KR20090064463A - Cathepsin proteases inhibitors - Google Patents

Abstract

The invention provides compounds (I) and pharmaceutical compositions thereof, and more particularly, bicyclic carbamates, which are useful as cathepsin inhibitors, and methods for using such compounds or pharmaceutically acceptable salts thereof, wherein X is 0 or S; R1 is OR2, halo, (CR2)nR3, nitro, cyano, amino, aniido, sulfonamide, or an optionally substituted C1-6alkyl, C2-6alkenyl, or C3-6 alkynyl;R2 is H, (CR2)nR3, or an optionally substituted C1-6alkyl, C2-6 alkenyl or C3-6 alkynyl; R3 is an optionally substituted aryl, heteroaryl, carbocyciic ring or heterocyclic ring; m is 1-3; and n is 0-4.

Description

Cathepsin Protease Inhibitor {CATHEPSIN PROTEASES INHIBITORS}

Cross-Reference to the Related Application

This application claims the benefit of US Provisional Patent Application No. 60 / 862,500, filed October 23, 2006, which is incorporated herein in its entirety.

Most cathepsins are endopeptidase belonging to the papain-like cysteine protease family. Cathepsin is usually present at high concentrations in the lysosome and endosome compartments and plays a role in a wide range of physiological processes. Among its roles are non-specific functions, such as the degradation of both internalized and cellular proteins, and more specific functions in the treatment of enzymes and hormones. Various diseases are associated with overexpression of specific proteases. For example, inhibition of certain cathepsin proteases is believed to be therapeutically relevant in pathological diseases associated with cellular homeostasis, apoptosis, tumor invasion and metastasis, bone uptake and antigen presentation.

Most of the small molecule inhibitors of the protease, including small molecule inhibitors for cathepsin, are peptidomimetic that recognizes specific pockets of enzymes and defines substrate selectivity, and electrophilicity, which makes important contacts for catalytic domains. It consists of a "warhead". The nature of the electrophilic group determines the classification of the small molecules as reversible and irreversible inhibitors. Some electrophilic groups have been commonly used for the inhibition of proteases by covalent interactions of enzymes with inhibitors. For example, aldehydes, semicarbazones, nitriles and ketones usually cause reversible inhibition, while halomethylketone, epoxide and Michael acceptor are some of the functional groups used for irreversible inhibition.

To date, relatively few protease inhibitors have been successfully progressed through clinical trials. Major problems include the non-drug-like nature of the peptide portion of the inhibitor and / or the lack of selectivity due largely to non-specific interactions of electrophilic functional groups with endogenous nucleophiles in vivo. In order to circumvent this problem commonly found in the classical approach of substrate-based drug design, new molecular concepts need to be found.

Description of the invention

The present invention provides compounds and pharmaceutical compositions thereof that may be useful as inhibitors of cathepsin proteases.

In one aspect, the present invention provides a compound of Formula 1 or a pharmaceutically acceptable salt thereof.

Where

X is O or S;

R ¹ is OR ^{2, _{halo, (CH 2) n R 3}} , nitro, cyano, amino, amido, a sulfonamide, or an optionally substituted C _{1 - 6} alkyl, C _{2 -6} alkenyl or C _{3 -6} Alkynyl;

R ² is _{H, (CH 2) n R} 3, or an optionally substituted C _{1 - 6} alkyl, C _{2 -6} alkenyl or C _{3 -6} alkynyl;

R ³ is optionally substituted aryl, heteroaryl, carbocyclic ring or heterocyclic ring;

m is 1 to 3;

n is 0-4.

In one embodiment, m is 1. For example, R ¹ is halo or OR ^2, where R ² may be an optionally substituted phenyl, benzyl or C _{1 -6} alkyl. In some instances, the compound of formula 1 has cis stereoconformation. In another example, the compound of Formula 1 has a trans stereoconfiguration.

In another aspect, the present invention provides a pharmaceutical composition comprising a compound of formula 1 and a pharmaceutically acceptable excipient.

In another aspect, the present invention provides a method for inhibiting cathepsin protease by administering to a system or subject in need of inhibition of cathepsin protease, a therapeutically effective amount of a compound of Formula 1, or a pharmaceutically acceptable salt or pharmaceutical composition thereof. It provides a method of inhibiting cathepsin protease, including.

In addition, the present invention is directed to a system or subject in need of treatment for a disease or condition mediated by cathepsin protease activity, wherein the cathepsin protease- Provided is a method of treating a disease or condition comprising treating the mediated disease or condition. In one embodiment, the present invention is directed to treating a disease or condition mediated by papain-like cathepsin protease, including but not limited to cell homeostasis, apoptosis, tumor invasion and metastasis, bone uptake and antigen presentation. Provide a method. For example, the compounds of the present invention can be used to treat osteoporosis, arthritis, asthma, autoimmune diseases and tumors.

In another aspect, the present invention provides the use of a compound of formula 1 for the manufacture of a medicament for the treatment of a disease mediated by cathepsin protease, more specifically papain-like cathepsin protease.

In the above method using the compound of the present invention, the compound of formula 1 may be administered in a system comprising a cell or tissue. In other embodiments, the compound of formula 1 may be administered to a human or animal subject.

Justice

"Alkyl" refers to a moiety as a structural element of a given moiety and other groups (eg, halo-substituted alkyl and alkoxy) and may be straight or branched. As used herein, the term “optionally substituted alkyl, alkenyl or alkynyl” is optionally halogenated (eg CF ₃ ), or one or more carbon atoms is substituted or replaced by a heteroatom such as NR, O or S (Eg, —OCH ₂ CH ₂ O—, alkylthiol, thioalkoxy, alkylamine, etc.).

"Aryl" refers to a monocyclic or fused bicyclic aromatic ring containing carbon atoms. For example, aryl can be phenyl or naphthyl. "Arylene" means a divalent radical derived from an aryl group.

