretion. For example, examples of the promoter include lacZ promoter and araB promoter. As a signal sequence for secretion of scFv, a pelB signal sequence (Lei, SP "et al. J. BacterioL, 1987, 169: 4379-4383) may be used when expressed in the periplasm of E. coli. The signal sequence of the g13 protein of M13 phage can also be used for secretion during cleansing.

[0022] The scFv expressed as described above can be purified to homogeneity by separating the inside and outside of the cell and the host force. Since the scFv expressed here has an E tag sequence added to its C-terminus, it can be easily purified in a short time using affinity chromatography using an anti-E tag antibody. In addition, it is possible to purify by combining the separation and purification methods used in normal proteins. For example, antibodies can be separated and purified by combining column chromatography such as ultrafiltration, salting out, gel filtration, Z ion exchange, and Z hydrophobic chromatography.

[0023] Next, the phage random peptide library used in the production of the IL-18-like peptide of the present invention. As long as a peptide molecule having a random amino acid sequence is displayed on the phage surface, the library can be constructed of any kind of phage vector. Specifically, a known method (science 249, 386-390, published 1990. Mol. Biol., 248, ρ58-78, 1995. Proc. Natl. Acad. Sci. USA, 90, 10638-42 and 10643-47, 1993. Published by Nichiya, Nanotechnology, 16 卷, pp. 278-270, 1998). Examples of the gene inserted to be displayed on the phage surface include those encoding a peptide molecule consisting of 9 or 12 amino acids or 7 amino acids having cystine at both ends. Furthermore, the peptide molecule having a random amino acid sequence may be inserted into any phage constituent protein as long as it is displayed on the surface of the phage. For example, those inserted into gene 3 protein (see, for example, Japanese Patent Application No. 2000-297098) or gene 8 protein are preferred. In the case of gene 3 protein, if 9 or 12 peptide molecules with random amino acid force are inserted between the N-terminal Ala and the spacer Gly-Gly-Gly-Ser It is possible to minimize the influence of amino acid sequences extending at both ends of the generated peptide molecule on the tertiary structure of the inserted peptide molecule.

[0024] The IL-18-like peptide production method of the present invention starts by screening a fuzzy random peptide library using the aforementioned anti-IL-18 antibody. As a screening method, a general panning method in which an antibody is immobilized on a plate and a column column method in which an antibody is immobilized can be used. Also non-ung? Phage recovery? By repeating the phage growth-panning process twice or more, the target phage clone can be concentrated. By such a screening method, phage clones displaying peptides that bind to anti-IL-18 antibody on the surface thereof can be enriched. Usually, the phage clones thus obtained can be classified into several clone populations depending on the difference in binding force with the antibody and the kind of amino acid sequence retained. The nucleotide sequence analysis of the gene of the obtained phage clone can be easily performed by a conventional method such as a dideoxy method.

[0025] The IL-18-like peptide of the present invention can negatively control IL-18. late In addition, it is useful as an inhibitor of IL-18 activity, and for the prevention and treatment of inflammation such as atopic dermatitis and immune abnormalities.

The pharmaceutical composition of the present invention and the IL-18 activity inhibitor comprise the IL-18-like peptide of the present invention, which is an active ingredient, if necessary, as appropriate pharmaceutically acceptable additives (eg, carrier, excipient) Mixed with pharmacologically necessary ingredients such as pills, diluents, etc., and applied to the skin as an ointment, orally in the form of powder, granule, tablet, capsule, etc., or in the form of injection It can be administered parenterally. The dose can be increased or decreased depending on the type of active ingredient (type of peptide), route of administration, patient condition and disease type, body weight, age, sex, etc. Generally, it is the amount of peptide per day for adults. Apply 001-1000mg, once or several times a day, or it is desirable to administer.

[0026] In order to describe the present invention in more detail, reference examples, examples and experimental examples are listed below. The present invention is not limited to these.

Reference example 1

(Construction of phage library from healthy people)

The phage library was constructed using peripheral blood lymphocytes from 20 healthy volunteers as a starting material with reference to the method reported by JD Marks et al. (J. Mol. Biol, 222: 581-597, 1991). ,It was constructed. The constructed sub-library of νΗ (γ) —VK, νΗ (γ) —V, VH), VK, VH) —V are 1.1 Χ 10 ⁶ , 2.1 Χ 10 ⁸ , 8.4, respectively. Χ

5.3 Χ 10 ⁷ It was evaluated as having diversity of clones.

[0027] Reference Example 2

(Panjung)

Human IL-1 18 was dissolved in 0. 1M NaHCO lmL and placed in a 35mm dish (Iwaki) at 4 ° C.

Three

-The reaction was fixed. After blocking with 0.5% gelatin ZPBS at 20 ° C for 2 hours, the plate was washed 6 times with 0.1% Tween20 PBS. To this was added 0.9 mL (lxl0 ¹² tu / mL) of an antibody phage library (single-chain antibody-displayed phage solution) derived from a healthy person, and allowed to react.

