| PATENT NUMBER | This data is not available for free |
| PATENT GRANT DATE | 31.12.02 |
| PATENT TITLE |
Chondrongenesis promotors and indolin-2-one derivatives |
| PATENT ABSTRACT |
A chondrogenesis promoter comprising as an active ingredient a compound represented by general formula (I) or a salt thereof: ##STR1## wherein R.sup.1 represents a halogen atom, a lower alkyl group, a lower alkoxy group, etc.; R.sup.2 represents a hydrogen atom, a lower alkyl group, a lower alkenyl group, a lower alkynyl group, a lower alkoxy group, an acyl group, an aryl group, a heterocyclic group, etc.; R.sup.1 represents a lower alkyl group, a cycloalkyl group, an aryl group, a heterocyclic group, etc.; R.sup.4 represents a hydrogen atom, a lower alkyl group, an aryl group, a heterocyclic group, etc.; X and Y represent --CH.sub.2 --, --NH--or --O--; and n represents an integer of 0-4. |
| PATENT INVENTORS | This data is not available for free |
| PATENT ASSIGNEE | This data is not available for free |
| PATENT FILE DATE | September 21, 2001 |
| PATENT CT FILE DATE | January 26, 2000 |
| PATENT CT NUMBER | This data is not available for free |
| PATENT CT PUB NUMBER | This data is not available for free |
| PATENT CT PUB DATE | August 3, 2000 |
| PATENT FOREIGN APPLICATION PRIORITY DATA | This data is not available for free |
| PATENT REFERENCES CITED |
Masayuki Azuma, et al., "Induction of Cells with a Chondrocyte-like Phenotype by Treatment with 1.alpha.,25-Dihydroxyvitamin D.sub.3 a Human Salivary Acinar Cell Line," Cancer Research, vol. 49, Oct. 1, 1989, pp. 5435-5442. Warner M. Burch and Harold E. Lebovitz, "Triiodothyronine Stimulation of in Vitro Growth and Maturation of Embryonic Chick Cartilage," Endocrinology, vol. 111, No. 2, 1982, pp. 462-468. Paul Dieppe, "Therapeutic Targets in Osteoarthritis," The Journal of Rheumatology, vol. 22:1, Supplement 43, 1995, pp. 136-139. Harrie L. Glansbeek, et al., "Stimulation of Articular Cartilage Repair in Established Arthritis by Local Administration of Transforming Growth Factor-.beta. into Murine Knee Joints," Laboratory Investigation, vol. 78, No. 2, Feb. 1998, pp. 133-142. H. Ide and H. Aono, "Retonic Acid Promotes Proliferation and Chondrogenesis in the Distal Mesodermal Cells of Chick Limb Bud," Development Biology, vol. 130, 1998, pp. 767-773. Yukio Kato, et al., "Effect of Vanadate on Cartilage-Matrix Proteoglycan Synthesis in Rabbit Coastal Chondrocyte Cultures," The Journal of Cell Biology, vol. 104, Feb. 1987, pp. 311-319. Lawrence Kent, Charles J. Malemud and Roland W. Moskowitz, "Differential Response of Articular Chondrocyte Populations to Thromboxane B.sub.2 and Analogs of Prostaglandin Cyclic Endoperoxides," Prostaglandins, Vo. 19, No. 3, Mar. 1980, pp. 391-406. H. Kitamura et al., "AG-041R, A Novel Indoline-2-One Derivative, Induces Systemic Cartilage Hyperplasia in Rats", Eur. J. Pharmacol, vol. 418, No. 3, 2001, pp. 225-230, XP002187328. (Abstract Only). David J. McQuillan, et al., "The Relation of Protein Synthesis to Chondroitin Sulphate Biosynthesis in Cultured Bovine Cartilage," Biochem. J., vol. 224, 1984, pp. 977-988. R. P. Miller, M. Husain, and S. Lohin, "Long Acting cAMP Analogues Enhance Sulfate Incorporation into Matrix Proteoglycans and Suppress Cell Division of Fetal Ral Chondrocytes in Monolayer Culture," J. Cell. Physiol., vol. 100, 1979, pp. 63-76. Shinobu Nakanishi and Edwin M. Uyeki, "Benzamide on Chondrocytic Differentiation in Chick Limb Bud Cell Culture," J. Embryol. exp. Morph., vol. 85, 1985, pp. 163-175. Rani S. Sellers, Diane Peluso, and Elisabeth A. Morris, "The Effect of Recombinant Human Bone Morphogenetic Protein-2 (rhBMP-2) on the Healing of Full-Thickness Defects of Articular Cartilage," The Journal of Bone and Joint Surgery, vol. 79-A, No. 10, Oct. 1997, pp. 1452-1463. Y. Shimomura, T. Yoneda, and F. Suzuki, "Osteogenesis by Chondrocytes from Growth Cartilage of Rat Rib," Calcif. Tiss. Res., vol. 19, 1975, pp. 179-187. Henk M. Van Beuningen, et al., "Transforming Growth Factor-.beta.1 Stimulates Articular Chondrocyte Proteoglycan Synthesis and Induces Osteophyte Formation in the Murine Knee Joint," Laboratory Investigation, vol. 71, No. 2, 1994, pp. 279-290. Shigeyuki Wakitani, et al., "Mesenchymal Cell-Based Repair of Large Full-Thickness Defects of Articular Cartilage", The Journal of Bone and Joint Surgery, vol. 76-A, No. 4, Apr. 1994, pp. 579-592. Weiqun Yan, et al., "Stimulation by Concanavalin A of Cartilage-Matrix Proteoglycan Synthesis in Chondrocyte Cultures," The Journal of Biological Chemistry, vol. 265, No. 17, Jun. 15, 1990, pp. 10125-10131. |
| PATENT CLAIMS |
What is claimed is: 1. A method for promoting chondrogenesis comprising oral or parenteral administration, to a patient in need thereof, of an effective amount of an agent comprising as an active ingredient a compound represented by general formula (I) or a salt thereof: ##STR172## wherein R.sup.1 represents a halogen atom, a lower alkyl group, a lower alkoxy group, a hydroxyl group, a nitro group, a trifluoromethyl group, a lower alkylthio group, an acyl group, a carboxyl group, a mercapto group or an amino group with an optional substituent; R.sup.2 represents a hydrogen atom, a lower alkyl group with an optional substituent, a lower alkenyl group with an optional substituent, a lower alkynyl group with an optional substituent, a lower alkoxy group with an optional substituent, an acyl group with an optional substituent, an aryl group with an optional substituent or a heterocyclic group with an optional substituent; R.sup.3 represents a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent, an aryl group with an optional substituent or a heterocyclic group with an optional substituent; R.sup.4 represents a hydrogen atom, a lower alkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, --OR.sup.5, --SR.sup.5 or --NR.sup.6 NR.sup.7 wherein R.sup.5, R.sup.6 and R.sup.7 may be the same or different and each represents a hydrogen atom, a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, a lower alkoxy group or an amino group with an optional substituent, or R.sup.6 and R.sup.7 may together form a group represented by --(CH.sub.2).sub.m -- or --(CH.sub.2).sub.1 NR.sup.8 (CH.sub.2).sub.k -- wherein k, 1 and m each represents an integer of 1-8 and R.sup.8 represents a hydrogen atom or a lower alkyl group; X and Y may be the same or different and each represents --CH.sub.2 --, --NH-- or --O--; and n represents an integer of 0-4. 