JP2007254411A - Aggrecanase production inhibitor - Google Patents

Aggrecanase production inhibitor Download PDF

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JP2007254411A
JP2007254411A JP2006082606A JP2006082606A JP2007254411A JP 2007254411 A JP2007254411 A JP 2007254411A JP 2006082606 A JP2006082606 A JP 2006082606A JP 2006082606 A JP2006082606 A JP 2006082606A JP 2007254411 A JP2007254411 A JP 2007254411A
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aggrecanase
cryptoxanthin
arthritis
production inhibitor
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Akira Ito
晃 伊東
Keisuke Imada
啓介 今田
Takashi Sato
隆 佐藤
Ayana Tsuchida
綾菜 土田
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Tokyo University of Pharmacy and Life Sciences
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/01—Hydrocarbons
    • A61K31/015—Hydrocarbons carbocyclic
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/045—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/045—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates
    • A61K31/047—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates having two or more hydroxy groups, e.g. sorbitol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/045—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates
    • A61K31/07—Retinol compounds, e.g. vitamin A
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/12—Ketones
    • A61K31/122—Ketones having the oxygen directly attached to a ring, e.g. quinones, vitamin K1, anthralin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/02—Stomatological preparations, e.g. drugs for caries, aphtae, periodontitis
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
    • A61P19/10—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease for osteoporosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/02—Nutrients, e.g. vitamins, minerals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00

Abstract

<P>PROBLEM TO BE SOLVED: To provide a medicine for inhibiting production of aggrecanase. <P>SOLUTION: The aggrecanase production inhibitor contains at least one kind of carotenoid selected from the group consisting of compounds expressed by general formulae (1), (2) and (3), therapeutically permissible salts, ethers, esters and isomers thereof, wherein R<SP>1</SP>shows a hydrogen, an alkyl group, a cycloalkyl group, an aryl group, a heteroaryl group, an aralkyl group, a halogen, a hydroxy group or an alkoxy group, for which a heteroatom may be substituted; R<SP>2</SP>and R<SP>2</SP>' show each a hydrogen atom, a hydroxy group or an alkoxy group. A halogen atom may be substituted for an alkene hydrogen in the structural formulae. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

本発明はアグリカナーゼ阻害剤に関する。更に詳細には、本発明は天然物由来カロテノイド色素を含有するアグリカナーゼ産生阻害剤に関する。   The present invention relates to an aggrecanase inhibitor. More specifically, the present invention relates to an aggrecanase production inhibitor containing a natural product-derived carotenoid pigment.

平成１６年国民生活基礎調査の結果では、特に女性の有訴者の症状として「肩こり」、「腰痛」に続いて「手足の関節が痛む」が３位であり（人口千人あたり７２．７人）、極めて多くの人が関節痛を訴えていることが分かる。   According to the results of the 2004 National Life Basic Survey, “stress in shoulders” and “back pain” followed by “pain in limb joints” was the third most common symptom of female complainants (72.7 per 1,000 population). People), you can see that so many people complain of joint pain.

関節疾患としては、主に関節リウマチや変形性関節症などが挙げられるが、特に変形性関節症は中年期以降にもっとも多く見られる関節疾患である。これらの疾患においては炎症を抑えるために非ステロイド性抗炎症剤やステロイド製剤が一般的に使用され、また関節痛を抑える目的でヒアルロン酸製剤の関節内注射が用いられるが、その薬物療法は現在でも十分満足できるものとは言い難い。   Examples of joint diseases include rheumatoid arthritis and osteoarthritis, but osteoarthritis is the most common joint disease since middle age. In these diseases, non-steroidal anti-inflammatory drugs and steroid preparations are generally used to suppress inflammation, and hyaluronic acid preparations are used for intraarticular injection for the purpose of suppressing joint pain. But it's hard to say that it's satisfactory enough.

関節表面を覆う軟骨はII型コラーゲンならびに主にヒアルロン酸とアグリカンにより形成されるプロテオグリカン会合体により構成され、この構造が関節の滑らかな可動に寄与していると考えられている。関節リウマチや変形性関節症においては、この関節軟骨の分解が亢進し、激しい痛みとともに関節可動が制限される。関節軟骨の分解に深く関わる酵素としてコラゲナーゼに代表されるマトリックスメタロプロテアーゼが挙げられるが、関節リウマチや変形性関節症の関節液中にはむしろマトリックスメタロプロテアーゼ以外のタンパク質分解酵素によるアグリカン分解産物が多く検出される（例えば、非特許文献１参照）ことから、アグリカンを分解する新たな酵素がこれら疾患に深く関わっているものと考えられていた。   The cartilage covering the joint surface is composed of type II collagen and proteoglycan aggregates formed mainly by hyaluronic acid and aggrecan, and this structure is thought to contribute to the smooth movement of the joint. In rheumatoid arthritis and osteoarthritis, the degradation of the articular cartilage is enhanced, and joint movement is restricted with severe pain. Matrix metalloproteases typified by collagenase are examples of enzymes that are deeply involved in the degradation of articular cartilage, but there are many aggrecan degradation products by proteolytic enzymes other than matrix metalloproteases in rheumatoid arthritis and osteoarthritic joint fluid. Since it is detected (see, for example, Non-Patent Document 1), it has been considered that a new enzyme that degrades aggrecan is deeply involved in these diseases.

１９９９年にアグリカンを特異的に分解する２種類のアグリカナーゼが発見された（例えば、非特許文献１および非特許文献２参照）。２種類のアグリカナーゼ（アグリカナーゼ−１およびアグリカナーゼ−２）は、いずれもA disintegrin and metalloproteinase with thrombospondin motifs（ADAMTS）と呼ばれる酵素群に属し、一般的にアグリカナーゼ−１はADAMTS-4、アグリカナーゼ−２はADAMTS-5と呼ばれる。また両アグリカナーゼは同時に産生されるTissue inhibitor of metalloproteinases (TIMP)-3により活性が阻害される（例えば、非特許文献３参照）。ADAMTS-5については最近変形性関節症や関節炎における関節破壊に中心的な役割を担っていることがADAMTS-5ノックアウトマウスを用いた実験で証明されている（例えば、非特許文献４および非特許文献５参照）。   Two kinds of aggrecanases that specifically degrade aggrecan were discovered in 1999 (see, for example, Non-Patent Document 1 and Non-Patent Document 2). Two types of aggrecanases (aggrecanase-1 and aggrecanase-2) all belong to an enzyme group called A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS). Generally, aggrecanase-1 is ADAMTS-4 and aggrecanase-2 is ADAMTS. Called -5. The activity of both aggrecanases is inhibited by Tissue inhibitor of metalloproteinases (TIMP) -3 produced simultaneously (see, for example, Non-Patent Document 3). ADAMTS-5 has recently been proved to play a central role in joint destruction in osteoarthritis and arthritis in experiments using ADAMTS-5 knockout mice (for example, Non-Patent Document 4 and Non-Patent Documents). Reference 5).

アグリカナーゼは、アグリカンのみならず脳の構成プロテオグリカンであるブレビカンをも分解し、脳神経系のがん細胞である神経膠芽腫細胞の浸潤・転移にも深く関わっていることが報告されている（例えば、非特許文献６参照）。   It has been reported that aggrecanase degrades not only aggrecan but also brebican, a constituent proteoglycan of the brain, and is also deeply involved in invasion and metastasis of glioblastoma cells, which are cancer cells of the cranial nervous system (for example, Non-patent document 6).

