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Anti-human ccr1 monoclonal antibody

A monoclonal antibody and antibody technology, applied in the direction of antibodies, anti-animal/human immunoglobulins, anti-inflammatory agents, etc., can solve problems such as failure to show effectiveness

Pending Publication Date: 2020-04-03
KYOWA HAKKO KIRIN CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For these low-molecular inhibitors, clinical trials have been conducted on patients with autoimmune or inflammatory diseases such as rheumatoid arthritis, multiple sclerosis, and chronic obstructive pulmonary disease, but none of them have shown efficacy (Non-Patent Document 14 )

Method used

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  • Anti-human ccr1 monoclonal antibody
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  • Anti-human ccr1 monoclonal antibody

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0316] (1) Antigen preparation

[0317] Human CCR1 or human CCR1-expressing cells as an antigen can be obtained by introducing an expression vector containing cDNA encoding full-length or partial-length human CCR1 into Escherichia coli, yeast, insect cells, animal cells, or the like. In addition, human CCR1 can also be obtained by purifying human CCR1 from various human cell lines, human cells, human tissues, etc. that express human CCR1 in large amounts. In addition, these human cell lines, human cells, human tissues, etc. can also be used as antigens as they are. In addition, a synthetic peptide having a partial sequence of human CCR1 can also be prepared by a chemical synthesis method such as the Fmoc method or the tBoc method, and used as an antigen. A known tag such as FLAG or His may be added to the C-terminus or N-terminus of human CCR1 or a synthetic peptide having a partial sequence of human CCR1.

[0318] Human CCR1 used in the present invention can be obtained by ...

Embodiment 1

[0462] [Example 1] Production of expression vectors for human and mouse CCR1

[0463] (1) Preparation of each CCR1 gene

[0464] DNAs encoding human or mouse CCR1 or CCR1-CCR3 chimeric receptors (GenScript Japan, Inc.) of the following 1 to 7 were synthesized. Restriction enzyme sites (BamHI and NotI) and Kozak sequences for integration into each vector were added at the time of synthesis.

[0465] 1. cDNA sequence (sequence number 1) encoding human CCR1 (hereinafter referred to as hCCR1)

[0466] 2. The cDNA sequence (sequence number 3) of coding mouse CCR1 (hereinafter referred to as mCCR1)

[0467] 3. cDNA sequence (SEQ ID NO: 5) encoding human CCR3 (hereinafter referred to as hCCR3)

[0468] 4. A cDNA sequence encoding a chimeric receptor (hereinafter referred to as NC3-hCCR1) obtained by substituting the amino acid sequence at positions 1 to 31 of human CCR1 with the corresponding N-terminal amino acid sequence of human CCR3 (SEQ ID NO: 6 )

[0469] 5...

Embodiment 2

[0478] [Example 2] Production of CCR1-expressing cell lines

[0479] (1) Production of hCCR1 expressing cells

[0480] hCCR1 / Tn-pMug-Hygro and the Tol2 transposase expression vector TPEX_pMug (International Publication No. 2013 / 005649) as the plasmid DNA prepared in Example 1 were co-introduced into CHO-S (Thermo Fisher Scientific) to prepare expressing cell lines. Gene transfer was carried out using Fugene HD (Promega) as follows. Prepare 1 × 10 inoculum in a 6-well plate 5 Cells / mL were 2.5 mL each, and a mixture of hCCR1 / Tn-pMug-Hygro, TPEX_pMug, and Fugene HD was added to the culture medium 24 hours later. 72 hours after the addition, 1 mg / mL of hygromycin (Invitrogen) was added, and drug selection was performed for about 2 weeks. The cells that had acquired drug resistance were recovered, and expression analysis was performed by flow cytometry (FACS Calibur, BD Biosciences). As a result, the expression of the introduced hCCR1 was confirmed. This cell line is r...

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Abstract

The present invention addresses the problem of providing a monoclonal antibody or fragment thereof that binds to human CC chemokine receptor 1 (CCR1) and blocks the activation of human CCR1. The present invention relates to: a monoclonal antibody or a fragment thereof that binds to the extracellular region of human CCR1 and blocks the activation of human CCR1 by human CC chemokine ligand 15 (CCL15); a hybridoma that produces said antibody; a nucleic acid having a base sequence encoding said antibody or fragment thereof; a transformed cell including a vector that includes said nucleic acid; a method for producing said antibody or fragment thereof using said hybridoma or said transformed cell; a therapeutic agent and a diagnostic agent including said antibody or fragment thereof; and a treatment method and a diagnosis method for CCR1-related diseases using said antibody or fragment thereof.

Description

technical field [0001] The present invention relates to inhibition of activation of human CCR1 induced by human CC chemokine ligand (hereinafter referred to as human CCL)15 by binding to the extracellular region of human CC chemokine receptor 1 (hereinafter referred to as human CCR1). Activated monoclonal antibody or the antibody fragment, hybridoma producing the antibody, nucleic acid having a base sequence encoding the antibody or the antibody fragment, a transformed cell containing a vector containing the nucleic acid, using the hybridoma or the transformed cell A method for producing the antibody or the antibody fragment, a therapeutic agent and a diagnostic agent comprising the antibody or the antibody fragment, and a method for treating and diagnosing CCR1-related diseases using the antibody or the antibody fragment. Background technique [0002] CCR1 has aliases such as cluster of differentiation (CD) 191, CKR-1, HM145, macrophage inflammatory protein 1α receptor (Mac...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/28A61K39/395A61P29/00A61P35/00A61P37/02C12N1/15C12N1/19C12N1/21C12N5/10C12N5/12C12N15/13C12P21/08G01N33/53
CPCA61P29/00A61P35/00A61P37/02A61P43/00C07K16/2869C07K2317/70C07K2317/33C07K2317/30C07K2317/24C07K2317/92C07K2317/56C07K2317/565G01N2333/70596G01N33/6872C07K16/2866C12N5/10C12N5/12A61K39/395G01N33/53G01N33/577
Inventor 甲斐正之小川进也武藤诚河田健二平位秀世坂井义治前川平
Owner KYOWA HAKKO KIRIN CO LTD
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