The term "heteroaryl" as used herein is aryl as defined above wherein at least one of the ring members is a heteroatom. Examples of heteroaryl include pyridyl, indolyl, indazolyl, quinoxalinyl, quinolinyl, benzofuranyl, benzopyranyl, benzothiopyranyl, benzo [1,3] dioxol, imidazolyl, benzo- Imidazolyl, pyrimidinyl, furanyl, oxazolyl, isoxazolyl, triazolyl, tetrazolyl, pyrazolyl, thienyl, and the like.

As used herein, the term “carbocyclic ring” refers to a saturated or partially unsaturated monocyclic, fused bicyclic or bridged polycyclic ring containing, for example, a carbon atom which may be optionally substituted by = O. do. Examples of carbocyclic rings include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclopropylene, cyclohexanone, and the like.

The term "heterocyclic ring" as used herein is a carbocyclic ring as defined above in which one or more ring carbons are replaced with heteroatoms. For example, the heterocyclic ring is N, O, S, -N = , -S-, -S (O), -S (O) 2 - or -NR- (wherein, R is hydrogen, C _{1 - 4} alkyl or protecting group). Examples of heterocyclic rings include morpholino, pyrrolidinyl, pyrrolidinyl-2-one, piperazinyl, piperidinyl, piperidinylone, 1,4-dioxa-8-aza-spiro [4.5] Death-8-day, and the like.

Unless indicated otherwise, substituents are considered “optionally substituted” which means that the substituents are, for example, optionally halogenated alkyl, alkenyl, alkynyl, alkoxy, alkylamine, alkylthio, alkynyl, amide, amino ( Mono- and di-substituted amino groups), aryl, aryloxy, arylthio, carbonyl, carbocyclic, cyano, cycloalkyl, halogen, heteroalkyl, heteroalkenyl, heteroalkynyl, heteroaryl, heterocy Click, hydroxy, isocyanato, isothiocyanato, mercapto, nitro, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N- Amido, S-sulfonamido, N-sulfonamido, C-carboxy, O-carboxy, perhaloalkyl, perfluoroalkyl, silyl, sulfonyl, thiocarbonyl, thiocyanato, trihalo Selected individually and independently from methanesulfonyl, and protected compounds thereof Means that the group which may be substituted by or more group (s). Such protecting groups capable of forming protected compounds of such substituents are known to those skilled in the art and described in Greene and Wuts, Protective Groups in Organic Synthesis, 3 ^rd Ed., John Wiley & Sons, New York, NY, 1999 and It may be found in references such as Kocienski, Protective Groups, Thieme Verlag, New York, NY, 1994, which are incorporated herein in their entirety.

As used herein, the term “co-administration” or “combination administration”, etc., is meant to encompass the administration of selected therapeutic agents to a single patient and does not necessarily require that the agents be administered in the same route or at the same time. It is intended to include).

The term "pharmaceutical combination" as used herein refers to a product obtained from mixing or combining the active ingredients, and includes both fixed and unfixed combinations of the active ingredients. The term "fixed combination" means that the active ingredient, such as the compound of formula 1 and the co-formulation, is administered to the patient simultaneously in a single form or in a single dosage form. The term “unfixed combination” means that the active ingredient, eg, the compound of formula 1 and the co-formulation are administered to the patient simultaneously, jointly, or sequentially as a separate individual without any particular time limit, wherein the administration is Providing a therapeutically effective level of active ingredient in the patient's body. Unfixed combinations also apply to cocktail therapy, eg the administration of three or more active ingredients.

The term "therapeutically effective amount" means the amount of a compound that will cause a biological or medical response to be sought by a researcher, veterinarian, physician or other clinician in a cell, tissue, organ, system, animal or human.

The term “administration” of a compound of interest and / or “administering a subject compound” should be understood to mean providing a compound of the invention, including prodrugs of a compound of the invention, to an individual in need thereof.

The present invention provides compounds and compositions that inhibit cathepsin protease, more specifically papain-like cathepsin protease. In addition, the present invention provides a therapeutically effective amount of a compound of formula 1 to a system or subject in need of treatment for a disease or condition mediated by cathepsin protease activity, particularly a disease or condition mediated by papain-like cathepsin protease, or a pharmaceutical thereof A disease or condition mediated by cathepsin protease activity, in particular by papain-like cathepsin protease, comprising treating the papain-like cathepsin protease-mediated disease or condition by administering a phase tolerable salt or pharmaceutical composition Provided are methods for treating a mediated disease or disorder.

Papain-like cysteine protease has been identified as an important proteolytic activator in degenerative, invasive and immune system related disorders (Lecaille et al., Chem. Rev. 2002, 102: 4459-4488; Broemme et al., Curr. Pharm.Des. 2002, 8: 1639-1658). For example, cathepsin K is the major bone-degenerative activator in osteoclasts, and its selective inhibition may be beneficial for treating osteoporosis and certain forms of arthritis. Cathepsin S plays an important role in the presentation of MHC class II dependent antigens, and inhibition of cathepsin S significantly reduces the response to antigens, thereby cathepsin S is a drug target for asthma and certain autoimmune diseases. (Riese et al., J. Clin. Invest. 1998, 101: 2351-2363). Cathepsins are also involved in tumor invasion and metastasis, and tumors of the central nervous system such as astrocytoma, glioblastoma and meningioma (Berquin et al., Perspect. Drug Discovery Design 1995, 2: 371-388; Levicar et al. , J. Neurooncol. 2002, 58: 21-32).

In one aspect, the compound of the present invention is a compound of formula (I) or a pharmaceutically acceptable salt thereof.

<Formula 1>

Where

X is O or S;

R ¹ is OR ^{2, _{halo, (CH 2) n R 3}} , nitro, cyano, amino, amido, a sulfonamide, or an optionally substituted C _{1 - 6} alkyl, C _{2 -6} alkenyl or C _{3 -6} Alkynyl;

R ² is _{H, (CH 2) n R} 3, or an optionally substituted C _{1 - 6} alkyl, C _{2 -6} alkenyl or C _{3 -6} alkynyl;

R ³ is optionally substituted aryl, heteroaryl, carbocyclic ring or heterocyclic ring;

m is 1 to 3;

n is 0-4.