一方、これらアグリカナーゼの活性を抑制する薬剤については既に幾つか報告されている。例えば、特許文献１には、（2R, 3R）1-[4-(2, 4-ジクロロ-ベンジルオキシ)-ベンゼンスルホニル]-3-ヒドロキシ-3-メチル-ピペリジン-2-カルボン酸ヒドロキシアミドなどのカルボン酸ヒドロキシアミド誘導体からなるアグリカナーゼ阻害剤が、また、特許文献２にはフィブロネクチン，またはフィブロネクチンのCOOH末端に位置する約40キロダルトンの断片からなるアグリカナーゼ阻害剤が、更に、特許文献３には、N-ヒドロキシ-Nα-メチル-Nα-（4-フェノキシベンゼンスルフォニル）-2-[2-(ピリミジン-2，4-ジオン-1イル)エチル]グリシンアミドなどのスルホンアミド誘導体からなるアグリカナーゼ阻害剤が記載されている。しかし、これらのアグリカナーゼ阻害剤は何れも薬物治療あるいは予防に応用されるまでには至っていない。また、これら疾患は長年にわたって徐々に進行して行くことからも、予防を含めて長期的に治療を続ける必要があり、高い安全性が要求される。   On the other hand, some drugs that suppress the activity of these aggrecanases have already been reported. For example, Patent Document 1 discloses (2R, 3R) 1- [4- (2,4-dichloro-benzyloxy) -benzenesulfonyl] -3-hydroxy-3-methyl-piperidine-2-carboxylic acid hydroxyamide, etc. An aggrecanase inhibitor comprising a carboxylic acid hydroxyamide derivative of the above, and Patent Document 2 discloses fibronectin or an aggrecanase inhibitor comprising a fragment of about 40 kilodalton located at the COOH terminus of fibronectin. , An aggrecanase inhibitor comprising a sulfonamide derivative such as N-hydroxy-Nα-methyl-Nα- (4-phenoxybenzenesulfonyl) -2- [2- (pyrimidin-2,4-dione-1yl) ethyl] glycinamide Is described. However, none of these aggrecanase inhibitors have been applied to drug treatment or prevention. Moreover, since these diseases progress gradually over many years, it is necessary to continue treatment for a long term including prevention, and high safety is required.

特開２００１−１１４７６５号公報JP 2001-114765 A 特開２００４−２５６４３６号公報JP 2004-256436 A 特開２００１−１６３８８５号公報Japanese Patent Laid-Open No. 2001-163885 Tottorella MD, Burn TC, Pratta MA et al.; Science; vol. 284:1664-1666; 1999Tottorella MD, Burn TC, Pratta MA et al .; Science; vol. 284: 1664-1666; 1999 Abbaszade I, Liu R-Q, Yang F et al.; J Biol Chem; vol. 274:23443-23450; 1999Abbaszade I, Liu R-Q, Yang F et al .; J Biol Chem; vol. 274: 23443-23450; 1999 Kashiwagi M, Tortorella M, Nagase H et al.; J Biol Chem; vol. 276:12501-12504; 2001Kashiwagi M, Tortorella M, Nagase H et al .; J Biol Chem; vol. 276: 12501-12504; 2001 Glasson SS, Askew R, Sheppard B et al.; Nature; vol.434:644-648; 2005Glasson SS, Askew R, Sheppard B et al .; Nature; vol.434: 644-648; 2005 Stanton H, Rogerson FM, East CJ et al.; Nature; vol.434:648-652; 2005Stanton H, Rogerson FM, East CJ et al .; Nature; vol. 434: 648-652; 2005 Nakada M, Miyamori H, Kita D et al.; Acta Neuropathol (Berl); vol. 110:239-246; 2005Nakada M, Miyamori H, Kita D et al .; Acta Neuropathol (Berl); vol. 110: 239-246; 2005

従って、本発明の目的はアグリカナーゼの産生を阻害する薬剤を提供することである。   Accordingly, an object of the present invention is to provide a drug that inhibits the production of aggrecanase.

本発明者らは、天然物由来カロテノイド色素が、ヒト滑膜細胞および軟骨細胞において、軟骨基質アグリカンを特異的に分解するアグリカナーゼ-1（ADAMTS-4）および-2(ADAMTS-5)の発現を極めて効果的に抑制することを発見し、本発明を完成させるに至った。すなわち、前記課題を解決するための手段として本発明は、天然物由来カロテノイド色素を含有するアグリカナーゼ産生阻害剤を提供する。   The present inventors show that natural product-derived carotenoid pigments express aggrecanase-1 (ADAMTS-4) and -2 (ADAMTS-5) that specifically degrade cartilage matrix aggrecan in human synovial cells and chondrocytes. It has been found that it is extremely effectively suppressed, and the present invention has been completed. That is, as a means for solving the above problems, the present invention provides an aggrecanase production inhibitor containing a natural product-derived carotenoid pigment.

本発明のアグリカナーゼ産生阻害剤は、アグリカナーゼによるアグリカンの分解に起因すると思われる関節リウマチや変形性関節症などの疾患の予防及び／又は治療用薬剤として利用できるばかりか、がん細胞の浸潤・転移の抑制及び肺癌、子宮頸部癌、食道癌、膀胱癌、大腸癌、皮膚癌などの発癌リスクの低減にも有用である。更に、本発明のアグリカナーゼ産生阻害剤を含有する機能性食品はこれら疾患の予防にも有用である。   The aggrecanase production inhibitor of the present invention can be used not only as a preventive and / or therapeutic agent for diseases such as rheumatoid arthritis and osteoarthritis, which may be caused by the degradation of aggrecan by aggrecanase, but also infiltration / metastasis of cancer cells It is also useful for the suppression of cancer and the risk of developing cancer such as lung cancer, cervical cancer, esophageal cancer, bladder cancer, colon cancer, skin cancer and the like. Furthermore, the functional food containing the aggrecanase production inhibitor of the present invention is also useful for preventing these diseases.

本発明のアグリカナーゼ産生阻害剤は、天然物由来カロテノイド色素を含有する。カロテノイド色素は様々な野菜類及び果物類に含有されるが、特に温州ミカン、パパイヤ、柿、桃、枇杷などの果物類及びピーマン（特に、赤ピーマン）、トマト、人参、カボチャなどの緑黄色野菜類に豊富に含まれる。   The aggrecanase production inhibitor of the present invention contains a natural product-derived carotenoid pigment. Carotenoid pigments are contained in various vegetables and fruits, especially fruits such as Wenzhou mandarin oranges, papayas, persimmons, peaches, persimmons and green and yellow vegetables such as peppers (especially red peppers), tomatoes, carrots, and pumpkins. Abundantly included.

本発明のアグリカナーゼ産生阻害剤は、下記の一般式（１）、（２）及び（３）、



（前記各式中、Ｒ１は、水素、アルキル基、シクロアルキル基、アリール基、ヘテロアリール基、アラルキル基、ハロゲン、ヒドロキシ基又はアルコキシ基を示し、それらはヘテロ原子で置換されていてもよい；Ｒ２およびＲ２’は、水素、ヒドロキシ基又はアルコキシ基を示す。構造式中のアルケン水素はハロゲン原子で置換されていてもよい。）で示される化合物、その治療上許容し得る塩、エーテル、エステル及び異性体からなる群から選択される少なくとも一種類のカロテノイドを含有する。 The aggrecanase production inhibitor of the present invention has the following general formulas (1), (2) and (3),



(In the above formulas, R 1 represents hydrogen, an alkyl group, a cycloalkyl group, an aryl group, a heteroaryl group, an aralkyl group, a halogen, a hydroxy group, or an alkoxy group, which may be substituted with a heteroatom. R 2 and R 2 ′ represent hydrogen, a hydroxy group or an alkoxy group. The alkene hydrogen in the structural formula may be substituted with a halogen atom.), A therapeutically acceptable salt thereof, It contains at least one carotenoid selected from the group consisting of ethers, esters and isomers.

前記一般式（１）、（２）及び（３）の定義における「異性体」という用語は、立体異性、幾何異性、光学異性、平面異性、配位異性、配位位置異性などの全ての異性体を含む意味で使用されている。   The term “isomer” in the definitions of the general formulas (1), (2) and (3) means all isomerism such as stereoisomerism, geometric isomerism, optical isomerism, plane isomerism, coordination isomerism, coordination position isomerism, etc. Used to include the body.