2. A method for promoting chondrogenesis according to claim 1, wherein R.sup.1 is a halogen atom, a lower alkyl group, a lower alkoxy group or a nitro group; R.sup.2 is a hydrogen atom, a lower alkyl group with an optional substituent, a lower alkenyl group with an optional substituent or an aryl group with an optional substituent; R.sup.3 is a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent or an aryl group with an optional substituent; R.sup.4 is a lower alkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, --OR.sup.5 or --NR.sup.6 R.sup.7 wherein R.sup.5, R.sup.6 and R.sup.7 are as previously defined; X is --CH.sub.2 --, --NH-- or --O--; Y is --CH.sub.2 -- or --NH--; and n is 0 or 1. 3. A method for promoting chondrogenesis according to claim 1, wherein R.sup.2 is a lower alkyl group with an optional substituent which is optionally substituted with a halogen atom; R.sup.3 is an aryl group with an optional substituent; R.sup.4 is --NR.sup.6 R.sup.7 wherein R.sup.6 and R.sup.7 may be the same or different and each represents a hydrogen atom or an aryl group with an optional substituent; X and Y may be the same or different and each represents --CH.sub.2 -- or --NH--; and n is 0. 4. A method for promoting chondrogenesis according to claim 3, wherein R.sup.2 is a lower alkyl group substituted at the same carbon with two lower alkoxy groups which are optionally substituted with 1-5 halogen atoms or the group --O--Z--O-- wherein Z represents a lower alkylene group optionally substituted with 1-10 halogen atoms; R.sup.3 is an aryl group which has a lower alkyl group or an amino group which amino group optionally has a lower alkyl group; and R.sup.4 is --NR.sup.6 R.sup.7 wherein R.sup.6 and R.sup.7 may be the same or different and each represents a hydrogen atom or an aryl group which has a lower alkyl group or an amino group which amino group optionally has a lower alkyl group. 5. A method for promoting chondrogenesis according to claim 4, wherein R.sup.2 is a group represented by general formula (II): ##STR173## wherein R.sup.10 and R.sup.11 may be the same or different, and each represents a lower alkyl group optionally substituted with 1-5 halogen atoms, or general formula (III): ##STR174## wherein Z represents a lower alkylene group optionally substituted with 1-10 halogen atoms. 6. A method for promoting chondrogenesis according to claim 5, wherein either or both R.sup.10 and R.sup.11 are lower alkyl groups with 1-5 halogen atoms. 7. A method for promoting chondrogenesis according to claim 5, wherein R.sup.2 is a 2,2-diethoxyethyl group, a 2,2-dimethoxyethyl group, a 2,2-diisopropoxyethyl group, a 2,2-bis(2-fluoroethoxy)ethyl group or a 2,2-bis(2-chloroethoxy)ethyl group; X is --NH--; and Y is --CH.sub.2 --. 8. A method for promoting chondrogenesis according to claim 5, wherein R.sup.3 is a 4-methylphenyl group; X is --NH--; and Y is --CH.sub.2 --. 9. A method for promoting chondrogenesis according to claim 5, wherein R.sup.4 is --NHR.sup.7 wherein R.sup.7 is a 4-methylphenyl group, a 4-ethylphenyl group or a 4-(N,N-dimethylamino)phenyl group; X is --NH--; and Y is --CH.sub.2 --. 10. A method for promoting chondrogenesis according to claim 5, wherein the combination of R.sup.2, R.sup.3 and R.sup.4 is any of the following: R.sup.2 is a 2,2-diethoxyethyl group, R.sup.3 is a 4-methylphenyl group and R.sup.4 is --NHR.sup.7 wherein R.sup.7 is a 4-methylphenyl group; R.sup.2 is a 2,2-diethoxyethyl group, R.sup.3 is a 4-methylphenyl group and R.sup.4 is --NHR.sup.7 wherein R.sup.7 is a 4-ethylphenyl group; R.sup.2 is a 2,2-diethoxyethyl group, R.sup.3 is a 4-methylphenyl group and R.sup.4 is --NHR.sup.7 wherein R.sup.7 is a 4-(N,N-dimethylamino)phenyl group; R.sup.2 is a 2,2-dimethoxyethyl group, R.sup.3 is a 4-methylphenyl group and R.sup.4 is --NHR.sup.7 wherein R.sup.7 is a 4-methylphenyl group; R.sup.2 is a 2,2 diisopropoxyethyl group, R.sup.3 is a 4-methylphenyl group and R.sup.4 is --NHR.sup.7 wherein R.sup.7 is a 4-methylphenyl group; R.sup.2 is a 2,2-bis(2-fluoroethoxy)ethyl group, R.sup.3 is a 4-methylphenyl group and R.sup.4 is --NHR.sup.7 wherein R.sup.7 is a 4-methylphenyl group; or R.sup.2 is a 2,2-bis(2-chloroethoxy)ethyl group, R.sup.3 is a 4-methylphenyl group and R.sup.4 is --NHR.sup.7 wherein R.sup.7 is a 4-methylphenyl group. 11. A method for promoting chondrogenesis comprising as an active ingredient an optically active form of a compound according to claim 1, or a salt thereof. 12. A method for repairing cartilage comprising oral or parenteral administration, to a patient in need thereof, of an effective amount of an agent comprising as an active ingredient a compound according to claim 1. 13. A method for promoting bone fracture repair comprising oral or parenteral administration, to a patient in need thereof, of an effective amount of an agent comprising as an active ingredient a compound according to claim 1. 14. An indolin-2-one derivative represented by the following general formula (IV): ##STR175## wherein R.sup.12 represents a lower alkyl group substituted at the same carbon with two lower alkoxy groups which is substituted with 1-5 halogen atoms, or a salt thereof. 