In one embodiment, the compounds of the present invention can be used to inhibit papain-like cathepsin protease. Table 1 shows the apparent inhibition constants (K _i (app), μM) of various compounds of the present invention for the inhibition of various cathepsins (St _rel is relative stereoconfiguration).

Cyclic carbamates (8) and (12) to (16) exhibited inhibitory activity over a wide range of papain-like cathepsin proteases. In this family of proteases, the apparent K _i value was the lowest for cathepsin B, followed by low for cathepsin S, C, L and K. Moderate activity was shown for cathepsin V, X, F and H. The compound has been shown to be inactive against other families of cysteine proteases (eg caspase) and members of the serine protease family (eg hepcin, thrombin, MT-SP1 or trypsin).

In general, a decrease in K _i value was observed when the trans coordination stereoisomer was compared to the cis coordination stereoisomer (compare compounds (8) and (12)). When compared to phenylsubstituted analog (14), analog (12) containing more hydrophobic dimethylphenyl groups was shown to be consistently higher activity in the cathepsin panel. Thiocarbamate (13) showed a substantially lower inhibitory effect than oxo-carbamate. Alkyl-substituted analogs (15) (where R is benzyl) and (16) (where R is isopropyl) exhibited a K _i value that is generally about 10 times higher than compounds (12) or (14). The low nanomolar activity of the benzyl substituted analog (15) against cathepsin B is exceptional, indicating that it is the most selective (100-fold) inhibitor of cathepsin B in the family.

Although the mechanism is not essential for practicing the present invention, the inhibition mechanism by the bicyclic carbamate of the present invention was investigated in dialysis studies. In sum, cathepsin B was first incubated at 37 ° C. with 0.2 μM (200 × K _i ) of compound (12) for complete inhibition of catalytic activity (2.9 rfu / sec for inhibitor treatment group; 121 rfu for vehicle control) / sec). The mixture was then dialyzed thoroughly at 4 ° C. to remove unbound inhibitors and then remeasured cathepsin activity at 37 ° C. (51 rfu / sec for inhibitor treatment group; 71 rfu / sec for vehicle control). After dialysis, a clear recovery of cathepsin B activity in the compound (12) treated sample suggests that compound (12) is a partially reversible inhibitor. However, careful examination of the progress curve showed that substrate conversion was accelerated during the analysis (indicated by dashed arrows). Since the substrate conversion of the control is not affected by the incubation temperature, this rate increase may be due to the temperature sensitive release of the covalently bound compound (12) from the active site.

Potential adduct formation between cathepsin B and the inhibitor (compound (12)) was analyzed by trypsin / chymotrypsin digestion followed by mass spectrometry. Two active site fragments of the test enzyme (cathepsin B) were identified as ¹⁹ EIRDQGSCGSC ^* W ³⁰ and ²² DQGSCGSC ^* W ³⁰ from untreated cathepsin B samples. For inhibitor treated cathepsin B, both fragments obtained a mass of 247, corresponding to a single covalent bond of compound (12) to the catalytic cysteine residue (C ^* ). The stability of the adduct was sensitive to temperature rise. Compared to the control maintained on ice, 2-hour incubation at 37 ° C. promoted dissociation of the enzyme-inhibitor adduct, resulting in a 4-fold increase in metabolite presence. Multiple reaction monitoring (MRM) properties of the metabolite were identical to that of the dissociation product (11) presented.

In summary, the present invention provides compounds of formula 1 which are selective for the cathepsin family and have subtype selectivity in the order B> S> C, L, K> V, X, F, H. The preference for cis-coordinated hydrophobic R-groups for carbamate oxygen was observed, with apparent Ki values for cathepsin B in the range below 10 nanomolar concentrations. The carbamate functional group in the backbone is substantially unstable and provides a weak point for nucleophilic attack by the active site cysteine thiol. Then, the ring-opening reaction occurs to the bicyclic compound and covalently binds to the catalytic cysteine, resulting in inhibition of the enzyme. The hypothesis is based on mass spectrometry analysis of the degraded enzyme. After incubation with compound (12), the fragment containing the catalytic cysteine has an inhibitor covalently. The non-linear rate recovery and the properties of the metabolite via MRM in dialysis studies suggest that the inhibitor is slowly hydrolyzed at ambient temperature to fall off the enzyme as its ring-opened synthetic precursor (11).

Dosing and Pharmaceutical Compositions

In general, the compounds of the present invention will be administered in a therapeutically effective amount, alone or in combination with one or more therapeutic agents, in any general and acceptable manner known in the art. The therapeutically effective amount can vary widely depending on the severity of the disease, the age and relative health of the subject, the potency of the compound used and other factors. In general, it is shown that satisfactory results are obtained systemically at a daily dosage of about 0.03 to 2.5 mg / kg body weight. The designated daily dosage in larger mammals, such as humans, is conveniently administered in the range of about 0.5 mg to about 100 mg, eg, up to four divided doses or delayed administration per day. Suitable unit dosage forms for oral administration comprise from about 1 to 50 mg of active ingredient.

The compounds of the present invention can be used in any conventional route as pharmaceutical compositions, especially in the intestine, for example orally (eg in the form of tablets or capsules) or parenterally (eg, injectable solutions). In the form of preparations or suspensions), topically (eg in the form of lotions, gels, ointments or creams) or intranasally or suppository forms.

Pharmaceutical compositions comprising a compound of the invention in free form or in a pharmaceutically acceptable salt form together with one or more pharmaceutically acceptable carriers or diluents may be prepared in a conventional manner by mixing, granulating or coating methods. For example, oral compositions may comprise a) diluents such as lactose, dextrose, sucrose, mannitol, sorbitol, cellulose and / or glycine; b) lubricants such as silica, talc, stearic acid, magnesium or calcium salts thereof, and / or polyethylene glycol; For tablets c) with a binder such as magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose and / or polyvinylpyrrolidone; And if necessary d) disintegrants, for example starch, agar, alginic acid or its sodium salt, or effervescent mixtures; And / or e) tablets or gelatin capsules, including with absorbents, colorants, flavors and sweeteners. Injectable compositions can be aqueous isotonic solutions or suspensions, and suppositories can be prepared from fatty emulsions or suspensions.