前記一般式（１）で示される化合物の代表例を挙げる。
Ｒ１＝ＣＨ３，Ｒ２＝Ｈ，Ｒ２’＝Ｈ：α−カロテン；
Ｒ１＝ＣＨ３，Ｒ２＝ＯＨ，Ｒ２’＝Ｈ：α−クリプトキサンチン（α−カロテン−３−オール）；
Ｒ１＝ＣＨ３，Ｒ２＝ＯＨ，Ｒ２’＝ＯＨ：ルテイン（α−カロテン−３，３’−ジオール） The typical example of a compound shown by the said General formula (1) is given.
R 1 = CH 3 , R 2 = H, R 2 ′ = H: α-carotene;
R 1 = CH 3 , R 2 = OH, R 2 ′ = H: α-cryptoxanthin (α-carotene-3-ol);
R 1 = CH 3 , R 2 = OH, R 2 ′ = OH: Lutein (α-carotene-3,3′-diol)

前記一般式（２）で示される化合物の代表例を挙げる。
Ｒ１＝ＣＨ３，Ｒ２＝Ｈ，Ｒ２’＝Ｈ：β−カロテン；
Ｒ１＝ＣＨ３，Ｒ２＝ＯＨ，Ｒ２’＝Ｈ：β−クリプトキサンチン（β−カロテン−３−オール）；
Ｒ１＝ＣＨ３，Ｒ２＝ＯＨ，Ｒ２’＝ＯＨ：ゼアキサンチン（β，β−カロテン−３，３’−ジオール） The typical example of a compound shown by the said General formula (2) is given.
R 1 = CH 3 , R 2 = H, R 2 ′ = H: β-carotene;
R 1 = CH 3 , R 2 = OH, R 2 ′ = H: β-cryptoxanthin (β-carotene-3-ol);
R 1 = CH 3 , R 2 = OH, R 2 ′ = OH: Zeaxanthin (β, β-carotene-3,3′-diol)

前記一般式（３）で示される化合物の代表例を挙げる。
Ｒ１＝ＣＨ３，Ｒ２＝Ｈ，Ｒ２’＝Ｈ：カンタキサンチン（β，β−カロテン−４，４’−ジオン）；
Ｒ１＝ＣＨ３，Ｒ２＝ＯＨ，Ｒ２’＝ＯＨ：アスタキサンチン（３，３’−ジヒドロキシ−β，β−カロテン−４，４’−ジオン） The typical example of a compound shown by the said General formula (3) is given.
R 1 = CH 3 , R 2 = H, R 2 ′ = H: canthaxanthin (β, β-carotene-4,4′-dione);
R 1 = CH 3 , R 2 = OH, R 2 ′ = OH: Astaxanthin (3,3′-dihydroxy-β, β-carotene-4,4′-dione)

前記一般式（１）〜（３）で示されるカロテノイド色素化合物は全て天然物に由来するものであり、細胞毒性が極めて低いという利点を有し、当業者に公知、慣用の方法により野菜類、果物類などの植物群から容易に抽出、分離することができる。   The carotenoid pigment compounds represented by the general formulas (1) to (3) are all derived from natural products and have the advantage of extremely low cytotoxicity. Vegetables, It can be easily extracted and separated from a group of plants such as fruits.

従来の薬剤が単にアグリカナーゼの酵素活性を抑制する薬理効果しか有しなかったのに対して、前記一般式（１）〜（３）で示されるカロテノイド色素化合物からなる本発明の薬剤は、ヒト滑膜細胞および軟骨細胞において、軟骨基質アグリカンを特異的に分解するアグリカナーゼ-1（ADAMTS-4）および-2(ADAMTS-5)の遺伝子の発現を抑制する薬理効果を有する点で画期的である。すなわち、従来の薬剤が、既に発生してしまったアグリカナーゼの酵素活性を抑制する対処療法的薬剤であるのに対して、本発明の薬剤はアグリカナーゼ自体の発現を阻止する根治的薬剤である点で明確に相違する。   Whereas conventional drugs merely have a pharmacological effect of suppressing the enzyme activity of aggrecanase, the drugs of the present invention comprising the carotenoid pigment compounds represented by the above general formulas (1) to (3) It is revolutionary in that it has a pharmacological effect to suppress the expression of aggrecanase-1 (ADAMTS-4) and -2 (ADAMTS-5) genes that specifically degrade cartilage matrix aggrecan in membrane cells and chondrocytes . That is, the conventional drug is a coping therapy drug that suppresses the enzymatic activity of aggrecanase that has already occurred, whereas the drug of the present invention is a radical drug that blocks the expression of aggrecanase itself. Clearly different.

前記一般式（１）〜（３）で示されるカロテノイド色素化合物からなる本発明の薬剤は、関節リウマチや変形性関節症などの疾患の他、癌細胞の浸潤・転移抑制にも有用である。特に、肺癌、子宮頸部癌、食道癌、膀胱癌、大腸癌、皮膚癌に対して有意な発癌抑制効果を発揮する。更に、骨関節症、関節損傷、若年性関節リウマチ、反応性関節炎、乾癬性関節炎、神経病性関節症（シャルコー関節）、血友病性関節症、感染性関節炎、強直性脊椎炎、ライター症候群、痛風、急性ピロリン酸関節炎（偽性通風）、炎症性腸疾患、クローン病、歯周疾患、骨粗鬆症及び人工関節インプラントの弛みなどの疾患に対しても有用である。機能性食品とした場合、これら疾患の予防効果が期待できる。   The drug of the present invention comprising the carotenoid pigment compounds represented by the general formulas (1) to (3) is useful for suppressing invasion / metastasis of cancer cells in addition to diseases such as rheumatoid arthritis and osteoarthritis. In particular, it exerts a significant carcinogenic inhibitory effect on lung cancer, cervical cancer, esophageal cancer, bladder cancer, colon cancer, and skin cancer. Furthermore, osteoarthritis, joint damage, juvenile rheumatoid arthritis, reactive arthritis, psoriatic arthritis, neuropathic arthropathy (Charcot joint), hemophilic arthropathy, infectious arthritis, ankylosing spondylitis, Reiter syndrome It is also useful for diseases such as gout, acute pyrophosphate arthritis (pseudo ventilation), inflammatory bowel disease, Crohn's disease, periodontal disease, osteoporosis and loosening of artificial joint implants. When it is a functional food, it can be expected to prevent these diseases.

本発明のアグリカナーゼ産生阻害剤を医薬品として使用する場合、疾患の治療及び／又は予防の目的のために、薬理学的に有効量の前記一般式（１）〜（３）で示されるカロテノイド色素化合物を、製剤学的に許容され得る他の成分と共に製剤化することができる。製剤学的に許容され得る他の成分は、例えば、担体、希釈剤、賦形剤、滑沢剤、結合剤、溶解補助剤、等張化剤、酸化防止剤、防腐剤、界面活性剤、乳化剤、着色剤、香料、甘味料などである。   When the aggrecanase production inhibitor of the present invention is used as a pharmaceutical, a pharmacologically effective amount of the carotenoid pigment compound represented by the general formulas (1) to (3) for the purpose of treating and / or preventing a disease Can be formulated with other pharmaceutically acceptable ingredients. Other pharmaceutically acceptable ingredients include, for example, carriers, diluents, excipients, lubricants, binders, solubilizers, tonicity agents, antioxidants, preservatives, surfactants, Emulsifiers, colorants, flavors, sweeteners and the like.

本発明のアグリカナーゼ産生阻害剤を医薬品として使用する場合、有効成分である前記一般式（１）〜（３）で示されるカロテノイド色素化合物の配合量は、製剤学的に許容され得る他の成分を含めた総重量を基準にして、一般的に、０．０１重量％〜１５重量％の範囲内であり、０．１重量％〜５重量％であることが好ましい。   When the aggrecanase production inhibitor of the present invention is used as a pharmaceutical, the compounding amount of the carotenoid pigment compound represented by the general formulas (1) to (3), which is an active ingredient, is different from that of other pharmaceutically acceptable ingredients. In general, it is in the range of 0.01% to 15% by weight, preferably 0.1% to 5% by weight, based on the total weight included.

本発明のアグリカナーゼ産生阻害剤を医薬品として使用する場合、任意の剤形で使用することができる。例えば、顆粒剤、細粒剤、錠剤、カプセル剤、丸剤、軟膏、ゲル、ペースト、クリーム、噴霧剤、溶液剤、懸濁液剤などの剤形である。投与経路としては、経口、経腸経鼻、及び経皮投与の他、静注、皮下注、筋肉注、関節膣注などの注射投与を含む全ての投与経路が利用できる。   When the aggrecanase production inhibitor of the present invention is used as a pharmaceutical product, it can be used in any dosage form. Examples thereof include dosage forms such as granules, fine granules, tablets, capsules, pills, ointments, gels, pastes, creams, sprays, solutions, suspensions and the like. As the administration route, all administration routes including injection administration such as intravenous injection, subcutaneous injection, intramuscular injection, articular vaginal injection and the like can be used in addition to oral, enteral, nasal and transdermal administration.