[0028] 0. l% Tween20—After washing 10 times with PBS, 1. OmL of glycine buffer (pH 2.2) was added to elute single-chain antibody-displayed phages that bind to IL-18. Eluted fur After adjusting pH by adding 1M Tris (hydroxymethyl) aminomethane-HCL, pH9.1, E. coli was infected with E. coli TGI in the logarithmic growth phase. TG1 after infection is centrifuged at 3000 x g for 10 min. The supernatant is removed, suspended in 200 L of 2 x YT medium, and suspended in a SOBAG plate (2 o / o glucose, 100 / z gZmL ampicillin-containing SOB Plate) and cultured in an incubator at 30 ° C. The resulting colonies were suspended and recovered in an appropriate amount of 2 X YT medium using a Cost ar.

[0029] 50 μL of this TG1 solution was planted in 30 mL of 2Χ YTAG medium, rescued using helper phage, and a phage library after screening was prepared. Healthy person-derived fuzzy library 11 (7) — ¥ ¥ 11 (7) — ¥ e, ¥ 11)-¥ ¥ 11)-¥, for each, use the aforementioned IL-18 fixed plate to measure panning I went around. After the fourth round of panning, clones were arbitrarily extracted from the SOBAG plate, confirmed for scFv expression, confirmed for specificity by IL-18 ELISA, and analyzed for nucleotide sequence. Reference Example 3

(Screening IL—18 ELISA)

ELISA for screening isolated clones was performed as follows. HI-18 and human MIP-1α were immobilized on an ELISA plate and used for screening. Place 2 μgZmL human IL-18 or human ΜΙΡ-1α 2.5 gZmL human serum albumin (HS A) in a 40 μL / well ELISA plate (Nune) and let stand at 4 ° C for 16 hours. Immobilized. P BSAOO / z LZwell containing 0.5% BSA, 0.5% gelatin and 5 ° C skim milk was placed in an ELISA plate and allowed to stand at 4 ° C for 2 hours for blocking.

[0030] After the sample solution containing the scFv-displayed phage was reacted in 40 μL Zwell, the sample solution was discarded and washed 5 times with a washing solution. It was reacted with an anti-Ml3 monoclonal antibody (Pharmacia biotech) labeled with piotin and reacted with an anti-mouse IgG antibody labeled with alkaline phosphatase (AP). After washing with washing solution 5 times, add 50 μL LZwell of chromogenic substrate solution [PBS solution containing (lg / mL p-nitrophenyl phoophato) (Wako), 10% diethanolamine (Wako)] Color was developed at 37 ° C for 5-10 minutes. Multi Plate Auto Reader Ni—2001 (Inte r Med), the absorbance at 405 nm was measured. As a result, it was confirmed that all the clones evaluated were specific to the force IL-18.

[0031] Reference Example 4

(Expression and purification of human anti-IL 18 scFv)

Plasmid DNA was recovered from scFv clone # 10 reacting with HI-18 isolated in Reference Examples 2 and 3, and Escherichia coli HB1251 was transformed according to a conventional method. These E. coli cells are pre-cultured overnight in 2 X YT medium containing 2% glucose, and then partially transferred to glucose-free 2 X YT medium, and added to a final concentration of ImL IPTG and further cultured overnight to induce scFv expression. went. After completion of the culture, the cells were collected by centrifugation, suspended in PBS containing ImL EDTA, and left on ice for 30 minutes. Next, the mixture was centrifuged at 8,900 × g for 30 minutes, and the supernatant was collected, filtered through a 0.45 μm filter, and used as a starting material for scFv purification with a periplasmic fraction.

[0032] The purification starting material thus prepared was purified by affinity chromatography using an anti-Etag antibody according to a conventional method. After dialysis with PBS, endotoxin was removed with Dotoxi-gel (PIERCE) according to the attached protocol. After concentration with Centricon (Amicon) with a molecular weight cut of 10,000, it was filtered through a 0.45 μm filter to obtain a purified sample.

[0033] Example 1

(1) A commercially available phage 'display' library (NEB; Ph. D.-C7C and Ph. D. -12) was used. Micropanning was performed according to the report of Fukumoto et al. (Nichiya's Biotechnology, 16 卷, pp. 278-270, 1998).

Subsequently, the phage random peptide library was pan-ung using the anti-human IL-18 monoclonal antibody (hl8-108) obtained according to the above Reference Example. The anti-IL-18 antibody was used as a cage for selecting phages. The library was screened to isolate phage clones that bind to anti-IL-18 antibody.