15. An indolin-2-one derivative or a salt thereof according to claim 14, wherein R.sup.12 is a group represented by general formula (V): ##STR176## wherein R.sup.13 and R.sup.14 may be the same or different, and each represents a lower alkyl group substituted with 1-5 halogen atoms. 16. An indolin-2-one derivative or a salt thereof according to claim 15, wherein R.sup.13 and R.sup.14 may be the same or different, and each represents an ethyl group substituted with a halogen atom. 17. An indolin-2-one derivative or a salt thereof according to claim 15, wherein R.sup.12 is a 2,2-bis(2-fluoroethoxy)ethyl group or a 2,2-bis(2-chloroethoxy)ethyl group. 18. A method for promoting chondrogenesis comprising contacting an agent comprising as an active ingredient a compound represented by the general formula (I) or a salt thereof with a chondrocyte or a precursor thereof: ##STR177## wherein R.sup.1 represents a halogen atom, a lower alkyl group, a lower alkoxy group, a hydroxyl group, a nitro group, a trifluoromethyl group, a lower alkylthio group, an acyl group, a carboxyl group, a mercapto group or an amino group with an optional substituent; R.sup.2 represents a hydrogen atom, a lower alkyl group with an optional substituent, a lower alkenyl group with an optional substituent, a lower alkynyl group with an optional substituent, a lower alkoxy group with an optional substituent, an acyl group with an optional substituent, an aryl group with an optional substituent or a heterocyclic group with an optional substituent; R.sup.3 represents a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent, an aryl group with an optional substituent or a heterocyclic group with an optional substituent; R.sup.4 represents a hydrogen atom, a lower alkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, --OR.sup.5, --SR.sup.5 or --NR.sup.6 NR.sup.7 wherein R.sup.5, R.sup.6 and R.sup.7 may be the same or different and each represents a hydrogen atom, a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, a lower alkoxy group or an amino group with an optional substituent, or R.sup.6 and R.sup.7 may together form a group represented by --(CH.sub.2).sub.m -- or --(CH.sub.2).sub.1 NR.sup.8 (CH.sub.2).sub.k -- wherein k, l and m each represents an integer of 1-8 and R.sup.8 represents a hydrogen atom or a lower alkyl group; X and Y may be the same or different and each represents --CH.sub.2 --, --NH-- or --O--; and n represents an integer of 0-4. -------------------------------------------------------------------------------- |
| PATENT DESCRIPTION |
TECHNICAL FIELD The present invention relates to pharmaceutically useful indolin-2-one derivatives or their salts, and to chondrogenesis promoters, cartilage repair agents and cartilage diagnostic reagents containing the indolin-2-one derivatives or their pharmaceutically acceptable salts. BACKGROUND ART Cartilage in the body generally consists of chondrocytes and fibrocytes, which are specialized connective tissue cells, and an amorphous gel-like matrix in which they are embedded, and it forms a part of the supportive tissue of the body. In warm-blooded animals including humans, cartilage forms the skeleton, joints, tracheae, auricula, nose and the like. That is, it performs a central role in functions that are indispensable to the survival of warm-blooded animals, including acting as a template for bone during growth (growth cartilage), and contributing to smooth joint movement (articular cartilage), respiration (tracheal cartilage, nasal cartilage) and hearing (auricular cartilage). Thus, degeneration or destruction of these types of cartilage causes various degrees of detriment to the body depending on the site and severity of degeneration or destruction. For example, among the aforementioned functions in which cartilage plays a role (growth, joints, respiration, hearing, etc.), the smooth movement of joints is particularly impaired by degeneration or destruction of articular cartilage in such conditions as chronic rheumatoid arthritis or osteoarthritis. Degeneration or destruction of articular cartilage is believed to be the major cause of the walking difficulty that results from such diseases. The prospect of suppressing articular degeneration or destruction or of promoting chondrogenesis has been raised as a possible method of treating conditions such as chronic rheumatoid arthritis and osteoarthritis (J. Rheum. 22(1), Suppl. 43:136-139, 1995, Lab. Invest. 78(2):133-142). Several different organism-derived substances and low molecular substances are known to have effects of promoting chondrogenesis or of inducing proliferation of chondrocytes. Substances that have been reported to have chondrogenesis-promoting effects include growth factors such as TGF-.beta., (Transforming growth factor .beta.), BMP-2 (J. Bone Joint Surg. 79-A(10):1452-1463, 1997), concanavalin A which is a type of lectin (J. Biol. Chem., 265:10125-10131, 1990) and osteogenin (BMP-7), as well as low molecular substances such as vitamin D derivatives (1.alpha., 25-D.sub.3) (Cancer Res., 49:5435-5442, 1989), vitamin A derivatives (retinoic acid) (Dev. Biol., 130:767-773, 1988), vanadates (J. Cell Biol., 104:311-319, 1987), benzamides (J. Embryol. Exp. Morphol., 85:163-175, 1985), benzyl .beta.-D-xyloside (Biochem. J., 224:977-988, 1984), triiodothyronines (T.sub.3) (Endocrinology, 111:462-468, 1982), prostaglandin derivatives (PGE2, U44069) (Prostaglandin, 19:391-406, 1980), dbcAMP (J. Cell. Physiol., 100:63-76, 1979) and 8-Br-cAMP(J. Cell. Physiol., 100:63-76, 1979). Of these organism-derived substances and low molecular substances, TGF-.beta., holds the most promise as a useful treatment agent, and TGF-.beta..sub.1, which is one isoform of TGF-.beta., has been reported to promote chondrogenesis when intraarticularly administered (Lab. Invest. 