本発明のアグリカナーゼ産生阻害剤を医薬品として使用する場合、有効成分である前記一般式（１）〜（３）で示されるカロテノイド色素化合物の量に換算して、通常、０．０５ｍｇ〜１０００ｍｇの投与量で、一日に１〜数回程度投与することができる。例えば、β−カロテンの場合、３．１１ｍｇ／日の投与量で、２１．０μｇ／ｄｌの血清濃度が、また、β−クリプトキサンチンの場合、０．０７ｍｇ／日の投与量で、１２．３〜１３．３μｇ／ｄｌの血清濃度が得られる。特に、β−クリプトキサンチンは吸収されやすく、低投与量でも高い血清濃度が得られるので好ましい。この投与量は、患者の年齢、体重、性別に応じて適宜変化させることができ、また、投与経路及び患者の病状の程度に応じて適宜変更することができる。   When the aggrecanase production inhibitor of the present invention is used as a pharmaceutical product, it is usually administered in an amount of 0.05 mg to 1000 mg in terms of the amount of the carotenoid pigment compound represented by the general formulas (1) to (3) as active ingredients. The dose can be administered 1 to several times a day. For example, in the case of β-carotene, a serum concentration of 21.0 μg / dl at a dose of 3.11 mg / day, and in the case of β-cryptoxanthin at a dose of 0.07 mg / day, 12.3 A serum concentration of ˜13.3 μg / dl is obtained. In particular, β-cryptoxanthin is preferable because it is easily absorbed and a high serum concentration can be obtained even at a low dose. This dose can be appropriately changed according to the age, weight and sex of the patient, and can be appropriately changed according to the administration route and the degree of the patient's medical condition.

本発明のアグリカナーゼ産生阻害剤を機能性食品として使用する場合、その食品の形状としては、当業者に周知の全ての固体状、半固体状、液状、懸濁液状、ゲル状、糊状、粉末状、顆粒状食品を含むことができる。   When the aggrecanase production inhibitor of the present invention is used as a functional food, the shape of the food is all solid, semi-solid, liquid, suspension, gel, paste, and powder known to those skilled in the art. And granular foods.

温州ミカンの全形をミキサーにかけて粉砕し、これをアセトンと混合し、上層を粗カロテノイド色素化合物抽出液として得た。これをロータリーエバポレータで濃縮、乾固した後、５０％エタノールに再溶解し、オクタデシルシリカゲルを担体とする逆相系カラム、溶離液としてメタノール／１０ｍＭ燐酸（４：６→６：４）を用い、紫外線吸収検出器（波長：３４０ｎｍ）でモニターしながら分取を行った。得られた分画を濃縮、乾固することにより目的のカロテノイド色素化合物を得た。   The whole form of Satsuma mandarin was pulverized with a mixer and mixed with acetone, and the upper layer was obtained as a crude carotenoid pigment compound extract. This was concentrated and dried on a rotary evaporator, then redissolved in 50% ethanol, using a reverse phase column with octadecyl silica gel as a carrier, methanol / 10 mM phosphoric acid (4: 6 → 6: 4) as the eluent, Sorting was performed while monitoring with an ultraviolet absorption detector (wavelength: 340 nm). The obtained fraction was concentrated and dried to obtain the desired carotenoid pigment compound.

α−カロテン：
融点：１８７．５℃ α-Carotene:
Melting point: 187.5 ° C

β−クリプトキサンチン（β−カロテン−３−オール）：
融点：１５８℃〜１５９℃ β-cryptoxanthin (β-carotene-3-ol):
Melting point: 158 ° C-159 ° C

ゼアキサンチン（β，β−カロテン−３，３’−ジオール）：
融点：２０７℃ Zeaxanthin (β, β-carotene-3,3′-diol):
Melting point: 207 ° C

カンタキサンチン（β，β−カロテン−４，４’−ジオン）：
融点：２１７℃ Canthaxanthin (β, β-carotene-4,4′-dione):
Melting point: 217 ° C

アスタキサンチン（３，３’−ジヒドロキシ−β，β−カロテン−４，４’−ジオン）：
融点：１８２℃〜１８３℃ Astaxanthin (3,3′-dihydroxy-β, β-carotene-4,4′-dione):
Melting point: 182 ° C-183 ° C

ヒト滑膜細胞およびヒト関節軟骨細胞のアグリカナーゼ−１（ＡＤＡＭＴＳ−４）およびアグリカナーゼ−２（ＡＤＡＭＴＳ−５）発現に対するβ−クリプトキサンチンの作用を検証した。   The effect of β-cryptoxanthin on aggrecanase-1 (ADAMTS-4) and aggrecanase-2 (ADAMTS-5) expression in human synovial cells and human articular chondrocytes was examined.