The composition may be sterile and / or contain adjuvant such as preservatives, stabilizers, wetting agents or emulsifiers, dissolution accelerators, osmotic pressure regulating salts and / or buffers. In addition, they may also contain other ingredients of therapeutic value. Formulations suitable for transdermal application include an effective amount of a compound of the invention in combination with a carrier. The carrier may comprise an absorbable pharmacologically acceptable solvent to assist passage through the skin of the host. For example, transdermal devices may be provided in the back member, optionally in a reservoir containing the compound, optionally in a rate control barrier that allows the compound to be delivered to the skin of the host at a controlled rate over a delayed period of time, and to the skin. In the form of a bandage comprising means for securing the device. Matrix transdermal formulations may also be used. For example, formulations suitable for topical application to the skin and eyes may be aqueous solutions, ointments, creams or gels known in the art. It may contain solubilizers, stabilizers, tonicity enhancing agents, buffers and preservatives.

Compounds of the invention can be administered in therapeutically effective amounts with one or more therapeutic agents (pharmaceutical combinations). When the compound of the present invention is administered in combination with other therapeutic agents, the dosage of the co-administered compound will of course vary depending on the type of co-drug used, the particular drug used, the disease to be treated and the like.

For example, the compounds of the present invention can be used in combination with chemotherapeutic agents to treat cell proliferative disorders and tumors. Examples of chemotherapeutic agents that can be used in the compositions and methods of the present invention include anthracyclines, alkylating agents (eg mitomycin C), alkyl sulfonates, aziridine, ethyleneimine, methylmelamine, nitrogen mustard, nitrosourea, Antibiotics, anti-metabolites, folic acid analogs (eg, dihydrofolate reductase inhibitors such as methotrexate), purine analogs, pyrimidine analogs, enzymes, grapephytotoxins, platinum-containing agents, interferons and interleukins, It is not limited to this. Specific examples of known chemotherapeutic agents that can be used in the compositions and methods of the present invention include, but are not limited to, busulfan, improsulfan, pifosulfan, benzodepa, carbocuone, meturedepa, uredepa, altretamine, triethylene Melamine, triethylenephosphoramide, triethylenethiophosphoramide, trimethylolomelamine, chlorambucil, chlornaphazine, cyclophosphamide, esturamustine, ifosfamide, mechlorethamine, mechlorethamine Oxide Hydrochloride, Melphalan, Norshamvikin, Fennesterine, Prednisostin, Trophosphamide, Urasyl Mustard, Carmustine, Chlorozotocin, Potemustine, Romustine, Nimustine, Lanimustine, Dacavazin, Mannomustine, mitobronitol, mitolactol, fibrobromine, alaccinomycin, actinomycin F (1), anthracycin, azaserine, bleomycin, cocktinomycin, carrubicin, karji Filin, chromomycin, dactinomycin, daunorubicin, daunomycin, 6-diazo-5-oxo-1-norleucine, doxorubicin, epirubicin, mitomycin C, mycophenolic acid, nogalamycin, Olibomycin, peplomycin, plicamycin, porphyromycin, puromycin, streptomycin, streptozosin, tubercidin, ubenimex, ginostatin, zorubicin, denophtherin, methotrexate, pterotropin, Trimetrexate, fludarabine, 6-mercaptopurine, thiamiprine, thioguanine, ancitabine, azacytidine, 6-azauridine, carmopur, cytarabine, dideoxyuridine, doxyfluri Dean, Enositabine, Phloxuridine, Fluorouracil, Tegapur, L-Asparaginase, Fulmoszyme, Aceglaton, Aldophosphamide Glycoside, Aminolevulinic Acid, Amsacrine, Vesturabsil , Bisantrene, carboplatin, cisplatin, depopamide, Demecolsin, diajikuon, elponnitine, elliptinium acetate, etogluside, etoposide, flutamide, gallium nitrate, hydroxyurea, interferon-alpha, interferon-beta, interferon-gamma, interleukin-2, Lentinane, ronidamine, mitoguazone, mitoxantrone, furdamol, nitracrine, pentostatin, phenamect, pyrarubicin, grapephylic acid, 2-ethylhydrazide, procarbazine, lazoic acid, sizopyran, Spirogermanium, paclitaxel, tamoxifen, teniposide, tenuazone acid, triazcuone, 2,2 ', 2 "-trichlorotriethylamine, urethane, vinblastine, vincristine and vindesine, including but not limited to Do not.

In addition, when combined with other immunomodulatory or anti-inflammatory substances, for example cyclosporin, rapamycin or ascomycin, or immunosuppressive analogs thereof, such as cyclosporin A (CsA), cyclosporin G, FK-506, Rapamycin, or a comparable compound, corticosteroid, cyclophosphamide, azathioprine, methotrexate, brequinar, leflunomide, mizoribin, mycophenolic acid, mycophenolate mofetil, 15-deoxys Such as ferguline, immunosuppressive antibodies, in particular leukocyte receptors such as MHC, CD2, CD3, CD4, CD7, CD25, CD28, B7, CD45, CD58 or their ligands, or CTLA4Ig Synergistic effects may occur when used in combination with other immunomodulatory compounds.

The invention also provides a pharmaceutical combination, eg a kit, comprising a) a first agent which is a compound of the invention in free form or in a pharmaceutically acceptable salt form described herein, and b) at least one co-formulation. do. The kit may comprise instructions for its administration.

Process for the preparation of the compound of the present invention

Compounds of the present invention can be prepared according to Scheme 1 below.