71(2):279-290, 1994). Also, since TGF-.beta..sub.1 suppresses arthritis-induced loss of proteoglycans in articular cartilage, or stated differently, it inhibits destruction of articular cartilage due to its anabolic effect on articular cartilage when administered intraarticularly in experimental animal models with induced arthritis, its possibility as a useful treatment agent for articular disease such as rheumatism has been suggested (Lab. Invest. 78(2):133-142, 1998). However, even TGF-.beta. which holds the most promise as a useful treatment agent has been reported to provoke synovitis even while promoting chondrogenesis, and this therefore poses a serious problem for its use as a treatment agent for articular diseases (Lab. Invest. 71(2):279-290, 1994), for which reason it has not been applied in the clinic as a treatment agent for such conditions. In summary, then, no practical treatment agent therapy exists that is based on promoting chondrogenesis. DISCLOSURE OF THE INVENTION It is an object of the present invention to overcome the aforementioned drawbacks of the prior art by providing a chondrogenesis promoter and cartilage repair agent that are able to promote chondrogenesis or induce proliferation of chondrocytes. It is another object of the invention to provide a reagent with chondrogenesis promoting action which is useful for biological, physical or chemical research on cartilage. It is yet another object of the invention to provide indolin-2-one derivatives that are useful as chondrogenesis promoters. It is still yet another object of the invention to provide indolin-2-one derivatives that are useful as bone fracture repair promoters. As a result of diligent research aimed at achieving these objects, the present inventors have completed the present invention upon the discovery that indolin-2-one derivatives having a specific structure exhibit a chondrogenesis promoting effect. In other words, a chondrogenesis promoter according to the invention comprises as an active ingredient a compound represented by general formula (I) or a salt thereof: ##STR2## wherein R.sup.1 represents a halogen atom, a lower alkyl group, a lower alkoxy group, a hydroxyl group, a nitro group, a trifluoromethyl group, a lower alkylthio group, an acyl group, a carboxyl group, a mercapto group or an amino group with an optional substituent; R.sup.2 represents a hydrogen atom, a lower alkyl group with an optional substituent, a lower alkenyl group with an optional substituent, a lower alkynyl group with an optional substituent, a lower alkoxy group with an optional substituent, an acyl group with an optional substituent, an aryl group with an optional substituent or a heterocyclic group with an optional substituent; R.sup.3 represents a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent, an aryl group with an optional substituent or a heterocyclic group with an optional substituent; R.sup.4 represents a hydrogen atom, a lower alkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, --OR.sup.5, --SR.sup.5 or --NR.sup.6 R.sup.7 wherein R.sup.5, R.sup.6 and R.sup.7 may be the same or different and each represents a hydrogen atom, a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, a lower alkoxy group or an amino group with an optional substituent, and R.sup.6 and R.sup.7 may together form a group represented by --(CH.sub.2).sub.m -- or --(CH.sub.2).sub.1 NR.sup.8 (CH.sub.2).sub.k -- wherein k, 1 and m each represent an integer of 1-8 and R.sup.8 represents a hydrogen atom or a lower alkyl group; X and Y may be the same or different and each represents --CH.sub.2 --, --NH--or --O--, and n represents an integer of 0-4. A cartilage repair agent according to the invention also comprises as an active ingredient a compound represented by general formula (I) above or a salt thereof. A reagent for biological, physical or chemical research on cartilage according to the invention also comprises as an active ingredient a compound represented by general formula (I) above or a salt thereof. An indolin-2-one derivative according to the invention is represented by the following general formula (IV): ##STR3## wherein R.sup.12 represents a lower alkyl group substituted at the same carbon with two lower alkoxy groups which is optionally substituted with 1-5 halogen atoms. Also, R.sup.12 of the indolin-2-one derivative according to the invention may be represented by general formula (V): ##STR4## wherein R.sup.13 and R.sup.14 may be the same or different, and each represents a lower alkyl group which is optionally substituted with 1-5 halogen atoms. BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1A is a photomicrograph showing a hematoxylin/eosin double stained auricular tissue sample from a rat after repeated oral administration of a vehicle control (3% gum Arabic) for 4 weeks, and FIG. 1B is a photomicrograph showing a hematoxylin/eosin double stained auricular tissue sample from a rat after repeated oral administration of compound A (2 g/kg) for 4 weeks. FIG. 2A is a photomicrograph showing a hematoxylin/eosin double stained trachea tissue sample from a rat after repeated oral administration of a vehicle control (3% gum Arabic) for 4 weeks, and FIG. 2B is a photomicrograph showing a hematoxylin/eosin double stained trachea tissue sample from a rat after repeated oral administration of compound A (2 g/kg) for 4 weeks. FIG. 3A is a photomicrograph showing a hematoxylin/eosin double stained sternal xiphoid process tissue sample from a rat after repeated oral administration of a vehicle control (3% gum Arabic) for 4 weeks, and FIG. 3B is a photomicrograph showing a hematoxylin/eosin double stained sternal xiphoid process tissue sample from a rat after repeated oral administration of compound A (2 g/kg) for 4 weeks. FIG. 4A is a photomicrograph showing a hematoxylin/eosin double stained knee joint tissue sample from a rat after repeated oral administration of a vehicle control (3% gum Arabic) for 4 weeks, and FIG. 4B is a photomicrograph