Anti-il-23 antibodies, genetically engineered products thereof, and uses thereof

Abstract

The application belongs to the field of antibodies, and particularly relates to anti-IL-23 antibodies and genetically engineered products and applications thereof. The anti-IL-23 antibody is a human-mouse chimeric antibody, the amino acid sequence of the heavy chain CDR region is shown as SEQ ID NO. 1-SEQ ID NO. 3, the amino acid sequence of the light chain CDR region is shown as SEQ ID NO. 4-SEQ ID NO. 6, the amino acid sequence of the heavy chain variable region is shown as SEQ ID NO. 7, and the amino acid sequence of the light chain variable region is shown as SEQ ID NO. 8. The anti-IL-23 antibody provided in the application has high affinity, strong specificity, is directed against the p19 subunit, and has excellent blocking effect. The application provides technical support for the prevention, adjuvant treatment or treatment of autoimmune and chronic inflammatory diseases.

Description

Technical Field

[0001] This invention belongs to the field of antibodies, specifically relating to anti-IL-23 antibodies and their genetically engineered products and applications. Background Technology

[0002] IL-23 (interleukin-23) is a cytokine secreted by activated dendritic cells, macrophages, monocytes, and neutrophils, belonging to the IL-12 family. It consists of p19 and the 40kD subunit of IL-12, linked by disulfide bonds to form a heterodimeric cytokine, which promotes the proliferation of CD45RO+ memory T cells and the production of gamma interferon.

[0003] Mouse models, genetic studies in patients, and clinical trials have all demonstrated that IL-23 is a major mediator in autoimmune and chronic inflammatory diseases, including inflammatory bowel disease, psoriasis, rheumatoid arthritis, and multiple sclerosis. Blocking the IL-23 signaling pathway has become an effective strategy for treating these diseases, as it can alleviate symptoms and improve patients' quality of life by reducing the production of pro-inflammatory cytokines. Numerous experiments have confirmed that IL-23 stimulates IL-17 secretion and proliferation, leading to inflammatory responses and participation in immune responses; the IL-23 / IL-17 inflammatory axis plays a crucial role in the occurrence and development of various autoimmune diseases and some kidney diseases.

[0004] Anti-IL-23 antibodies are specific antibody drugs designed to target the cytokine interleukin-23 (IL-23), primarily used to treat various autoimmune and inflammatory diseases. Anti-IL-23 antibodies selectively block the IL-23 pathway without affecting other cytokines such as IL-12, reducing off-target effects. Furthermore, anti-IL-23 antibodies exhibit good efficacy and safety, significantly improving patients' disease symptoms and quality of life.

[0005] For IL-23-mediated diseases, several drugs targeting IL-23 have been developed and are used in clinical treatment. These drugs inhibit inflammatory responses and disease progression by blocking the IL-23 signaling pathway, providing patients with new treatment options. However, due to the complexity and diversity of these diseases, there is still a need to develop anti-IL-23 antibodies with better blocking effects. Summary of the Invention

[0006] To address the aforementioned problems, this invention provides a novel anti-IL-23 antibody that exhibits higher affinity, better blocking effect, and higher safety.

[0007] On one hand, the present invention provides an anti-IL-23 antibody, comprising a heavy chain and a light chain, wherein the heavy chain includes a heavy chain variable region, and the light chain includes a light chain variable region; the heavy chain variable region includes HCDR1, HCDR2, and HCDR3, and the light chain variable region includes LCDR1, LCDR2, and LCDR3, wherein:

[0008] (1) HCDR1 is the amino acid sequence shown in SEQ ID NO.1;

[0009] (2) HCDR2 is the amino acid sequence shown in SEQ ID NO.2;

[0010] (3) HCDR3 is the amino acid sequence shown in SEQ ID NO.3;

[0011] (4) LCDR1 is the amino acid sequence shown in SEQ ID NO.4;

[0012] (5) LCDR2 is the amino acid sequence shown in SEQ ID NO.5;

[0013] (6) LCDR3 is the amino acid sequence shown in SEQ ID NO.6.

[0014] Specifically, the amino acid sequence of the heavy chain variable region can be SEQ ID NO.7 or a sequence having more than 80% sequence similarity to SEQ ID NO.7; the amino acid sequence of the light chain variable region can be SEQ ID NO.8 or a sequence having more than 80% sequence similarity to SEQ ID NO.8.