［実験材料及び実験方法］
1． 培養器具および試薬
細胞培養用プラスチック器具は旭テクノグラス社製; Dulbeco's modified Eagle's medium (DMEM) はInvirtogen社製; Ca2+ and Mg2+-free phosphate buffered saline (PBS (-)) は日水製薬社製; fetal bovine serum (FBS) はBioWhittaker社製; penicillin Gは万有製薬社製; streptomycin sulfate は明治製菓社製; trypsin は DIFCO Laboratories Co.製; lactalbumin hydrolysate (LAH), bovine serum albumin (BSA), peroxydase-conjugated goat anti-(sheep IgG) IgG, dimethyl sulphoxide (DMSO) および diethyl pyrocarbonate (DEPC)はSigma社製; Isogen はニッポンジーン社製; mouse anti-[human cyclooxygenase (COX-1)] monoclonal antibody, mouse anti-(human COX-2) monoclonal antibodyはCayman社製を各々購入し使用した。
Recombinant human interleukin-1β (rhIL-1β, 2 x 107 units/mg) は大塚製薬より提供を受けた。
その他の試薬は全て特級試薬を使用した。
2．ヒト関節滑膜細胞および軟骨細胞の培養および薬物処理
ヒト滑膜細胞(Applied Cell Biology Research Instituteより購入)および軟骨細胞(Cambrex Bio Science Walkersville社より購入)をそれぞれ 6-multiwell plate に播種し、DMEM/10% (v/v) FBS中でコンフルエントまで培養した。この細胞をDMEM/0.2% (w/v) LAH で洗浄後、DMEM/0.2% (w/v) LAH中に薬物を添加し、 細胞培養系に添加した。なお、β-cryptoxanthinは、すべて DMSO溶液として DMSOの終濃度が 0.1% (v/v)になるように添加した。また、β-cryptoxanthin無添加群にも同濃度のDMSOを添加した。5% CO2 -95% air 気相下、37℃で24時間インキュベーション後、培養液ならびに細胞画分を回収し実験に供するまで -20℃で保存した。
3．リアルタイムRT-PCR法
1) 総RNAの抽出
総RNAの抽出はIsogenを用いて添付文書に従い実施した。Ribonucleaseの混入を防ぐため操作は清潔なディスポーザブルのグローブを着用して行った。乾熱滅菌可能な器具については 180℃ で9時間以上乾熱滅菌し、不可能な器具については未使用のものをオートクレーブで3回処理した。使用した水はオートクレーブで3回処理した0.2% DEPC溶液を用いた。
2) 逆転写反応
総RNA (1 μg)からcDNAを合成するためQuantitect Reverse Transcription kit (Qiagen社製)を用いて添付文書に従い逆転写反応を実施した。
3) リアルタイムPCR法
逆転写反応により得られたcDNA（総RNA量として25 ngに相当する量）を用いて ADAMTS-4 mRNA、ADAMTS-5 mRNAならびにglyceraldehyde-3-phospate dehydrogenase (GAPDH) mRNAに特異的なpolymerase chain reaction (PCR)用Primer (Quantitect Primer Assay, Qiagen社製)を用いて, ABI Prism sequence detection 7000 (Applied Biosystems社製)によりPCR反応ならびにデータ解析を行った。データはGAPDH mRNA発現量により補正した後、無処理群の遺伝子発現量を1とした相対値として表した。
4．プロスタグランジンE2量の測定
培養液中プロスタグランジンE2量の定量は、Prostaglandin E2 EIA kit-Monoclonal (Cayman社製)を用いて添付文書に従って実施した。
5．ウエスタンブロット法
細胞質画分（タンパク質量として10μg）に20% (w/v) trichloroacetic acid (TCA) 溶液を終濃度 3.3% (w/v)となるよう加え、4℃で12 時間以上放置した後、8,000 x gで 5 分間遠心分離した。得られた沈殿を diethyletherで1回洗浄し、風乾後sample 溶液（63 mM Tris-HCl (pH6.8)/2% sodium dodecylsulfate (SDS)/10% glycerol/1% 2-mercaptoethanol/0.001% bromophenol blue）に溶解した。SDS-polyacrylamide gel electrophoresis (PAGE)にて試料中タンパク質を分離後、ゲルを転写用緩衝液 (20 mM Tris /150 mM glycine/20% (w/v) methanol/0.1% (w/v) SDS (pH 9.2))に浸したニトロセルロース膜に密着させ、セミドライ型転写装置を用い2 mA/cm2で1 時間タンパク質の転写を行った。転写後、ニトロセルロース膜をブロッキング溶液[10% (w/v) fatty-free dry milk/10 mM Tris-HCl/0.9% NaCl/0.02% NaN3 (pH 7.5)]に浸し、10 分間ブロッキングを行った。続いてニトロセルロース膜をイオン交換水およびPBS-T 緩衝液 [0.1% Tween 20/PBS(-)]にて数回洗浄し、1% (w/v) BSA/PBS (-) で希釈した1次抗体[mouse anti-(human COX-1)モノクローナル抗体またはmouse anti-(human COX-2)モノクローナル抗体]溶液に4℃で12時間以上浸した。1次抗体結合後、ブロッキングおよび洗浄を同様に行い、peroxydase-conjugated goat anti -(mouse IgG)IgGに室温で1 時間浸した。2次抗体結合後、PBS-T 緩衝液にて数回洗浄し、ECL-Western blot detection reagents (アマシャム社製)に浸し、正確に1分間反応させた。反応終了後、直ちにImage Analyzer LAS-1000 plus (富士写真フィルム社製)を用いて検出した。
6．生細胞数の測定
細胞増殖能はCell counting kit-8 (Dojindo社製) を用いて、添付の操作方法に従って測定した。すなわち、ヒト滑膜細胞を96-well plate に細胞数 1 x 104 cells/100μL/well で播種し、DMEM/10% (v/v) FBS中で5% CO2 - 95% air 気相下、37℃で24 時間培養後、βクリプトキサンチンを含むDMEM/10% (v/v) FBSで24時間処理を行った。次にCell couting kit-8を10μL/well添加し、5% CO2 - 95% air 気相下、37℃で2時間インキュベーションし 430 nm の吸光度を測定した。 [Experimental materials and experimental methods]
1． Culture equipment and reagents Plastic equipment for cell culture is manufactured by Asahi Techno Glass; Dulbeco's modified Eagle's medium (DMEM) is manufactured by Invirtogen; Ca 2+ and Mg 2+ -free phosphate buffered saline (PBS (-)) is Nissui Pharmaceutical Fetal bovine serum (FBS) manufactured by BioWhittaker; penicillin G manufactured by Banyu Pharmaceutical; streptomycin sulfate manufactured by Meiji Seika; trypsin manufactured by DIFCO Laboratories Co .; lactalbumin hydrolysate (LAH), bovine serum albumin (BSA ), peroxydase-conjugated goat anti- (sheep IgG) IgG, dimethyl sulphoxide (DMSO) and diethyl pyrocarbonate (DEPC) are from Sigma; Isogen is from Nippon Gene; mouse anti- [human cyclooxygenase (COX-1)] monoclonal antibody Mouse anti- (human COX-2) monoclonal antibody was purchased from Cayman.
Recombinant human interleukin-1β (rhIL-1β, 2 x 10 7 units / mg) was provided by Otsuka Pharmaceutical.
All other reagents used special grade reagents.
2． Culture and drug treatment of human articular synovial cells and chondrocytes Human synovial cells (purchased from Applied Cell Biology Research Institute) and chondrocytes (purchased from Cambrex Bio Science Walkersville) were seeded on 6-multiwell plates, respectively, and DMEM / Cultured to confluence in 10% (v / v) FBS. After washing the cells with DMEM / 0.2% (w / v) LAH, a drug was added to DMEM / 0.2% (w / v) LAH and added to the cell culture system. Β-cryptoxanthin was added as a DMSO solution so that the final concentration of DMSO was 0.1% (v / v). The same concentration of DMSO was also added to the group without β-cryptoxanthin. After incubation for 24 hours at 37 ° C. in a gas phase of 5% CO 2 -95% air, the culture solution and cell fraction were collected and stored at −20 ° C. until use in experiments.
3． Real-time RT-PCR method
1) Extraction of total RNA Total RNA was extracted using Isogen according to the package insert. To prevent Ribonuclease contamination, the operation was performed wearing clean disposable gloves. For devices that can be sterilized by dry heat, they were sterilized by dry heat at 180 ° C for over 9 hours, and for those that were not possible, unused ones were treated three times in an autoclave. The water used was a 0.2% DEPC solution treated three times in an autoclave.
2) Reverse transcription reaction In order to synthesize cDNA from total RNA (1 μg), a reverse transcription reaction was performed using a Quantitect Reverse Transcription kit (Qiagen) according to the package insert.
3) Specific to ADAMTS-4 mRNA, ADAMTS-5 mRNA, and glyceraldehyde-3-phospate dehydrogenase (GAPDH) mRNA using cDNA obtained by real-time PCR reverse transcription (total RNA equivalent to 25 ng) PCR reaction and data analysis were performed by ABI Prism sequence detection 7000 (Applied Biosystems) using a standard primer chain for polymerase chain reaction (PCR) (Quantitect Primer Assay, Qiagen). Data was expressed as a relative value with the gene expression level of the untreated group taken as 1, after correction by the GAPDH mRNA expression level.
Four. Prostaglandin E 2 of measuring culture Prostaglandin E 2 of quantification was performed according to the attached document using Prostaglandin E 2 EIA kit-Monoclonal ( Cayman Inc.).
Five. Western blotting Add a 20% (w / v) trichloroacetic acid (TCA) solution to the cytoplasmic fraction (10 μg protein) to a final concentration of 3.3% (w / v) and leave it at 4 ° C for 12 hours or longer. And centrifuged at 8,000 xg for 5 minutes. The resulting precipitate was washed once with diethylether, air-dried and then sample solution (63 mM Tris-HCl (pH 6.8) / 2% sodium dodecylsulfate (SDS) / 10% glycerol / 1% 2-mercaptoethanol / 0.001% bromophenol blue ). After separating the protein in the sample by SDS-polyacrylamide gel electrophoresis (PAGE), the gel is transferred to a transfer buffer (20 mM Tris / 150 mM glycine / 20% (w / v) methanol / 0.1% (w / v) SDS ( The protein was adhered to a nitrocellulose membrane soaked in pH 9.2)), and protein transfer was performed at 2 mA / cm 2 for 1 hour using a semi-dry type transfer apparatus. After transfer, the nitrocellulose membrane is immersed in a blocking solution [10% (w / v) fatty-free dry milk / 10 mM Tris-HCl / 0.9% NaCl / 0.02% NaN 3 (pH 7.5)] and blocked for 10 minutes. It was. Subsequently, the nitrocellulose membrane was washed several times with ion-exchanged water and PBS-T buffer [0.1% Tween 20 / PBS (-)] and diluted with 1% (w / v) BSA / PBS (-) 1 It was immersed in a subsequent antibody [mouse anti- (human COX-1) monoclonal antibody or mouse anti- (human COX-2) monoclonal antibody] solution at 4 ° C. for 12 hours or more. After binding with the primary antibody, blocking and washing were performed in the same manner, and immersed in peroxydase-conjugated goat anti- (mouse IgG) IgG at room temperature for 1 hour. After binding the secondary antibody, it was washed several times with PBS-T buffer, immersed in ECL-Western blot detection reagents (Amersham), and reacted exactly for 1 minute. Immediately after the reaction was completed, detection was performed using Image Analyzer LAS-1000 plus (manufactured by Fuji Photo Film Co., Ltd.).
6． Viable cell count measurement
Cell proliferation ability was measured by using Cell counting kit-8 (manufactured by Dojindo) according to the attached operation method. That is, the human synovial cells were seeded at 96-well plate cell number 1 to x 10 4 cells / 100μL / well , DMEM / 10% (v / v) 5% in FBS CO 2 - 95% air gas phase under After culturing at 37 ° C. for 24 hours, the cells were treated with DMEM / 10% (v / v) FBS containing β cryptoxanthin for 24 hours. Next, Cell couting kit-8 was added at 10 μL / well, and the absorbance at 430 nm was measured by incubation at 37 ° C. for 2 hours in 5% CO 2 -95% air gas phase.