showing a hematoxylin/eosin double stained knee joint tissue sample from a rat after repeated oral administration of compound A (2 g/kg) for 4 weeks. FIG. 5A is a photomicrograph showing a hematoxylin/eosin double stained lumbar spine (disk) tissue sample from a rat after repeated oral administration of a vehicle control (3% gum Arabic) for 4 weeks, and FIG. 5B is a photomicrograph showing a hematoxylin/eosin double stained lumbar spine (disk) tissue sample from a rat after repeated oral administration of compound A (2 g/kg) for 4 weeks. FIG. 6 is a graph of the width of the femoral knee joint of a rat after repeated administration of compound A (1.0 mmol/L) or a vehicle control (50% DMSO physiological saline solution) at 50 .mu.l per day into the knee joint for 3 weeks. FIG. 7A is a photomicrograph showing a 0.3% Safranin O stained femoral knee joint tissue sample from a rat after repeated administration of a vehicle control (50% DMSO physiological saline solution) at 50 .mu.l per day into the knee joint for 3 weeks, and FIG. 7B is a photomicrograph showing a 0.3% Safranin O stained femoral knee joint tissue sample from a rat after repeated administration of compound A (1.0 mmol/L) at 50 .mu.l per day into the knee joint for 3 weeks. FIG. 8 is a graph showing the uptake of .sup.35 S-labeled sulfuric acid into glycosaminoglycans in rat primary culture articular chondrocytes to which compound A or a vehicle control (ethanol at 1% final concentration in medium) was added. FIG. 9 is a graph showing the uptake of .sup.3 H-labeled thymidine in rat primary culture articular chondrocytes to which compound A or a vehicle control (ethanol at 1% final concentration in medium) was added. FIG. 10A is a photomicrograph showing alcian blue/oil red O double stained confluent CL-1 cells after addition of a vehicle control (ethanol at 1% final concentration in medium) and culturing for 7 days, and FIG. 10B is a photomicrograph showing alcian blue/oil red O double stained confluent CL-1 cells after addition of compound A (10 .mu.mol/l final concentration in medium) and culturing for 7 days. FIG. 11 is a graph showing uptake of .sup.35 S-labeled sulfuric acid into confluent CL-1 cells during the final 24 hours of culturing of the CL-1 cells for 48 hours in medium containing compounds A and F at final concentrations of 1, 5 and 10 .mu.mol/L, compounds B and E at a final concentration of 10 .mu.mol/L or a vehicle control with a final concentration of 1% ethanol. FIG. 12 is a graph showing uptake of .sup.35 S-labeled sulfuric acid into confluent CL-1 cells during the final 24 hours of culturing of the CL-1 cells for 48 hours in medium containing compounds A, C, D and G at final concentrations of 10 .mu.mol/L or a vehicle control with a final concentration of 1% ethanol. FIG. 13A and FIG. 13B are graphs showing the results from histological examination of cartilage repair in deficient areas of the femoral patellar surface of rats with deficient areas reaching to the marrow were created, given repeated administration of compound A (1.0 mmol/L) or a vehicle control (50% DMSO physiological saline solution) at 50 .mu.L per day for 3 weeks beginning on the 7th day after the operation. FIG. 13A shows the scores for cell morphology, matrix-staining and thickness of cartilage, and FIG. 13B shows the total scores. FIG. 14A is a photomicrograph showing a 0.3% Safranin O stained tissue sample of a deficient area created in the femoral patellar surface reaching to the marrow of a rat given repeated intraarticular administration of a vehicle control (50% DMSO physiological saline solution) at 50 .mu.L per day for 3 weeks beginning on the 7th day after the operation, and FIG. 14B is a photomicrograph showing a 0.3% Safranin O stained tissue sample of a deficient area created in the femoral patellar surface reaching to the marrow of a rat given repeated intraarticular administration of compound A (1.0 mmol/L) at 50 .mu.L per day for 3 weeks beginning on the 7th day after the operation. FIG. 15A is a set of scanning electron photomicrographs showing partially menisectomized rabbits given repeated intraarticular administration of a vehicle control (50% DMSO physiological saline solution) at 500 .mu.L per day for 3 weeks beginning on the 7th day after the operation, and FIG. 15B is a set of scanning electron photomicrographs showing partially menisectomized rabbits given repeated intraarticular administration of compound A (3.0 mmol/L) at 500 .mu.L per day for 3 weeks beginning on the 7th day after the operation. FIG. 15A and FIG. 15B both contain photomicrographs of 6 individuals. FIG. 16A, FIG. 16B and FIG. 16C are graphs showing the injured articular cartilage surface area from scanning electron photomicrographs of partially menisectomized rabbits given repeated intraarticular administration of compound A (3.0 mmol/L) and a vehicle control (50% DMSO physiological saline solution) at 500 .mu.L per day for 3 weeks beginning on the 7th day after the operation. FIG. 16A shows the average area of mild lesions, FIG. 16B shows the average area of medium lesions and FIG. 16C shows the average of the total lesion area as the sum of the mild and medium lesion areas. FIG. 17A and FIG. 17B are graphs showing the results of histological examination of 0.3% Safranin O stained samples of lesions in partial excisions from the articular crescents of rabbits given repeated intraarticular administration of compound A (3.0 mmol/L) and a vehicle control (50% DMSO physiological saline solution) at 500 .mu.L per day for 3 weeks beginning on the 7th day after the operation. FIG. 17A shows the scores for loss of superficial layer, ulceration or erosion, fibrillation and cluster formation, and FIG. 17B shows the global assessment score. BEST MODE FOR CARRYING OUT THE INVENTION The present invention will now be explained in greater detail