[0015] More specifically, the amino acid sequence of the heavy chain variable region can be SEQ ID NO.7 or a sequence having more than 90% sequence similarity to SEQ ID NO.7; the amino acid sequence of the light chain variable region can be SEQ ID NO.8 or a sequence having more than 90% sequence similarity to SEQ ID NO.8.

[0016] Preferably, the amino acid sequence of the heavy chain variable region can be SEQ ID NO.7 or a sequence having more than 95% sequence similarity to SEQ ID NO.7; the amino acid sequence of the light chain variable region can be SEQ ID NO.8 or a sequence having more than 95% sequence similarity to SEQ ID NO.8.

[0017] More preferably, the amino acid sequence of the heavy chain variable region can be SEQ ID NO.7 or a sequence having more than 98% sequence similarity to SEQ ID NO.7; the amino acid sequence of the light chain variable region can be SEQ ID NO.8 or a sequence having more than 98% sequence similarity to SEQ ID NO.8.

[0018] More preferably, the amino acid sequence of the heavy chain variable region may be SEQ ID NO.7; and the amino acid sequence of the light chain variable region may be SEQ ID NO.8.

[0019] Specifically, the heavy chain amino acid sequence may be SEQ ID NO.9 or a sequence having more than 65% sequence similarity to SEQ ID NO.9; the light chain amino acid sequence may be SEQ ID NO.10 or a sequence having more than 65% sequence similarity to SEQ ID NO.10.

[0020] More specifically, the heavy chain amino acid sequence may be SEQ ID NO.9 or a sequence having more than 80% sequence similarity to SEQ ID NO.9; the light chain amino acid sequence may be SEQ ID NO.10 or a sequence having more than 80% sequence similarity to SEQ ID NO.10.

[0021] Preferably, the heavy chain amino acid sequence can be SEQ ID NO.9 or a sequence having more than 95% sequence similarity to SEQ ID NO.9; the light chain amino acid sequence can be SEQ ID NO.10 or a sequence having more than 95% sequence similarity to SEQ ID NO.10.

[0022] More preferably, the heavy chain amino acid sequence may be SEQ ID NO.9 or a sequence having more than 99% sequence similarity to SEQ ID NO.9; the light chain amino acid sequence may be SEQ ID NO.10 or a sequence having more than 99% sequence similarity to SEQ ID NO.10.

[0023] More preferably, the heavy chain amino acid sequence may be SEQ ID NO.9; the light chain amino acid sequence may be SEQ ID NO.10.

[0024] Specifically, the anti-IL-23 antibody is a human-mouse chimeric antibody.

[0025] In another aspect, the present invention provides the aforementioned nucleotide sequence of the anti-IL-23 antibody, wherein the nucleotide sequence of the heavy chain may be SEQ ID NO.11 or a sequence having more than 60% sequence similarity to SEQ ID NO.11; and the nucleotide sequence of the light chain may be SEQ ID NO.12 or a sequence having more than 60% sequence similarity to SEQ ID NO.12.

[0026] Specifically, the nucleotide sequence of the heavy chain may be SEQ ID NO.11 or a sequence having more than 70% sequence similarity to SEQ ID NO.11; the nucleotide sequence of the light chain may be SEQ ID NO.12 or a sequence having more than 70% sequence similarity to SEQ ID NO.12.

[0027] More specifically, the nucleotide sequence of the heavy chain may be SEQ ID NO.11 or a sequence having more than 90% sequence similarity to SEQ ID NO.11; the nucleotide sequence of the light chain may be SEQ ID NO.12 or a sequence having more than 90% sequence similarity to SEQ ID NO.12.

[0028] Preferably, the nucleotide sequence of the heavy chain can be SEQ ID NO.11 or a sequence having more than 97% sequence similarity to SEQ ID NO.11; the nucleotide sequence of the light chain can be SEQ ID NO.12 or a sequence having more than 97% sequence similarity to SEQ ID NO.12.

[0029] More preferably, the nucleotide sequence of the heavy chain may be SEQ ID NO.11; and the nucleotide sequence of the light chain may be SEQ ID NO.12.