In Scheme 1, the reagents and conditions are as follows: (a) CbzCl, 0.5 M Na ₂ CO ₃ / dioxane 5: 2, room temperature, 5h; (b) mCPBA, DCM, 0 ° C.-room temperature, 7 h; (c) ROH, 2 M NaOH / MeCN 1: 4, reflux, 48 h; (d) 1 atm H ₂ , Pd / C (cat), EtOH, room temperature, 3 h; (e) triphosgene, NEt ₃ , DCM, 0 ° C., 1 h; (f) NaH, room temperature, 18 h; (g) PPh ₃ , DEAD, p -nitrobenzoic acid, THF, 50 ° C., 48 h; (h) NaOH, MeOH, room temperature, 1 h; (i) 1 atm H ₂ , Pd / C (cat), EtOH, room temperature, 3 h; (j) OsO ₄ , NMO, citric acid, tBuOH / H ₂ O 1: 1, 50 ° C., 12 h; (k) HBr, HOAc, 0 ° C.-room temperature, 3 h; (l) 2,3-dimethylphenol, K ₂ CO ₃ , MeCN, 60 ° C., 12 h; (m) MeNH ₂ , MeOH, 50 ° C., 2 h; (n) CDI, DMAP, benzene, reflux, 4 h. Compounds (3) to (21) were racemic mixtures.

According to the synthetic procedure described by Paioni (US Pat. No. 4,160,837; incorporated herein in its entirety), tetrahydropyridine (2) was converted into a racemic Cbz-protected epoxide (3) in two steps. Switched. A mixture of regioisomers was obtained by ring opening of epoxide (3) (ROH is 2,3-dimethylphenol) under basic aqueous conditions. As expected for the predominant trans-biaxial ring opening, the ratio of 4-aryloxy piperidine (4) to 3-aryloxypiperidine (5) was about 5: 1. The regioisomer was chromatographed and deprotected by hydrogenolysis to yield compounds (6) and (7) in open form. Cyclization of compound (6) requires a two step procedure. First, piperidine-nitrogen is selectively carbonylated with triphosgene and triethylamine to give the corresponding chloroformate, followed by deprotonation of 3-hydroxy groups with sodium hydride to spontaneous intramolecular By causing cyclization, the racemate trans-4- (2,3-dimethyl-phenoxy) -6-oxa-1-aza-bicyclo [3.2.1] octan-7-one (8) was obtained. Attempts to cyclize the chloroformate intermediate of compound (7) to bicyclic compound (9) have been unsuccessful, presumably because the piperidine ring, which requires intramolecular closure, is in the form of an unfavorable boat. Seems to have done.

The 3-position stereocenter of compound (4) was reversed using a Mitsunobu reaction using p-nitrobenzoic acid as the nucleophile. Alternatively, the reverse could be achieved by oxidation / reduction chain reaction using Pyr.SO ₃ and Red-Al. Saponification of intermediate (10) in methanolic base followed by deprotection yielded piperidine (11). Cis-4- (2,3-dimethyl-phenoxy) -6-oxa-1-aza-bicyclo [3.2.1] octan-7-one (12) was prepared by intramolecular cyclization similar to the procedure described above. Obtained.

Both stereoisomeric compounds (8) and (12) were isolated as stable white solids. In addition, the compounds are stable in basic aqueous solutions or neutral aqueous solutions, but spontaneous ring-opening reactions were observed under significant acidic conditions (pH 2 or below). Analogs (13) to (16) (Table 1 above) were synthesized in a similar manner to the synthesis of compound (12). For analog (13), thiophosgen was used instead of phosgene in the cyclization step, and for analogs (15) and (16), the solvent in the ring opening of the epoxide (3) was the Benzyl alcohol, isopropanol in the case of analogue (16) together with NaH. Synthetic alternatives via dihydroxylation result in protected tetrahydropyridine (17). Since no selective cyclization and alkylation of the deprotected diol occurred, the desired bicyclic carbamate analogs were obtained. The epoxide (3) was optionally treated with HBr to give bromide (18) which can be cyclized to carbamate (19), but no attempt to replace bromide was successful. Control compound (21) was synthesized in three steps starting from epibromohydrin (20).

In addition, the compounds of the present invention can be obtained in free form, in their salt form (when salt formers are present) or in ester form (when ester formers are present). Compounds of the invention having acidic groups are advantageously converted to salts by pharmaceutically acceptable bases (eg, aqueous alkali metal hydroxides) in the presence of an ethereal solvent or an alcoholic solvent (such as lower alkanols). Can be. The resulting salt can be converted to the free compound, for example by treatment with an acid. In addition, these salts or other salts can be used for the purification of the compounds obtained. Ammonium salts can be obtained by reaction with a suitable amine, for example diethylamine and the like.

In certain aspects, the compounds of the invention having basic groups can be converted to acid addition salts, in particular pharmaceutically acceptable salts. These include, for example, inorganic acids (eg sulfuric acid, phosphoric acid or hydrofluoric acid); Organic carboxylic acids (eg, C ₁ -C ₄ alkyl carboxylic acids which may be unsubstituted or substituted by halogen, such as acetic acid; saturated or unsaturated dicarboxylic acids such as oxalic acid, succinic acid, maleic acid or fumaric acid; Hydroxycarboxylic acids such as glycolic acid, lactic acid, malic acid, tartaric acid or citric acid); Amino acids (eg, aspartic acid or glutamic acid); Organic sulfonic acids (eg, C ₁ -C ₄ alkylsulfonic acids such as methanesulfonic acid); Or arylsulfonic acids which may be unsubstituted or substituted (eg by halogen). Salts formed by hydrochloric acid, methanesulfonic acid and maleic acid may be preferred.

In view of the close relationship between the free compounds and compounds in their salt or ester form, when a compound is mentioned in the context, it also means the corresponding salt or ester, if possible or appropriate under the given circumstances. In addition, the compounds (including their salts) may be obtained in the form of their hydrates, or may include other solvents used for their crystallization.

In addition, compounds of the present invention comprising free hydroxyl groups may exist in the form of pharmaceutically acceptable physiologically cleavable esters, and therefore they may be included within the scope of the present invention. Such pharmaceutically acceptable esters may be prodrug ester derivatives which are convertible to the corresponding compounds of the invention comprising free hydroxyl groups, preferably by solvolysis or cleavage under physiological conditions. Pharmaceutically acceptable suitable prodrug esters may be those derived from carboxylic acids, carbonate monoesters or carbamic acids, preferably esters derived from optionally substituted lower alkanoic acids or arylcarboxylic acids.

Certain compounds of the present invention may comprise asymmetric carbon atoms (optical centers) or double bonds, meaning that the racemates, diastereomers, enantiomers, geometric isomers and individual isomers are all within the scope of the present invention. It will be apparent to one skilled in the art.