with reference to the attached drawings as appropriate. The amounts indicated as "parts" and "%" values in the following explanations are based on weight unless otherwise specified. A chondrogenesis promoter or cartilage repair agent according to the invention comprises as an active ingredient a compound represented by the aforementioned general formula (I) or a salt thereof. Compounds represented by general formula (I) according to the invention are described in WO094/19322, and the same patent publication teaches that the compounds are CCK-B/gastrin receptor antagonists. The present inventors have found that compounds represented by general formula (I) have an unexpected chondrogenesis promoting effect, and the present invention has been completed on the basis of this discovery. The compounds represented by general formula (I) of the invention may be obtained by the method described in the aforementioned international patent publication, or by the method illustrated hereunder in the examples. A "halogen atom" according to the invention is a fluorine atom, a chlorine atom, a bromine atom or an iodine atom. A "lower alkyl group" according to the invention is a linear or branched alkyl group of 1 to 6 carbons, such as a methyl group, an ethyl group, a n-propyl group, an i-propyl group, a n-butyl group, a s-butyl group, a t-butyl group, a pentyl group, a hexyl group or the like. A "lower alkenyl group" is a linear or branched alkenyl group of 2 to 6 carbons, such as a vinyl group, an allyl group, a butenyl group, a pentenyl group, a hexenyl group or the like. A "lower alkynyl group" is a linear or branched alkynyl group of 2 to 6 carbons, such as an ethynyl group, a propynyl group, a butynyl group or the like. A "lower alkoxy group" is a linear or branched alkoxy group of 1 to 6 carbons, such as a methoxy group, an ethoxy group, a n-propoxy group, an i-propoxy group, a n-butoxy group, a s-butoxy group, a t-butoxy group, a pentoxy group, a hexoxy group or the like. An "acyl group" is a carbonyl group substituted with a hydrogen atom or with an alkyl group with an optional substituent, an aryl group with an optional substituent, an alkoxy group with an optional substituent, an amino group with an optional substituent or the like, for example, an alkylcarbonyl group such as an acetyl group, a propionyl group, a pivaloyl group, a cyclohexanecarbonyl group or the like, or an arylcarbonyl group such as a benzoyl group, a naphthoyl group, a toluoyl group or the like. An "aryl group" is a monovalent group which is an aromatic hydrocarbon minus one hydrogen atom, such as a phenyl group, a tolyl group, a xylyl group, a biphenyl group, a naphthyl group, an anthryl group, a phenanthryl group or the like. A "lower alkylene group" is a linear or branched alkylene group of 1 to 6 carbons, such as a methylene group, an ethylene group, a propylene group, a butylene group, a pentylene group, a hexylene group or the like. A "cycloalkyl group" is a cyclic saturated hydrocarbon group of 3 to 8 carbons, such as a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, a cyclohexyl group or a cycloheptyl group. And, substituted cycloalkyl groups include menthyl group, adamantyl group and the like. A "heterocyclic group" is an aromatic heterocyclic group with at least one hetero atom, such as a pyridyl group, a furyl group, a thienyl group, an imidazolyl group, a pyrazinyl group, a pyrimidyl group or the like. The aforementioned lower alkyl group, lower alkenyl group, alkynyl group, lower alkoxy group, acyl group, aryl group, cycloalkyl group and heterocyclic group may, if necessary, be substituted with one or more substituents. As examples of such substituents there may be a halogen atom, a lower alkyl group, a cycloalkyl group, an aryl group, a hydroxyl group, a lower alkoxy group which alkoxy group may be substituted with a halogen atom, an aryloxy group, a lower alkylthio group, a heterocyclic group, a formyl group which formyl group may be protected as an acetal, a lower alkylcarbonyl group, an arylcarbonyl group, a carboxyl group, a lower alkoxycarbonyl group, an amino group which amino group may have a lower alkyl group, etc., an imino group, a thioacetal group, a nitro group, a nitrile group, a trifluoromethyl group and the like. The compounds serving as the active ingredients in the chondrogenesis promoters and cartilage repair agents of the invention (indolin-2-one derivatives represented by the above general formula (I)) will now be explained in greater detail. R.sup.1 represents a halogen atom, a lower alkyl group, a lower alkoxy group, a hydroxyl group, a nitro group, a trifluoromethyl group, a lower alkylthio group, an acyl group, a carboxyl group, a mercapto group or an amino group with an optional substituent, and among these, R.sup.1 is preferably a halogen atom, a lower alkyl group, a lower alkoxy group or a nitro group. The subscript "n" represents an integer of 0 to 4. It is preferably 0 or 1, and most preferably 0. R.sup.2 represents a hydrogen atom, a lower alkyl group with an optional substituent, a lower alkenyl group with an optional substituent, a lower alkynyl group with an optional substituent, a lower alkoxy group with an optional substituent, an acyl group with an optional substituent, an aryl group with an optional substituent or a heterocyclic group with an optional substituent. R.sup.2 is preferably a hydrogen atom, a lower alkyl group with an optional substituent, a lower alkenyl group with an optional substituent or an aryl group with an optional substituent, and from the viewpoint of activity as a chondrogenesis promoter or cartilage repair agent, it is even more preferably a lower alkyl group with an optional substituent which is optionally substituted with a halogen atom. Among these, R.sup.2 is yet more preferably a lower alkyl group substituted at the same carbon with two