[0030] In another aspect, the present invention provides an expression vector comprising the aforementioned nucleotide sequence.

[0031] Specifically, the expression vector can be a plasmid, bacteriophage, or virus.

[0032] Preferably, the expression vector can be a plasmid.

[0033] In another aspect, the present invention provides a host cell comprising the aforementioned nucleotide sequence or expression vector.

[0034] Specifically, the host cells include, but are not limited to, prokaryotic cells or eukaryotic cells.

[0035] Preferably, the host cell can be a eukaryotic cell.

[0036] More preferably, the host cell may be a HEK293F cell.

[0037] In another aspect, the present invention provides a recombinant protein comprising the aforementioned anti-IL-23 antibody; the recombinant protein further comprises a bioactive protein or functional fragment thereof that assists in its expression and / or secretion, or prolongs its half-life in vivo.

[0038] Specifically, the bioactive protein or its functional fragment is selected from at least one of the following: immunoglobulin Fc domain, serum albumin, albumin-binding polypeptide, prealbumin, carboxyl-terminal peptide, elastin-like polypeptide, His tag, GST tag, MBP tag, FLAG tag, and SUMO tag.

[0039] In another aspect, the present invention provides an antibody formulation comprising the aforementioned anti-IL-23 antibody or recombinant protein.

[0040] In another aspect, the present invention provides a drug conjugate comprising the following (a) and (b):

[0041] (a) The aforementioned anti-IL-23 antibody or recombinant protein;

[0042] (b) The coupled part that combines with (a).

[0043] In another aspect, the present invention provides a pharmaceutical composition comprising the aforementioned anti-IL-23 antibody or recombinant protein or antibody preparation or pharmaceutical conjugate.

[0044] In another aspect, the present invention provides a kit comprising the aforementioned anti-IL-23 antibody or recombinant protein or antibody preparation or drug conjugate.

[0045] Specifically, the kit also includes a container for loading the antibody preparation.

[0046] In another aspect, the present invention provides a method for in vitro detection of IL-23 in samples for non-diagnostic purposes, the method comprising the following steps:

[0047] S1, including the aforementioned anti-IL-23 antibody, recombinant protein, antibody preparation, or drug conjugate, comes into contact with the sample to be tested;

[0048] S2, Detecting antigen-antibody complexes;

[0049] S3, Interpretation result.

[0050] In another aspect, the present invention provides the use of the aforementioned anti-IL-23 antibody, recombinant protein, antibody preparation, or drug conjugate, said use including at least one of the following:

[0051] (a) Preparation of reagents or kits for detecting IL-23;

[0052] (b) To prepare medicines for the prevention, adjunctive treatment and / or treatment of psoriasis, psoriatic arthritis, ulcerative colitis, Crohn's disease or plaque psoriasis;

[0053] (c) To prepare drugs for the prevention, adjuvant therapy and / or treatment of autoimmune diseases, tumors or viral infections.

[0054] The technical effects achieved by this invention are as follows:

[0055] (1) The antibody provided by the present invention can specifically bind to the Human IL23 p19 subunit.

[0056] (2) The antibody provided by the present invention has an affinity of 0.21 nM for Human IL23.

[0057] (3) The antibody provided by the present invention has a specific binding to monkey IL23 with an affinity of about 2.9 nM, but no obvious binding signal was detected with rat and mouse IL23.

[0058] (4) The antibody provided by the present invention can block the interaction between Human IL23 and Human IL23 receptor, and the blocking effect is obvious (the binding signal is reduced by about 98%). Attached Figure Description

[0059] Figure 1 This is a map of a 1pcDNA3.1 vector. Detailed Implementation

[0060] The present invention will be further described in detail below with reference to specific embodiments. The following embodiments are not intended to limit the present invention, but only to illustrate the present invention. Unless otherwise specified, the experimental methods used in the following embodiments are generally performed under conventional conditions. Unless otherwise specified, the materials and reagents used in the following embodiments are commercially available.