Anti-IL-18 antibody coated with 10 g of Imunotube (Nunk) was reacted with the previously prepared phage library [5 X 10 ¹² Transducing Unit (TU)] at room temperature for 1 hour (Panjung ). Unbound phages were washed with Tris-HCl buffered saline containing 0.5% Tween 20 (TBS; containing 50 mM Tris-HCl, 0.15 M sodium chloride, pH 7. After washing in 5), phages bound to various anti-IL-18 antibodies were eluted using 0.1 IN hydrochloric acid-glycine buffer (pH 2.2). After recovery, the solution was neutralized with 1M Tris-HCl (pH 9.1). The resulting phages were infected and propagated in E. coli ER2738. The amount of various anti-IL-18 antibodies used for panning was reduced to 5 μg, and the same procedure was repeated to select phage clones that bind more strongly and specifically to anti-IL-18 antibodies. Panjung was performed separately using the two types of libraries described above.

[0035] Phage clones obtained in each panning were subjected to primary screening by ELISA! An ELISA plate (manufactured by NUNC) coated with anti-E-tag antibody 80ngZ40 1Z hole was coated with 0.5% BSA, and then reacted with anti-IL-18 antibody 100η ₈ Ζ80 / ζ lZ hole. 0.1. Wash plates with PBS containing 1% Tween 20 and various phage clones 8 X 10 ¹ . Virion Z40 μ1 well and piotinized anti-M13 monoclonal antibody (manufactured by Pharmacia) were added. Alkaline phosphatase (AP) -labeled streptavidin (Vector Lab Co., Ltd.) diluted 1: 10,000 was reacted at room temperature for 1 hour. After washing, the substrate ρ-nitrophenol-sodium phosphate hexahydrate (Wako Pure Chemical Industries, Ltd.) Was added, and the absorbance at 405 nm was measured. Seven clones specifically bound with anti-IL18 antibody.

[0036] (2) Sequence analysis of DNA inserted into phage selected by panning Gene 3 DNA sequences of various peptides incorporated into protein were analyzed using DNA Sequencer (Applied Biosystems) . As a primer, a synthetic oligonucleotide having the base sequence of SEQ ID NO: 8 was used.

As a result, all of the peptides inserted between the N-terminal Ala of the amino acid sequence of gene 3 protein and the spacer Gly-Gly-Gly-Ser have different amino acid sequences. It was able to be classified into 7 types shown by No. 1, Sequence Listing SEQ ID No. 2, Sequence Listing Sequence No. 3, Sequence Listing Sequence No. 4, Sequence Listing Sequence No. 5, Sequence Listing Sequence No. 6, Sequence Listing Sequence No. 7.

[0037] A clear common sequence was conserved among the amino acid sequences of the seven types of peptides identified. Furthermore, a homology search between these peptides and IL-18 was performed using CLUSTAL W ver. 3.1. As a result, the primary sequence showed remarkable similarity.

Experimental example 1 Reactivity with the phage clone obtained in Example 1 was confirmed using an anti-IL 18 antibody (hl8-108) and an antibody with unknown antigen specificity (scFv: ml8-92) as a standard. The analysis was performed by the ELISA method of Example 1. As a result, all clones specifically bound to anti-IL-18 antibody and did not react with ml8-92 antibody. The results are shown in Figure 1.

INDUSTRIAL APPLICABILITY The IL-18-like peptide of the present invention is useful as an IL-18 activity inhibitor and for the prevention and treatment of inflammatory diseases such as atopic dermatitis, immune abnormal diseases and the like.

(Sequence table free text)

Sequence Listing SEQ ID NO: 8: Oligonucleotide designed to act as a sequencer primer.

Claims

The scope of the claims

[1] An IL-18-like peptide that can bind to an anti-IL-18 antibody that inhibits the IL-18 signal.

[2] The IL 18-like peptide according to claim 1, which is homologous to the amino acid sequence of IL-18.

[3] The IL-18-like peptide according to claim 1 or 2, wherein the IL-18-like peptide has a three-dimensional structure similar to that of IL-18, and controls or regulates the interaction between Z or IL-18 and its receptor.

[4] The IL18-like peptide according to claim 1, which is a polypeptide having a 9-residue amino acid sequence or a 12-residue amino acid sequence, or a 7-residue amino acid sequence having cysteine at both ends.

[5] The IL 18-like peptide according to claim 1, which is a polypeptide having any one of the amino acid sequences of SEQ ID NOs: (1) to (7) in the sequence listing.

[6] A fusion product of the IL-18-like peptide according to any one of claims 1 to 5 and the gene 3 protein.

[7] IL 18-like peptide according to any one of claims 1 to 5, or IL according to claim 6.

A pharmaceutical composition comprising a fusion of an 18-like peptide and gene 3 protein as an active ingredient.

[8] The IL 18-like peptide according to any one of claims 1 to 5, or the IL according to claim 6.

A therapeutic agent for immune diseases comprising a fusion of an 18-like peptide and gene 3 protein as an active ingredient.

[9] A phage random peptide library is screened using an anti-IL-18 antibody, and a phage clone expressing an IL-18-like peptide capable of binding to the anti-IL-18 antibody is isolated. A method for producing an IL 18-like peptide capable of binding to an anti-IL 18 antibody.