Detailed examples of the synthesis of compounds of formula 1 can be found in the following examples, which are intended to illustrate the invention and are not intended to be limiting.

Example  One

Trans-4- (2,3-dimethyl- Phenoxy ) -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7-on (8)

3,6- Dehydro -2H-pyridine-1- Carboxylic acid  Benzyl ester (2)

1,2,3,6-tetrahydropyridine (1.1 mL, 12 mmol) was dissolved in dioxane (10 mL). An aqueous solution of 0.5 M sodium carbonate (25 mL, 12.5 mmol) was added followed by benzyl chloroformate (1.8 mL, 12 mmol). The mixture was stirred at rt for 3 h and then diluted with ethyl acetate (50 mL). The organic layer was separated, washed with water and dried over magnesium sulfate. The solvent was removed in vacuo to give 3,6-dihydro-2H-pyridine-1-carboxylic acid benzyl ester as a colorless liquid.

7-oxa-3- Keep it up - Bicyclo [4.1.0] heptane -3- Carboxylic acid  Benzyl ester (3)

3,6-dihydro-2H-pyridine-1-carboxylic acid benzyl ester (2) (2.6 g, 12 mmol) was dissolved in dichloromethane (100 mL). M-chloroperoxide acid (4.5 g, 18 mmol) dissolved in dichloromethane (40 mL) was added dropwise over 20 minutes. The mixture was stirred at rt for 7 h, then the solution was washed three times with 1 M aqueous sodium carbonate and once with brine. The organic phase is separated, dried (MgSO ₄ ) and concentrated in vacuo to give the racemate epoxide 7-oxa-3-aza-bicyclo [4.1.0] heptan-3-carboxylic acid benzyl ester (3) Obtained as a colorless oil.

Trans-4- (2,3-dimethyl- Phenoxy ) -3-hydroxy-piperidine-1- Carboxylic acid  Benzyl ester (4) and trans-3- (2,3-dimethyl- Phenoxy ) -4-hydroxy-piperidine-1- Carboxylic acid  Benzyl ester (5)

Dissolve 2,3-dimethylphenol and 7-oxa-3-aza-bicyclo [4.1.0] heptane-3-carboxylic acid benzyl ester (3) (1.0 g, 4.3 mmol) in acetonitrile (20 mL) I was. 2 N aqueous sodium hydroxide (5 mL) was added and the mixture was heated to reflux for 48 h. Acetonitrile was removed under vacuum, and the aqueous residue was diluted with water (20 mL) and extracted twice with dichloromethane (50 mL). The combined organic layers were washed with water and brine, dried over MgSO ₄ and concentrated. The crude product was separated by flash chromatography (hexane / ethyl acetate gradient) to give trans-4- (2,3-dimethyl-phenoxy) -3-hydroxy-piperidine-1-carboxylic acid benzyl ester (4 ) And regioisomer trans-3- (2,3-dimethyl-phenoxy) -4-hydroxy-piperidine-1-carboxylic acid benzyl ester (5) were obtained as a white solid.

Trans-4- (2,3-dimethyl- Phenoxy ) -Piperidine-3-ol (6)

Trans-4- (2,3-dimethyl-phenoxy) -3-hydroxy-piperidine-1-carboxylic acid benzyl ester (4) (700 mg, 2.0 mmol) was dissolved in EtOH (10 mL) and , Catalytic amount of palladium (10% on charcoal) was added. After stirring for 3 hours at room temperature under 1 atm hydrogen, the mixture was filtered through CELITE® and washed with EtOH. The filtrate was concentrated in vacuo to give piperidinol (6) as a white solid.

Trans-4- (2,3-dimethyl- Phenoxy ) -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7-on (8)

Piperidinol (6) (50 mg, 0.23 mmol) was dissolved in DCM (10 mL). Triphosgen (45 mg, 0.15 mmol) was added and the mixture was cooled to 0 ° C. Triethylamine (96 mL, 0.69 mmol) was then added slowly in portions over 50 minutes. Sodium hydride (30 mg, 0.75 mmol) was added and the mixture was warmed to rt and stirred at rt for 18 h. The mixture was washed with 0.25 M phosphate buffer, pH 6.2, the organic layer was separated, dried (MgSO ₄ ) and concentrated. The crude product was purified on reverse phase HPLC (MeCN / H ₂ O gradient, no acidic additives) to afford compound (8) as a white solid.

Example  2

Trans-3- (2,3-dimethyl- Phenoxy ) -Piperidin-4-ol (7)

Substitute the reagent (4) as (5) and transfer the trans-3- (2,3-dimethyl-phenoxy) -piperidine-4- following the procedure for piperidinol (6) of Example 1 Ol (7) was obtained as a white solid.

Example  3

Sheath -4- (2,3-dimethyl- Phenoxy ) -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7-on (12)

Benzyl carbamate (4) (0.93 g, 2.6 mmol) was dissolved in THF (50 mL) together with triphenylphosphine (2.07 g, 7.9 mmol) and p-nitrobenzoic acid (1.32 g, 7.9 mmol). The mixture was cooled to 0 ° C. and then DEAD (1.24 mL, 7.9 mmol) was added slowly. The mixture was stirred at 50 ° C. for 48 h, concentrated and purified by flash chromatography (hexane / ethyl acetate gradient) to give cis-4- (2,3-dimethyl-phenoxy) -3- (4-nitro- Benzoyloxy) -piperidine-1-carboxylic acid benzyl ester was obtained as a white solid.

Sheath -4- (2,3-dimethyl- Phenoxy ) -3-hydroxy-piperidine-1- Carboxylic acid  benzyl Este Porn (10)

NaOH powder (0.4 g, 10 mmol) was dissolved in MeOH (20 mL). Cis-4- (2,3-dimethyl-phenoxy) -3- (4-nitro-benzoyloxy) -piperidine-1-carboxylic acid benzyl ester (0.5 g, 1 mmol) is added and the mixture is Stir at room temperature for 1 hour. The mixture was concentrated, DCM was added and the organic layer was washed twice with water. The organic layer was dried over MgSO ₄ and concentrated to give cis-4- (2,3-dimethyl-phenoxy) -3-hydroxy-piperidine-1-carboxylic acid benzyl ester as a white solid.