［結果］
ヒト滑膜細胞においてＡＤＡＭＴＳ−４ ｍＲＮＡ発現（図１参照）およびＡＤＡＭＴＳ−５ ｍＲＮＡ発現（図２参照）はＩＬ−１β（１０ｎｇ／ｍＬ）により促進した。これに対して、β−クリプトキサンチン（β−ＣＰＸ）は１μＭから１０μＭにかけて濃度依存的にＡＤＡＭＴＳ−４ ｍＲＮＡ発現およびＡＤＡＭＴＳ−５ ｍＲＮＡ発現を抑制した。軟骨細胞においてもＡＤＡＭＴＳ−４ ｍＲＮＡ発現はＩＬ−１βにより促進し、β−ＣＰＸは濃度依存的にＡＤＡＭＴＳ−４ ｍＲＮＡ発現を抑制した（図３参照）。ＡＤＡＭＴＳ−５ ｍＲＮＡ発現は滑膜細胞の場合と異なりＩＬ−１βによる影響を受けなかったが，β−ＣＰＸはＡＤＡＭＴＳ−５ ｍＲＮＡ発現を濃度依存的に抑制した（図４参照）。また、ヒト滑膜細胞におけるＰＧＥ２産生とＣＯＸ−１およびＣＯＸ−２の発現に対するβ−ＣＰＸの影響を検討した結果、ＩＬ−１βによるＰＧＥ２産生促進に対してβ−ＣＰＸは濃度依存的に抑制し（図５参照）、さらにβ−ＣＰＸはＣＯＸ−１の産生には影響を与えずにＩＬ−１βにより誘導されたＣＯＸ−２産生を抑制した（図６参照）。一方、β−ＣＰＸの細胞毒性について調べた結果、５０μＭのβ−ＣＰＸは細胞毒性を示したが、１μＭから２０μＭの間では全く細胞毒性を示さなかった（図７参照）。 [result]
In human synovial cells, ADAMTS-4 mRNA expression (see FIG. 1) and ADAMTS-5 mRNA expression (see FIG. 2) were promoted by IL-1β (10 ng / mL). In contrast, β-cryptoxanthin (β-CPX) suppressed ADAMTS-4 mRNA expression and ADAMTS-5 mRNA expression in a concentration-dependent manner from 1 μM to 10 μM. Also in chondrocytes, ADAMTS-4 mRNA expression was promoted by IL-1β, and β-CPX suppressed ADAMTS-4 mRNA expression in a concentration-dependent manner (see FIG. 3). Unlike the case of synovial cells, ADAMTS-5 mRNA expression was not affected by IL-1β, but β-CPX suppressed ADAMTS-5 mRNA expression in a concentration-dependent manner (see FIG. 4). In addition, as a result of examining the effect of β-CPX on PGE 2 production and expression of COX-1 and COX-2 in human synovial cells, β-CPX is dependent on the concentration of PGE 2 production by IL-1β. In addition, β-CPX suppressed COX-2 production induced by IL-1β without affecting COX-1 production (see FIG. 6). On the other hand, as a result of examining the cytotoxicity of β-CPX, 50 μM β-CPX showed cytotoxicity, but no cytotoxicity was observed between 1 μM and 20 μM (see FIG. 7).

［考察］
以上の結果から、天然物由来カロテノイド色素が、ヒト滑膜細胞および軟骨細胞において、軟骨基質アグリカンを特異的に分解するアグリカナーゼ−１（ＡＤＡＭＴＳ−４）および−２（ＡＤＡＭＴＳ−５）の発現を濃度依存的に抑制することが判明した。ＡＤＡＭＴＳ−４およびＡＤＡＭＴＳ−５は関節リウマチや変形性関節症における軟骨基質の破壊と密接に関連していることからして、天然物由来カロテノイド色素が関節破壊防御薬として有効であることが確認できる。
前掲の非特許文献６に記載されるように、アグリカナーゼは、アグリカンのみならず脳の構成プロテオグリカンであるブレビカンをも分解し、脳神経系のがん細胞である神経膠芽腫細胞の浸潤・転移にも深く関わっている。従って、本発明のアグリカナーゼ産生阻害剤は、これら脳神経系のがん細胞である神経膠芽腫細胞の浸潤・転移を抑制するための薬剤として有効であることが確認できる。
また、炎症メディエーターであるＰＧＥ２産生に対しても濃度依存的に抑制することが判明した。これらの事実は、本発明による天然物由来カロテノイド色素が、関節破壊防止物質として有効であることを立証している。
更に、本発明のアグリカナーゼ産生阻害剤は、ＣＯＸ−１の産生を抑制せず、ＣＯＸ−２の産生を選択的に抑制することができる。ＣＯＸ−１は正常組織において恒常的に産生され、例えば消化管粘膜保護作用に働くプロスタグランジンの産生に関与するのに対して、ＣＯＸ−２は炎症組織（変形性関節症，関節リウマチ等）や癌組織において産生誘導されることから、ＣＯＸ−２産生を選択的に阻害することは胃腸障害等の副作用が低い抗炎症薬となることを示唆する。 [Discussion]
Based on the above results, the natural product-derived carotenoid pigment has a concentration of expression of aggrecanase-1 (ADAMTS-4) and -2 (ADAMTS-5) that specifically degrade cartilage matrix aggrecan in human synovial cells and chondrocytes. It turned out to be dependently suppressed. Since ADAMTS-4 and ADAMTS-5 are closely related to the destruction of cartilage matrix in rheumatoid arthritis and osteoarthritis, it can be confirmed that natural product-derived carotenoid pigments are effective as joint destruction protective agents. .
As described in Non-Patent Document 6 described above, aggrecanase decomposes not only aggrecan but also brebican, which is a constituent proteoglycan of the brain, to infiltrate and metastasize glioblastoma cells, which are cancer cells of the cranial nervous system. Is also deeply involved. Therefore, it can be confirmed that the aggrecanase production inhibitor of the present invention is effective as a drug for suppressing invasion / metastasis of glioblastoma cells which are cancer cells of these cranial nervous systems.
Was also found to be dose-dependently inhibited against PGE 2 production is inflammation mediators. These facts prove that the natural product-derived carotenoid pigment according to the present invention is effective as a joint destruction preventing substance.
Furthermore, the aggrecanase production inhibitor of the present invention can selectively suppress the production of COX-2 without suppressing the production of COX-1. COX-1 is constitutively produced in normal tissues, and is involved in the production of prostaglandins that act to protect the gastrointestinal mucosa, whereas COX-2 is inflamed tissues (osteoarthritis, rheumatoid arthritis, etc.) Since the production is induced in cancer tissue, selective inhibition of COX-2 production suggests that it becomes an anti-inflammatory drug having low side effects such as gastrointestinal disorders.