lower alkoxy groups which are optionally substituted with 1-5 halogen atoms or the group --O--Z--O-- wherein Z represents a lower alkylene group optionally substituted with 1-10 halogen atoms, and still more preferably, a group represented by general formula (II): ##STR5## wherein R.sup.10 and R.sup.11 may be the same or different, and each represents a lower alkyl group optionally substituted with 1-5 halogen atoms, preferably either or both being lower alkyl groups with 1-5 halogen atoms, or general formula (III): ##STR6## wherein Z represents a lower alkylene group optionally substituted with 1-10 halogen atoms. R.sup.2 is more preferably a 2,2-diethoxyethyl group, a 2,2-dimethoxyethyl group, a 2,2-diisopropoxyethyl group, a 2,2-bis(2-fluoroethoxy)ethyl group or a 2,2-bis(2-chloroethoxy)ethyl group, among which a 2,2-diethoxyethyl group and a 2,2-bis(2-fluoroethoxy)ethyl group are most preferred, and a 2,2-diethoxyethyl group is particularly preferred. R.sup.3 represents a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent, an aryl group with an optional substituent or a heterocyclic group with an optional substituent. R.sup.3 is preferably a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent or an aryl group with an optional substituent, among which an aryl group with an optional substituent is particularly preferred. Preferred as the substituent is a lower alkyl group (preferably a methyl group and an ethyl group, and especially a methyl group) and an amino group optionally having a lower alkyl group, and a 4-methylphenyl group is especially preferred for R.sup.3. R.sup.4 represents a hydrogen atom, a lower alkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, --OR.sup.5, --SR.sup.5 or --NR.sup.6 R.sup.7. Here, R.sup.5, R.sup.6 and R.sup.7 may be the same or different and each represents a hydrogen atom, a lower alkyl group with an optional substituent, a cycloalkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent, a lower alkoxy group or an amino group with an optional substituent, and R.sup.6 and R.sup.7 may together form a group represented by --(CH.sub.2).sub.m -- or --(CH.sub.2).sub.1 NR.sup.8 (CH.sub.2).sub.k -- wherein k, 1 and m each represent an integer of 1-8 and R.sup.8 represents a hydrogen atom or a lower alkyl group. R.sup.4 is preferably a lower alkyl group with an optional substituent, an aryl group with an optional substituent, a heterocyclic group with an optional substituent or a group represented by --OR.sup.5 or --NR.sup.6 R.sup.7 wherein R.sup.5, R.sup.6 and R.sup.7 are as previously defined, and it is more preferably the group --NR.sup.6 R.sup.7 wherein R.sup.6 and R.sup.7 may be the same or different and each represents a hydrogen atom or an aryl group with an optional substituent. Among these, R.sup.4 is preferably a group represented by --NR.sup.6 R.sup.7 wherein R.sup.6 and R.sup.7 may be the same or different and each represents a hydrogen atom or an aryl group which aryl group has a lower alkyl group or amino group which amino group optionally has a lower alkyl group, and it is most preferably a group represented by --NHR.sup.7 wherein R.sup.7 is a 4-methylphenyl group, a 4-ethylphenyl group or a 4-(N,N-dimethylamino)phenyl group. X and Y may be the same or different and represent --CH.sub.2 --, --NH--or --O--, among which X is preferably --CH.sub.2 --, --NH--or --O-- and Y is preferably --CH.sub.2 --or --NH--. From the viewpoint of activity as a chondrogenesis promoter or cartilage repair agent, X and Y may be the same or different and are preferably --CH.sub.2 --or --NH--, and most preferably, X is --NH--and Y is --CH.sub.2 --. The indolin-2-one derivatives of the invention may be used in the form of pharmaceutically acceptable salts. As examples of such salts there may be mentioned inorganic salts such as hydrochloric acid salts, hydrobromic acid salts, hydroiodic acid salts, sulfuric acid salts and phosphoric acid salts; organic acid salts such as succinic acid salts, malonic acid salts, acetic acid salts, maleic acid salts, fumaric acid salts, citric acid salts, benzoic acid salts and salicylic acid salts; and metal salts such as sodium salts, potassium salts and magnesium salts. The indolin-2-one derivatives of the invention may also be optically active forms. When in optically active forms, the absolute configuration at the 3 position is preferably the R configuration. As examples of specific compounds which are indolin-2-one derivatives according to the invention there may be mentioned the compounds mentioned in the examples of WO94/19322, as well as the compounds mentioned in the examples of Japanese Patent Application Laid-Open No. 7-48349, namely, Compound Nos. 1-201-listed in the following tables (Tables 1-9). In Tables 1-9, R.sup.1 -R.sup.4, X, Y and n have the same definitions as for general formula (I) above. TABLE 1 As examples of specific compounds more suitable as chondrogenesis promoters and cartilage repair agents according to the invention there may be mentioned the following compounds. ##STR167## ##STR168## As examples of biometabolites of compound A there may be mentioned the following compounds, H and I. ##STR169## The compounds listed in Tables 1 to 9 may be synthesized by the process described in Japanese Patent Application Laid-Open No. 7-48349. Of the compounds A to G mentioned above, compounds A to D may be synthesized by the process described in Japanese Patent Application Laid-Open No. 7-48349. Compounds E, F and G may be synthesized, for example, by Reaction Path A shown below (corresponding to "Reaction Path 6" in Japanese Patent Application Laid-Open No. 7-48349), using as the starting material the aldehyde intermediate mentioned in the examples of the present application, synthesized according to the process described in Japanese Patent Application Laid-Open No. 7-48349. (Reaction Path