Sheath -4- (2,3-dimethyl- Phenoxy ) -Piperidine-3-ol (11)

Cis-4- (2,3-dimethyl-phenoxy) -3-hydroxy-piperidine-1-carboxylic acid benzyl ester (300 mg, 0.85 mmol) is dissolved in EtOH (20 mL) and a catalytic amount of Palladium (10% on charcoal) was added. After stirring for 3 hours at room temperature under 1 atm hydrogen, the mixture was filtered through Celite (R) and washed with EtOH. The filtrate was concentrated in vacuo to afford cis-4- (2,3-dimethyl-phenoxy) -piperidin-3-ol (11) as a white solid.

Sheath -4- (2,3-dimethyl- Phenoxy ) -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7-on (12)

Cis-4- (2,3-dimethyl-phenoxy) -piperidin-3-ol (11) (50 mg, 0.23 mmol) was dissolved in DCM (10 mL). Triphosgen (45 mg, 0.15 mmol) was added and the mixture was cooled to 0 ° C. Triethylamine (96 μL, 0.69 mmol) was then partitioned and added slowly over 50 minutes. After warming to room temperature, sodium hydride (30 mg, 0.75 mmol) was added and the mixture was stirred at room temperature overnight. The mixture was diluted with DCM, washed with water, the organic layer was separated, dried (MgSO ₄ ) and concentrated. The crude product was purified on reverse phase HPLC (MeCN / H ₂ O gradient, no acidic additives) to afford compound (12) as a white solid.

Example  4

Sheath -4- (2,3-dimethyl- Phenoxy ) -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7- Tion  (13)

Triphosphene was replaced with thiophosgen to give the title compound (13) as a white solid following the procedure for bicyclic carbamate (12).

Example  5

Sheath -4- Phenoxy -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7-on (14)

The title compound (14) was prepared as a white solid following the procedure for bicyclic carbamate (12) by replacing 2,3-dimethylphenol with phenol.

Example  6

Sheath -4- Benzyloxy -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7-on (15)

Epoxide (3) (1.0 g, 4.3 mmol) was dissolved in benzyl alcohol (5 mL). Sodium hydride (0.86 g, 21.5 mmol) was added and the mixture was stirred at 50 ° C. overnight. The solvent was then removed in vacuo and the residue was purified by flash chromatography (hexane / ethyl acetate gradient) to afford 3-hydroxy-4-isobutoxy-piperidine-1-carboxylic acid benzyl ester. . Following the procedure for bicyclic carbamate (12), the title compound (15) was prepared as a white solid.

Example  7

Sheath -4- Isobutoxy -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7-on (16)

The title compound (16) was prepared as a white solid following the procedure for bicyclic carbamate (15), replacing benzyl alcohol with isobutanol.

Example  8

Trans-4- Bromo -6-oxa-1- Keep it up - Bicyclo [3.2.1] octane -7-on (19)

7-oxa-3-aza-bicyclo [4.1.0] heptan-3-carboxylic acid benzyl ester (3) (1.0 g, 4.3 mmol) was dissolved in DCM (32 mL). The solution was cooled to 0 ° C., then hydrobromic acid (30% in acetic acid, 8 mL, 30 mmol) was added dropwise and the mixture was stirred at rt for 3 h. The product was precipitated and filtered to afford 4-bromo-piperidin-3-ol hydrobromide (18) as a white solid.

4-Bromo-piperidin-3-ol hydrobromide (18) (100 mg, 0.4 mmol) and triphosgene (80 mg, 0.27 mmol) were suspended in DCM (20 mL) and cooled to 0 ° C. Triethylamine (332 μL, 2.4 mmol) was then added in portions over 60 minutes. After warming to room temperature, the mixture was stirred at room temperature overnight. Purification by flash chromatography (hexane / ethyl acetate gradient) afforded compound (19) as a white solid.

Example  9

5- (2,3-dimethyl- Phenoxymethyl ) -3- methyl - Oxazolidine 2-one (21)

Epibromohydrin (20) (0.7 mL, 8.2 mmol), 2,3-dimethylphenol (1.0 g, 8.2 mmol) and K ₂ CO ₃ (1.13 g, 8.2 mmol) were suspended in MeCN (25 mL), Stir overnight at 60 ° C. The mixture was concentrated and the residue was diluted with DCM and washed twice with water. The organic layer was dried, concentrated and purified by flash chromatography (hexane / ethyl acetate gradient) to give 2- (2,3-dimethyl-phenoxymethyl) -oxirane as a colorless oil.

1- (2,3-dimethyl- Phenoxy ) -3- Methylamino -Propan-2-ol

2- (2,3-dimethyl-phenoxymethyl) -oxirane (0.3 g, 1.7 mmol) was dissolved in MeOH (2 mL). Methylamine (40% in H ₂ O, 0.3 mL, 3.4 mmol) was added and the mixture was stirred at 50 ° C. for 2 h. The solvent is removed in vacuo and the residue is purified on reverse phase HPLC (MeCN / H ₂ O gradient) to convert 1- (2,3-dimethyl-phenoxy) -3-methylamino-propan-2-ol into colorless oil. Obtained.

5- (2,3-dimethyl- Phenoxymethyl ) -3- methyl - Oxazolidine 2-one (21)

1- (2,3-dimethyl-phenoxy) -3-methylamino-propan-2-ol (180 mg, 0.86 mmol) was dissolved in benzene (10 mL). Carbonyl diimidazole (167 mg, 1.03 mmol) was added followed by a catalytic amount of DMAP. The mixture was heated to reflux for 4 hours. The solvent was removed in vacuo and the residue was purified on reverse phase HPLC (MeCN / H ₂ O gradient) to afford compound (21) as a colorless oil.