本発明のアグリカナーゼ産生阻害剤は、関節リウマチや変形性関節症などの疾患の他、脳神経系のがん細胞である神経膠芽腫細胞の浸潤・転移の抑制及び肺癌、子宮頸部癌、食道癌、膀胱癌、大腸癌、皮膚癌などの発癌リスクの低減にも有用である。更に、骨関節症、関節損傷、若年性関節リウマチ、反応性関節炎、乾癬性関節炎、神経病性関節症（シャルコー関節）、血友病性関節症、感染性関節炎、強直性脊椎炎、ライター症候群、痛風、急性ピロリン酸関節炎（偽性通風）、炎症性腸疾患、クローン病、歯周疾患、骨粗鬆症及び人工関節インプラントの弛みなどの疾患に対しても有用である。更に、機能性食品としてこれら疾患の予防にも有用である。   In addition to diseases such as rheumatoid arthritis and osteoarthritis, the aggrecanase production inhibitor of the present invention suppresses infiltration / metastasis of glioblastoma cells, which are cancer cells of the cranial nervous system, and lung cancer, cervical cancer, esophagus It is also useful for reducing the risk of developing cancer such as cancer, bladder cancer, colon cancer, and skin cancer. Furthermore, osteoarthritis, joint damage, juvenile rheumatoid arthritis, reactive arthritis, psoriatic arthritis, neuropathic arthropathy (Charcot joint), hemophilic arthropathy, infectious arthritis, ankylosing spondylitis, Reiter syndrome It is also useful for diseases such as gout, acute pyrophosphate arthritis (pseudo ventilation), inflammatory bowel disease, Crohn's disease, periodontal disease, osteoporosis and loosening of artificial joint implants. Furthermore, it is useful for the prevention of these diseases as a functional food.

ヒト滑膜細胞におけるＡＤＡＭＴＳ−４ ｍＲＮＡ発現に対するβ−クリプトキサンチンの効果を説明する特性図であり、図中、レーン１は無処理（対照）、２はＩＬ−１β（１０ｎｇ／ｍＬ）、３はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１μｍｏｌ／Ｌ）、４はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（５μｍｏｌ／Ｌ）、５はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１０μｍｏｌ／Ｌ）、６はＩＬ−１β（１０ｎｇ／ｍＬ）＋デキサメタゾン（１μｍｏｌ／Ｌ）をそれぞれ示す。It is a characteristic view explaining the effect of β-cryptoxanthin on ADAMTS-4 mRNA expression in human synovial cells, in which lane 1 is untreated (control), 2 is IL-1β (10 ng / mL), 3 is IL-1β (10 ng / mL) + β-cryptoxanthin (1 μmol / L), 4 is IL-1β (10 ng / mL) + β-cryptoxanthin (5 μmol / L), 5 is IL-1β (10 ng / mL) + β− Cryptoxanthine (10 μmol / L), 6 represents IL-1β (10 ng / mL) + dexamethasone (1 μmol / L), respectively. ヒト滑膜細胞におけるＡＤＡＭＴＳ−５ ｍＲＮＡ発現に対するβ−クリプトキサンチンの効果を説明する特性図であり、図中、レーン１は無処理（対照）、２はＩＬ−１β（１０ｎｇ／ｍＬ）、３はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１μｍｏｌ／Ｌ）、４はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（５μｍｏｌ／Ｌ）、５はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１０μｍｏｌ／Ｌ）、６はＩＬ−１β（１０ｎｇ／ｍＬ）＋デキサメタゾン（１μｍｏｌ／Ｌ）をそれぞれ示す。It is a characteristic view explaining the effect of β-cryptoxanthin on ADAMTS-5 mRNA expression in human synovial cells, in which lane 1 is untreated (control), 2 is IL-1β (10 ng / mL), 3 is IL-1β (10 ng / mL) + β-cryptoxanthin (1 μmol / L), 4 is IL-1β (10 ng / mL) + β-cryptoxanthin (5 μmol / L), 5 is IL-1β (10 ng / mL) + β− Cryptoxanthine (10 μmol / L), 6 represents IL-1β (10 ng / mL) + dexamethasone (1 μmol / L), respectively. ヒト関節軟骨細胞におけるＡＤＡＭＴＳ−４ ｍＲＮＡ発現に対するβ−クリプトキサンチンの効果を説明する特性図であり、図中、レーン１は無処理（対照）、２はＩＬ−１β（１０ｎｇ／ｍＬ）、３はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１μｍｏｌ／Ｌ）、４はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（５μｍｏｌ／Ｌ）、５はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１０μｍｏｌ／Ｌ）、６はＩＬ−１β（１０ｎｇ／ｍＬ）＋デキサメタゾン（１μｍｏｌ／Ｌ）をそれぞれ示す。It is a characteristic view explaining the effect of β-cryptoxanthin on ADAMTS-4 mRNA expression in human articular chondrocytes, in which lane 1 is untreated (control), 2 is IL-1β (10 ng / mL), 3 is IL-1β (10 ng / mL) + β-cryptoxanthin (1 μmol / L), 4 is IL-1β (10 ng / mL) + β-cryptoxanthin (5 μmol / L), 5 is IL-1β (10 ng / mL) + β− Cryptoxanthine (10 μmol / L), 6 represents IL-1β (10 ng / mL) + dexamethasone (1 μmol / L), respectively. ヒト関節軟骨細胞におけるＡＤＡＭＴＳ−５ ｍＲＮＡ発現に対するβ−クリプトキサンチンの効果を説明する特性図であり、図中、レーン１は無処理（対照）、２はＩＬ−１β（１０ｎｇ／ｍＬ）、３はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１μｍｏｌ／Ｌ）、４はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（５μｍｏｌ／Ｌ）、５はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１０μｍｏｌ／Ｌ）、６はＩＬ−１β（１０ｎｇ／ｍＬ）＋デキサメタゾン（１μｍｏｌ／Ｌ）をそれぞれ示す。It is a characteristic view explaining the effect of β-cryptoxanthin on ADAMTS-5 mRNA expression in human articular chondrocytes, in which lane 1 is untreated (control), 2 is IL-1β (10 ng / mL), 3 is IL-1β (10 ng / mL) + β-cryptoxanthin (1 μmol / L), 4 is IL-1β (10 ng / mL) + β-cryptoxanthin (5 μmol / L), 5 is IL-1β (10 ng / mL) + β− Cryptoxanthine (10 μmol / L), 6 represents IL-1β (10 ng / mL) + dexamethasone (1 μmol / L), respectively. ヒト滑膜細胞におけるプロスタグランジン（ＰＧ）Ｅ２産生に対するβ−クリプトキサンチンの効果を説明する特性図であり、図中、レーン１は無処理（対照）、２はＩＬ−１β（１０ｎｇ／ｍＬ）、３はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１μｍｏｌ／Ｌ）、４はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（５μｍｏｌ／Ｌ）、５はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１０μｍｏｌ／Ｌ）、６はＩＬ−１β（１０ｎｇ／ｍＬ）＋デキサメタゾン（１μｍｏｌ／Ｌ）をそれぞれ示し、また、レーン２における＃＃はレーン１に対して有意差（ｐ＜０．０１）がある事を示し、レーン３〜６における＊＊はレーン２に対して有意差（ｐ＜０．０１）がある事を示す。It is a characteristic diagram for explaining the effect of β- cryptoxanthin for prostaglandins (PG) 2 production E in human synovial cells, in the figure, Lane 1 untreated (control), 2 IL-1β (10ng / mL ), 3 is IL-1β (10 ng / mL) + β-cryptoxanthin (1 μmol / L), 4 is IL-1β (10 ng / mL) + β-cryptoxanthin (5 μmol / L), 5 is IL-1β (10 ng / L) mL) + β-cryptoxanthin (10 μmol / L), 6 represents IL-1β (10 ng / mL) + dexamethasone (1 μmol / L), and ## in lane 2 is significantly different from lane 1 (p <0.01), and ** in lanes 3 to 6 indicates that there is a significant difference (p <0.01) from lane 2. ヒト滑膜細胞におけるシクロオキシゲナーゼ（ＣＯＸ）−１及び−２産生に対するβ−クリプトキサンチンの効果を説明する特性図であり、図中、レーン１は無処理（対照）、２はＩＬ−１β（１０ｎｇ／ｍＬ）、３はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１μｍｏｌ／Ｌ）、４はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（５μｍｏｌ／Ｌ）、５はＩＬ−１β（１０ｎｇ／ｍＬ）＋β−クリプトキサンチン（１０μｍｏｌ／Ｌ）、６はＩＬ−１β（１０ｎｇ／ｍＬ）＋デキサメタゾン（１μｍｏｌ／Ｌ）をそれぞれ示す。FIG. 2 is a characteristic diagram illustrating the effect of β-cryptoxanthin on cyclooxygenase (COX) -1 and -2 production in human synovial cells, in which lane 1 is untreated (control) and 2 is IL-1β (10 ng / mL), 3 is IL-1β (10 ng / mL) + β-cryptoxanthin (1 μmol / L), 4 is IL-1β (10 ng / mL) + β-cryptoxanthin (5 μmol / L), 5 is IL-1β (10 ng) / ML) + β-cryptoxanthin (10 μmol / L), 6 represents IL-1β (10 ng / mL) + dexamethasone (1 μmol / L), respectively. ヒト滑膜細胞の生存率に対するβ−クリプトキサンチンの効果を説明する特性図であり、図中、レーン１は無処理（対照）、２はβ−クリプトキサンチン（１μｍｏｌ／Ｌ）、３はβ−クリプトキサンチン（５μｍｏｌ／Ｌ）、４はβ−クリプトキサンチン（１０μｍｏｌ／Ｌ）、５はβ−クリプトキサンチン（２０μｍｏｌ／Ｌ）、６はβ−クリプトキサンチン（５０μｍｏｌ／Ｌ）をそれぞれ示し、レーン６における＊＊＊はレーン１に対して有意差（ｐ＜０．００１）がある事を示す。It is a characteristic view explaining the effect of β-cryptoxanthin on the survival rate of human synoviocytes, in which lane 1 is untreated (control), 2 is β-cryptoxanthin (1 μmol / L), 3 is β- Cryptoxanthine (5 μmol / L), 4 represents β-cryptoxanthin (10 μmol / L), 5 represents β-cryptoxanthin (20 μmol / L), 6 represents β-cryptoxanthin (50 μmol / L). *** indicates that there is a significant difference with respect to lane 1 (p <0.001).