[0061] Example 1

[0062] The anti-IL-23 antibody provided by this invention is a human-mouse chimeric antibody that binds to human interleukin-23, named 19A1F1, and its specific sequence is as follows:

[0063] (1) Sequence of amino acids in the heavy chain complementarity-determining region (CDR region) (Kabat nomenclature system):

[0064] CDR area of ​​19A1F1:

[0065] SEQ ID No. 1: CDR1 amino acid sequence SDYAW;

[0066] SEQ ID No. 2: CDR2 amino acid sequence YIRYSGNTSYNPSLKS;

[0067] SEQ ID No. 3: CDR3 amino acid sequence AYAY.

[0068] (2) Sequence of amino acids in the light chain complementarity-determining region (CDR region) (Kabat nomenclature system):

[0069] CDR area of ​​19A1F1:

[0070] SEQ ID No. 4: CDR1 amino acid sequence RASESVDNYGISFMN;

[0071] SEQ ID No. 5: CDR2 amino acid sequence AASNQGS;

[0072] SEQ ID No. 6: CDR3 amino acid sequence QQSKEVPLT.

[0073] (3) Amino acid sequence of the heavy chain variable region:

[0074] 19A1F1:SEQ ID No. 7

[0075] DVQLQESGPGLVKPSQSLSLTCTVTGYSITSDYAWNWIRQFPGNKLEWM GYIRYSGNTSYNPSLKSRISITRDTSKNQFYLQLNSVTTEDTATYFCARAYAYW GQGTLVTVSA.

[0076] (4) Amino acid sequence of the light chain variable region:

[0077] 19A1F1:SEQ ID No. 8

[0078] DIVLIQSPTSLAVSLGQRATISCRASESVDNYGISFMNWFQQKPGQPPKLLI YAASNQGSGVPARFSGSGSGTDFSLNIHPMEEDDTAMYFCQQSKEVPLTFGAG TKLEIKR.

[0079] (5) Amino acid sequence of the heavy chain:

[0080] 19A1F1:SEQ ID No. 9

[0081] DVQLQESGPGLVKPSQSLSLTCTVTGYSITSDYAWNWIRQFPGNKLEWMGYIRYSGNTSYNPSLKSRISITRDTSKNQFYLQLNSVTTEDTATYFCARAYAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK。

[0082] (6) Amino acid sequence of the light chain:

[0083] 19A1F1: SEQ ID No.10

[0084] DIVLIQSPTSLAVSLGQRATISCRASESVDNYGISFMNWFQQKPGQPPKLLIYAASNQGSGVPARFSGSGSGTDFSLNIHPMEEDDTAMYFCQQSKEVPLTFGAGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC。

[0085] DNA sequence encoding the above amino acid sequence;

[0086] Heavy chain

[0087] 19A1F1: SEQ ID No.11

[0088]

[0089] Light chain

[0090] 19A1F1: SEQ ID No.12

[0091] GACATCGTGCTGATCCAGAGCCCCACCAGCCTGGCCGTGTCTTTAGGACAAAGGGCCACCATCAGCTGTAGGGCCAGCGAGAGCGTGGACAACTACGGCATCAGCTTCATGAACTGGTTCCAGCAGAAGCCCGGCCAGCCTCCCAAGCTGCTGATCTACGCTGCTTCCAACCAGGGATCTGGAGTTCCTGCTAGGTTCAGCGGCTCTGGCAGCGGAACCGACTTCAGCCTGAACATCCACCCCATGGAGGAGGACGACACCGCCATGTACTTCTGCCAGCAGAGCAAGGAGGTGCCCCTGACCTTCGGCGCTGGCACAAAGTTGGAGATCAAGAGGACTGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGT。

[0092] 1.1 Antigen preparation and immunization

[0093] (1) Preparation of human IL23 protein

[0094] To screen for the antigen hIL23A-hIL12B, human IL23A and human IL12B subunits were constructed into a pcDNA3.1(+) vector, with the two subunits covalently linked by a G4S linker and a His tag introduced at the N-terminus. For the immunogenic antigen hIL23-mIL12B, human IL23A and mouse IL12B subunits were constructed into a pcDNA3.1(+) vector, with the two subunits covalently linked by a G4S linker and a His tag introduced at the N-terminus. After transient transfection of HEK293F cells, the expression supernatant was collected, purified by metal ion chelation chromatography, and then concentrated by ultrafiltration. SDS-PAGE was used to verify protein purity.