Example  10

Cathepsin  B Decomposition  Mass spectrometry analysis

See Ricklider et al., Anal. Chem. 74: 3076-3083 (2002), using a LCQ Deca XP Plus mass spectrometer equipped with a home-built nanospray source for online desalination. LC-MS analysis. Peptide digests were loaded onto a 100 μm pretreatment column packed with 2 cm Monitor 5 μm C18 (Column Engineering, Ontario, Canada) and 5 μL per minute with 0.1 M HOAc. Desalted for minutes. After desalting, the pretreatment column is placed in line with a 75 μm inner diameter pulled tip (opening 5 μm) packed with the same packing material 8 cm and the ACN concentration is increased from 0% to 50% over 90 minutes. I was. The column was then washed with 95% ACN and then returned to 0%. The effluent from the HPLC pump was manually split prior to the pretreatment column to achieve a flow rate of 250 nL / min. MS-MS was acquired in three MS-MS scans of the three strongest precursor ions followed by a data-dependent scanning scheme by one full scan. Dynamic exclusion of preselected precursors was set to 1 minute. Tandem MS data were analyzed with TurboSequest (Thermo Electron). A customized database containing 8 proteins was searched using the following parameters: variable methionine oxidation, carboxamidomethyl adducts on cysteine, and carbamate adducts on cysteine (+247) or histidine (+247).

Example  11

Inhibitor-treated Cathepsin  From B LC - MS Of MS Peptide identified by

Cathepsin B was digested with trypsin and chymotrypsin. Peptides were identified by nano-LC-MS / MS as described in the method section. "^" Represents carboxymethylated cysteine and "#" represents modification by modified carbamate compound. When controlling the amount of injected material, the unmodified peptide was present in 118 times less than the modified peptide. As measured by Q-Tof analysis of the original protein, the enzyme precursors were further cleaved upon self-activation to yield active enzymes with alternative N-terminal sequence starting points. Cleavage at the -4 position for the expected major cleavage product is shown in Table 2 below.

Example  12

Bicyclic Carbamate  Effect of temperature on adduct

A sample of cathepsin B was inhibited with compound (12) and then dialyzed overnight to remove any non-cathepsin binding compound. The dialyzed sample was then analyzed by LC-MS for the presence of dissociation product (11). Prior to LC-MS analysis, samples were incubated on ice or at 37 ° C. for 2 hours. The effect of temperature on carbamate inhibited cathepsin B samples was assessed using LC-MS in combination with multiple reaction monitoring (MRM) on Applied Biosystems / MDS SCIEX 4000 Q TRAP. MRM transfer to the open form of carbamate was measured by injection of synthetic standards. Collision energy and exit cell potential were individually optimized for each transition selected. Cathepsin B samples were loaded onto a Phenomenex Luna C5 column (30 × 2 mm) (2% ACN, 0.1M HOAc) and using a linear gradient to 70% ACN, 0.1M HOAc for 5 minutes. Eluted at a flow rate of 300 μL / min. Eluate from the column was introduced into 4000 QTRAP using a TurboV ion source. Source parameters are as follows: Cur, 10; IS, 4500; TEM, 450; GS1, 30; GS2, 15.

Cathepsin B was inhibited with compound (12) and dialyzed overnight. Compound (11) was detected after dialysis (panel A) and incubated for 2 hours at ice (panel B) or 37 ° C. (panel C), followed by detection using LC-MS in combination with MRM. MRM transfer was selected from dissociation activated by collision of synthetic standard compound (11) (influx). Five MRM transitions were monitored for detection of compound (11) (222.1 to 100.0, 71.1, 69.0, 81.9 and 55.0) and only the strongest transitions were graphically shown for clarity (222.1 to 100.0).

Example  13

Deformed Carbamate - Cathepsin  Effect of temperature on stability of B adduct

The presence of metabolite of compound (12) was monitored by MRM. Synthetic analogs of the metabolite (Compound 11) were used to select and optimize MRM transitions. After incubation at 37 ° C. for 2 hours, the amount of compound 11 increased 4-fold.

The examples and embodiments described herein are for illustrative purposes only and various modifications and alterations in this respect will be suggested to those skilled in the art, and such variations and modifications are intended to cover the spirit and scope of the present application and the scope of the appended claims. It should be understood that it is included within. All documents, patents, and patent applications cited herein are hereby incorporated for all purposes.

Claims (10)

A compound of formula 1 or a pharmaceutically acceptable salt thereof. <Formula 1>

Where X is O or S; R ¹ is OR ^{2, _{halo, (CH 2) n R 3}} , nitro, cyano, amino, amido, a sulfonamide, or an optionally substituted C _{1 - 6} alkyl, C _{2 -6} alkenyl or C _{3 -6} Alkynyl; R ² is _{H, (CH 2) n R} 3, or an optionally substituted C _{1 - 6} alkyl, C _{2 -6} alkenyl or C _{3 -6} alkynyl; R ³ is optionally substituted aryl, heteroaryl, carbocyclic ring or heterocyclic ring; m is 1 to 3; n is 0-4. The compound of claim 1, wherein m is 1. The compound of claim 2, wherein R ¹ is halo. The compound of claim 2, wherein R ¹ is OR ² ; R ² is an optionally substituted C _{1 - 6} alkyl, C _{2 -6} alkenyl or C _{3 -6} alkynyl; Or an optionally substituted aryl, heteroaryl, carbocyclic ring or heterocyclic ring. The method of claim 4, wherein, R ² is an optionally substituted phenyl, benzyl or C _{1 -6} alkyl. The compound of any one of claims 1-5, having cis or trans stereoconformation. A pharmaceutical composition comprising a therapeutically effective amount of a compound of any one of claims 1 to 6 and a pharmaceutically acceptable carrier. The preparation of a medicament for the treatment of a disease or disorder mediated by cathepsin protease activity, which is cell homeostasis, apoptosis, tumor invasion and metastasis, bone uptake or antigen presentation. Use of a compound of claim 1, or a pharmaceutically acceptable salt or pharmaceutical composition thereof. Use according to claim 8, wherein the cathepsin protease is papain-like cathepsin protease. Use of a compound of any one of claims 1 to 6, or a pharmaceutically acceptable salt or pharmaceutical composition thereof, in the manufacture of a medicament for inhibiting cathepsin protease activity.