Claims (5)

下記の一般式（１）、一般式（２）及び一般式（３）、



（前記各式中、Ｒ１は、水素、アルキル基、シクロアルキル基、アリール基、ヘテロアリール基、アラルキル基、ハロゲン、ヒドロキシ基又はアルコキシ基を示し、それらはヘテロ原子で置換されていてもよい；Ｒ２およびＲ２’は、水素、ヒドロキシ基又はアルコキシ基を示す。構造式中のアルケン水素はハロゲン原子で置換されていてもよい。）で示される化合物、その治療上許容し得る塩、エーテル、エステル及び異性体からなる群から選択される少なくとも一種類のカロテノイドを含有するアグリカナーゼ産生阻害剤。 The following general formula (1), general formula (2) and general formula (3),



(In the above formulas, R 1 represents hydrogen, an alkyl group, a cycloalkyl group, an aryl group, a heteroaryl group, an aralkyl group, a halogen, a hydroxy group, or an alkoxy group, which may be substituted with a heteroatom. R 2 and R 2 ′ represent hydrogen, a hydroxy group or an alkoxy group. The alkene hydrogen in the structural formula may be substituted with a halogen atom.), A therapeutically acceptable salt thereof, An aggrecanase production inhibitor containing at least one carotenoid selected from the group consisting of ethers, esters and isomers. 前記カロテノイドが、α−カロテン、α−クリプトキサンチン、ルテイン、β−カロテン、β−クリプトキサンチン、ゼアキサンチン、カンタキサンチン及びアスタキサンチンからなる群から選択されることを特徴とする請求項１記載のアグリカナーゼ産生阻害剤。 The aggrecanase production inhibition according to claim 1, wherein the carotenoid is selected from the group consisting of α-carotene, α-cryptoxanthin, lutein, β-carotene, β-cryptoxanthin, zeaxanthin, canthaxanthin and astaxanthin. Agent. 前記カロテノイドがβ−クリプトキサンチンであることを特徴とする請求項１又は２記載のアグリカナーゼ産生阻害剤。 The aggrecanase production inhibitor according to claim 1 or 2, wherein the carotenoid is β-cryptoxanthin. アグリカナーゼ関連疾患の治療及び／又は予防薬として使用することを特徴とする請求項１〜３の何れかに記載のアグリカナーゼ産生阻害剤。 The aggrecanase production inhibitor according to any one of claims 1 to 3, which is used as a therapeutic and / or prophylactic agent for an aggrecanase-related disease. アグリカナーゼ関連疾患が、関節リウマチ、変形性関節症、癌、骨関節症、関節損傷、若年性関節リウマチ、反応性関節炎、乾癬性関節炎、神経病性関節症（シャルコー関節）、血友病性関節症、感染性関節炎、強直性脊椎炎、ライター症候群、痛風、急性ピロリン酸関節炎（偽性通風）、炎症性腸疾患、クローン病、歯周疾患、骨粗鬆症及び人工関節インプラントの弛みからなる群から選択されることを特徴とする請求項４記載のアグリカナーゼ産生阻害剤。
Aggrecanase-related diseases include rheumatoid arthritis, osteoarthritis, cancer, osteoarthritis, joint damage, juvenile rheumatoid arthritis, reactive arthritis, psoriatic arthritis, neuropathic arthropathy (Charcot joint), hemophilic joint From infectious arthritis, infectious arthritis, ankylosing spondylitis, Reiter syndrome, gout, acute pyrophosphate arthritis (pseudo-ventilation), inflammatory bowel disease, Crohn's disease, periodontal disease, osteoporosis and loosening of artificial joint implants The aggrecanase production inhibitor according to claim 4, wherein
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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2011042627A (en) * 2009-08-21 2011-03-03 Unitika Ltd Hyaluronic acid synthesis promoter
WO2014175226A1 (en) * 2013-04-22 2014-10-30 国立大学法人東京農工大学 Prophylactic or therapeutic composition for bone diseases
JP2015163642A (en) * 2015-06-03 2015-09-10 ユニチカ株式会社 Hyaluronic acid synthesis promoter

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20040076691A1 (en) * 2002-01-16 2004-04-22 David Haines Anti-inflammatory formulations
JP2004210725A (en) * 2002-12-27 2004-07-29 Shonan Institute For Medical & Preventive Science Cyclooxygenase inhibitor and food product
JP2005034135A (en) * 2003-06-30 2005-02-10 Meiji Seika Kaisha Ltd Functionally reinforcing composition for common food, health functional food or health supplemental food, and method for the same
US20050261254A1 (en) * 2004-04-14 2005-11-24 Lockwood Samuel F Carotenoid analogs or derivatives for the inhibition and amelioration of inflammation
JP2006104090A (en) * 2004-10-01 2006-04-20 Unitika Ltd Anti-osteoporosis composition

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1081137A1 (en) * 1999-08-12 2001-03-07 Pfizer Products Inc. Selective inhibitors of aggrecanase in osteoarthritis treatment
SE9903619D0 (en) * 1999-10-07 1999-10-07 Astacarotene Ab Use and method of treatment

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20040076691A1 (en) * 2002-01-16 2004-04-22 David Haines Anti-inflammatory formulations
JP2004210725A (en) * 2002-12-27 2004-07-29 Shonan Institute For Medical & Preventive Science Cyclooxygenase inhibitor and food product
JP2005034135A (en) * 2003-06-30 2005-02-10 Meiji Seika Kaisha Ltd Functionally reinforcing composition for common food, health functional food or health supplemental food, and method for the same
US20050261254A1 (en) * 2004-04-14 2005-11-24 Lockwood Samuel F Carotenoid analogs or derivatives for the inhibition and amelioration of inflammation
JP2006104090A (en) * 2004-10-01 2006-04-20 Unitika Ltd Anti-osteoporosis composition

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2011042627A (en) * 2009-08-21 2011-03-03 Unitika Ltd Hyaluronic acid synthesis promoter
WO2014175226A1 (en) * 2013-04-22 2014-10-30 国立大学法人東京農工大学 Prophylactic or therapeutic composition for bone diseases
JPWO2014175226A1 (en) * 2013-04-22 2017-02-23 国立大学法人東京農工大学 Composition for prevention or treatment of bone disease
JP2015163642A (en) * 2015-06-03 2015-09-10 ユニチカ株式会社 Hyaluronic acid synthesis promoter

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