[0095] (2) Animal Immunization

[0096] To obtain high-affinity and high-specificity antibodies against the human IL23A subunit, this experiment immunized several Balb / c mice with the chimeric protein hIL23A-mIL12B using a classic immunization schedule to induce an immune response. Mouse serum was collected, and serum titers were determined using ELISA with hIL23A-hIL12B protein as the antigen. Mice with high titers were selected for fusion.

[0097] 1.2 Hybridoma Fusion and Screening

[0098] Prior to fusion, the status of mouse myeloma cells SP2 / 0 was adjusted to ensure that their growth density did not exceed 1×10⁻⁶. 6 Cells / mL, animals were immunized 3 days in advance. Feeder cells were prepared one day in advance, with a plate density of 2.0 × 10⁶ cells / mL. 4 Cells / well. Fusion is performed using an electrofusion apparatus to ensure a spleen cell or lymph node cell to SP2 / 0 cell ratio between 10:1 and 5:1, with no more than 1 × 102 spleen cells seeded per well. 5 Seven days after fusion, the supernatant was harvested and the culture medium was replaced. The collected supernatant was then used to obtain one hybridoma cell line that showed positive results in all screening experiments using the ELISA method.

[0099] ELISA binding method:

[0100] Dilute human IL23 or human IL12 to 1 μg / mL with PBS (pH 7.4) to coat 96-well plates, adding 100 μL to each well. After blocking, add hybridoma culture supernatant and incubate. After washing, add horseradish peroxidase (HRP)-labeled affinity-purified goat anti-mouse IgG Fcγ fragment secondary antibody (Jackson Immunoresearch, Cat#115035062) and incubate. After washing, add chromogenic substrate and incubate. After stopping the reaction, read the absorbance at OD450.

[0101] 1.3 Preparation and activity identification of human-mouse chimeric antibodies

[0102] (1) Acquisition of mouse resistance genes

[0103] Total RNA was extracted from hybridoma cell lines using the Trizol (Ambion 15596-026) kit, and then analyzed using PrimeScript. TM II. 1st Strand cDNA Synthesis Kit (Takara, Cat#6210A) was used to reverse transcribe total RNA to prepare cDNA, and the heavy and light chain variable regions of the antibody were amplified using degenerate primers.

[0104] (2) Construction of human-mouse chimeric antibodies

[0105] The mouse anti-light chain variable region sequence was spliced ​​with the human IgG kappa CL region sequence to obtain the full-length light chain sequence, which was then constructed into a pcDNA3.1 vector containing a nitrogen-terminal signal peptide through whole-genome synthesis. Figure 1 Light chain expression vectors were obtained from )

[0106] The variable region sequence of the mouse anti-heavy chain was spliced ​​with the CH1, CH2 and CH3 region sequences of human IgG1 to obtain the full-length heavy chain sequence. The heavy chain expression vector was obtained by synthesizing the entire gene and constructing it into a pcDNA3.1 vector containing a nitrogen-terminal signal peptide.

[0107] (3) Expression and purification of human-mouse chimeric antibodies

[0108] Light and heavy chain expression vectors were used to transiently transfect HEK293F cells via co-transfection. After 7 days of culture, the supernatant was collected. The chimeric antibody in the supernatant was purified using a protein A affinity chromatography column and dialyzed into PBS, pH 7.4. The concentration was determined by the A280 method and analyzed by SDS-PAGE.

[0109] (4) Surface plasmon resonance assay to determine the binding specificity of human-mouse chimeric antibodies to human IL23.

[0110] The chimeric antibody was captured onto the surface of a Protein A chip (GE, Cat#29127556), and affinity sequencing was performed on the chip surface using 50 nM human IL23 or 50 nM human IL12 as analytical streams.

[0111] A complete affinity assay was performed using antibodies that specifically bind to human IL23. The chimeric antibody was captured onto the surface of a Protein A chip (GE, Cat#29127556), and affinity was measured by passing serially diluted human IL23 as the analytical stream through the chip surface.

[0112] 1.4 Competitive binding activity of the antibody against human IL23 and human IL23R

[0113] SPR method:

[0114] Human IL23R was immobilized onto the surface of a CM5 chip (GE, Cat#29149603) using an amino-immobilization method. The binding signal of 50 nM human IL23 flowing through the chip surface served as a positive control. The test sample was mixed with human IL23 and then flowed through the chip surface immobilized with human IL23R. The percentage decrease in the human IL23 binding signal was used as an indicator of whether the antibody had a blocking effect.