A) ##STR170## Compound H mentioned above may be synthesized according to "Reaction Path 7" in Japanese Patent Application Laid-Open No. 7-48349. Compound I may be obtained by the following process, either in a racemic form or optically active form. Specifically, a racemic form of compound I may be synthesized, for example, according to the following Reaction Path B (corresponding to "Reaction Path 5" in Japanese Patent Application Laid-Open No. 7-48349), using an isocyanate having a hydroxyl group protected with a suitable protecting group (for example, a substituted silyl group such as a triethylsilyl group, a t-butyldimethylsilyl group or the like). An optically active form of compound I may be synthesized, for example, according to the following Reaction Path B (corresponding to "Reaction Path 7" in Japanese Patent Application Laid-Open No. 7-48349), using an isocyanate having a hydroxyl group protected with a suitable protecting group (for example, a substituted silyl group such as a triethylsilyl group, a t-butyldimethylsilyl group or the like), or by optically separating a stereoisomeric mixture of compound I by a method well known to those skilled in the art (for example, a method using an optically active column). Identification, structure determination and purity determination of the obtained compound may be accomplished by ordinary methods (spectroscopic methods such as NMR, IR, etc. and high performance liquid chromatography or the like). (Reaction Path B) ##STR171## A chondrogenesis promoter according to the invention may be applied for a variety of uses without any particular restrictions, so long as the chondrogenesis promoting effect of the agent can be effectively utilized; it is particularly useful for treatment of chondropathy accompanying cartilage dysfunction due to degeneration or destruction of cartilage, treatment of damage or ablation of cartilage due to injury or surgery, or treatment of congenital cartilage hypoplasia or malformation. Examples of such chondropathic conditions include osteoarthritis, chronic rheumatoid arthritis, dissecting osteochondrosis, injury-induced articular cartilage damage, herniated intervertebral disk, and the like. Examples of congenital cartilage hypoplasia or malformation include anotia and microtia. An agent containing a compound according to the invention may be administered either orally or parenterally, but parenteral administration is preferred from the standpoint of avoiding unnecessary promotion of chondrogenesis outside of the site of administration, and from the standpoint of the effect. The agent may be formulated in a manner suitable for the method of administration. A pharmaceutical composition comprising a compound of the invention as an active ingredient may be formulated using an ordinary formulation technique. The pharmaceutical composition may be used in various forms depending on the purpose of use, such as in the form of capsules, granules, cream, powder, syrup, tablets, injection or ointment, and either in solid or liquid form. The carrier or excipient used for the formulation may be a solid or liquid substance. As examples there may be mentioned lactose, magnesium stearate, starch, talc, gelatin, agar, pectin, gum Arabic, olive oil, sesame oil, ethylene glycol and the like, as well as any other commonly used substance. The content of the compound of the invention in the formulation will differ depending on the form of preparation, but usually a concentration of 0.00001-80 wt % is preferred. A pharmaceutical composition of a compound of the invention may administered in various dosages depending on the type of warm-blooded animal, such as a human, the severity of symptoms and the clinical diagnosis; in most cases, however, the daily oral dosage will be 0.01-2 g/kg and the daily parenteral dosage will be 0.0000001-0.01 g/kg. This dosage may be given at once or spread over several times every 1 to 7 days, with appropriate adjustment depending on the severity of symptoms and the clinical diagnosis. As shown in the examples provided below, agents according to the invention were shown to have chondrogenesis promoting effects when administered orally or parenterally to rats. This indicates the usefulness of these agents as therapeutic agents for cartilage disease. Also, the chondrogenesis promoting effects exhibited by compounds represented by general formula (I) on tracheal cartilage, interspinal disks, auricular cartilage and sternal cartilage indicate that the compounds represented by general formula (I) can constitute useful therapies as repair agents for diseases characterized by cartilage defect and degeneration or destruction of cartilage. Moreover, treatment of undifferentiated mesenchymal cells (CL-1) or chondrocytes with chondrogenesis promoters represented by general formula (I) can yield chondrocytes (or more abundant chondrocytes if it is chondrocytes that are treated), as well as the extracellular matrix or cartilage-like tissue. The chondrogenesis promoters described in the present application may be used for analysis of extracellular matrix metabolism of chondrocytes and the mechanism of differentiation into chondrocytes (biological properties), analysis of the components constituting the extracellular matrix (chemical properties), and analysis of properties such as viscoelasticity (physical properties). Furthermore, the indolin-2-one derivatives represented by general formula (I) of the invention that exhibit chondrogenesis promoting effects may, in some cases, promote calcified chondrogenesis by the same chondrogenesis promoting effect, thus eliciting, in such cases, a bone fracture cure promoting effect via eventual endochondral ossification by the compounds. That is, these compounds are also expected to be useful as bone fracture repair promoters. |
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