[0115] 1.5 Determination of antibody species cross-binding activity

[0116] The chimeric antibody to be tested was captured using a Protein A chip. Serially diluted mouse IL23, monkey IL23 and rat IL23 were used as analytical streams and passed through the chip surface immobilized with the antibody to determine the affinity between the antibody and IL23.

[0117] 1.6 Antibody inhibits the ability of IL23-stimulated mouse spleen cells to secrete IL17.

[0118] Single-cell suspensions of splenocytes from Balb / c mice were prepared and resuspended in RPMI-1640 complete medium (86% (v / v) RPMI-1640, 1% (v / v) non-essential amino acids, 1% (v / v) sodium pyruvate, 2.5 mM HEPES, 1% (v / v) L-glutamine, 0.00035% (wt) 2-mercaptoethanol, 1% (v / v) penicillin / streptomycin, 10% (v / v) heat-inactivated fetal bovine serum and 50 ng / mL human IL-2 (R and D Systems). Splenocytes were resuspended at 5 × 10⁶ cells / mL. 6 A cell suspension of 100 cells / mL was added to a 96-well cell culture plate at a rate of 100 μL / well.

[0119] After incubating serially diluted antibodies with 10 pM human IL23 at 37°C for 90 minutes, 100 μL of the solution was added to each well of a 96-well cell culture plate containing spleen cells. Wells containing only human IL23 without antibody served as positive controls. Cells were then returned to 37°C and cultured in 5% CO2 for 48–72 hours. The cell supernatant was collected and analyzed using a Mouse IL17A assay kit to calculate the IC50 value of the antibody.

[0120] 1.7 Results

[0121] 1.7.1 Binding of positive clones to human IL23 protein

[0122] One positive clone, 19A1F1, was obtained through screening. The ELISA results of this clone showed that it binds to Human IL23 but not to Human IL12. This clone is an antibody that specifically binds to the p19 subunit of Human IL23 (Table 1).

[0123] Table 1

[0124]

[0125]

[0126] 1.7.2 Affinity determination of positive clones with human IL23 and human IL12

[0127] The affinity of the positive clone 19A1F1 for specific binding was determined using SPR technology. The results showed that the affinity of this clone for Human IL23 was 0.21 nM, and there was no obvious specific binding signal with Human IL12 at a concentration of 50 nM (Table 2).

[0128] Table 2

[0129]

[0130] 1.7.3 Affinity of positive clones to mouse IL23, rat IL23 and monkey IL23

[0131] The species binding ability of 19A1F1 was identified. SPR affinity assay showed that the clone specifically binds to monkey IL23 with an affinity of approximately 2.9 nM, but no obvious binding signal was detected with rat and mouse IL23 (Table 3).

[0132] Table 3

[0133]

[0134] 1.7.4 Neutralization of IL23 activity by positive clones

[0135] The results are shown in Table 4:

[0136] Table 4

[0137]

[0138] SPR results showed that 19A1F1 could block the interaction between Human IL23 and its receptor, and the blocking effect was significant (the binding signal was reduced by approximately 98%) (Table 4). Simultaneously, the IL17A secretion assay showed that in the presence of 19A1F1, the ability of IL23 to stimulate mouse spleen cells to secrete IL17A was significantly reduced, with an IC50 value of 1.3 μg / mL (Table 4).

Claims

1. An anti-IL-23 antibody, characterized in that, It includes a heavy chain and a light chain, wherein the heavy chain includes a heavy chain variable region, and the light chain includes a light chain variable region; the heavy chain variable region includes HCDR1, HCDR2, and HCDR3, and the light chain variable region includes LCDR1, LCDR2, and LCDR3, wherein: (1) HCDR1 is the amino acid sequence shown in SEQ ID NO.1; (2) HCDR2 is the amino acid sequence shown in SEQ ID NO.2; (3) HCDR3 is the amino acid sequence shown in SEQ ID NO.3; (4) LCDR1 is the amino acid sequence shown in SEQ ID NO.4; (5) LCDR2 is the amino acid sequence shown in SEQ ID NO.5; (6) LCDR3 is the amino acid sequence shown in SEQ ID NO.

6.

2. The anti-IL-23 antibody according to claim 1, characterized in that, The amino acid sequence of the heavy chain variable region is SEQ ID NO.7 or a sequence having more than 80% sequence similarity to SEQ ID NO.7; the amino acid sequence of the light chain variable region is SEQ ID NO.8 or a sequence having more than 80% sequence similarity to SEQ ID NO.

8.

3. The anti-IL-23 antibody according to claim 2, characterized in that, The amino acid sequence of the heavy chain is SEQ ID NO.9 or a sequence having more than 65% sequence similarity to SEQ ID NO.9; the amino acid sequence of the light chain is SEQ ID NO.10 or a sequence having more than 65% sequence similarity to SEQ ID NO.

10.

4. The anti-IL-23 antibody according to any one of claims 1-3, characterized in that, The anti-IL-23 antibody is a human-mouse chimeric antibody.

5. A nucleotide sequence expressing the anti-IL-23 antibody according to any one of claims 1-4, characterized in that, The nucleotide sequence of the heavy chain is SEQ ID NO.11 or a sequence having more than 60% sequence similarity to SEQ ID NO.11; the nucleotide sequence of the light chain is SEQ ID NO.12 or a sequence having more than 60% sequence similarity to SEQ ID NO.

12.

6. An expression carrier, characterized in that, Includes the nucleotide sequence described in claim 5.

7. The carrier according to claim 6, characterized in that, The expression vector is a plasmid, bacteriophage, or virus.

8. A host cell, characterized in that, Includes the nucleotide sequence of claim 5 or the expression vector of claim 6 or 7.

9. The host cell according to claim 8, characterized in that, The host cell is a prokaryotic cell or a eukaryotic cell.

10. A recombinant protein, characterized in that, The recombinant protein comprises the anti-IL-23 antibody as described in any one of claims 1-4; the recombinant protein further comprises a bioactive protein or a functional fragment thereof that assists in its expression and / or secretion, or prolongs its half-life in vivo.

11. The recombinant protein according to claim 10, characterized in that, The bioactive protein or its functional fragment is selected from at least one of the following: immunoglobulin Fc domain, serum albumin, albumin-binding peptide, prealbumin, carboxyl-terminal peptide, elastin-like peptide, His tag, GST tag, MBP tag, FLAG tag, and SUMO tag.

12. An antibody preparation, characterized in that, Includes the anti-IL-23 antibody according to any one of claims 1-4 or the recombinant protein according to claim 10 or 11.

13. A drug conjugate, characterized in that, Including the following (a) and (b): (a) The anti-IL-23 antibody according to any one of claims 1-4 or the recombinant protein according to claim 10 or 11; (b) The coupled part that combines with (a).

14. A pharmaceutical composition, characterized in that, Includes the anti-IL-23 antibody according to any one of claims 1-4, the recombinant protein according to claim 10 or 11, the antibody preparation according to claim 12, or the drug conjugate according to claim 13.

15. A reagent kit, characterized in that, Includes the anti-IL-23 antibody according to any one of claims 1-4, the recombinant protein according to claim 10 or 11, the antibody preparation according to claim 12, or the drug conjugate according to claim 13.

16. A method for in vitro detection of IL-23 in samples for non-diagnostic purposes, characterized in that, The method includes the following steps: S1. The anti-IL-23 antibody according to any one of claims 1-4, the recombinant protein according to claim 10 or 11, the antibody preparation according to claim 12, or the drug conjugate according to claim 13 is brought into contact with the sample to be tested; S2, Detecting antigen-antibody complexes; S3, Interpretation result.

17. Use of the anti-IL-23 antibody according to any one of claims 1-4, or the recombinant protein according to claim 10 or 11, or the antibody preparation according to claim 12, or the drug conjugate according to claim 13, wherein the use comprises at least one of the following: (a) Preparation of reagents or kits for detecting IL-23; (b) To prepare medicines for the prevention, adjunctive treatment and / or treatment of psoriasis, psoriatic arthritis, ulcerative colitis, Crohn's disease or plaque psoriasis; (c) To prepare drugs for the prevention, adjuvant therapy and / or treatment of autoimmune diseases, tumors or viral infections.

Images