MXPA97001832A - New profarmacos for inflamator tumors and disease therapy - Google Patents

Abstract

The present invention relates to compounds of the formula (I) Glycosyl-Y [-C (= Y) -X-] pW (R) nZC (= Y) - active substance (I) which are suitable for the treatment of diseases cancerous, autoimmune diseases and chronic inflammatory diseases such as rheumatoid arthritis

Description

New prodrugs for the therapy of tumors and inflammatory diseases The effect of drugs (in the Anglo-Saxon field, drugs) is very often that, in the case of certain diseases, enzymes, cytokines or other factors are inhibited, which have been overexpressed pathologically, in terms of their activity promoting the disease. However, the inhibitory effect of the drugs extends not only to the pharmacological target structures (enzymes, cytokines, factors) in diseased tissue, but also inhibits the activities that occur in healthy tissues. This results in unwanted side effects or side effects, which have been observed in the case of many drugs. In order to mitigate the side effects of the drugs, experimental systems were developed, which allow a more selective release of drugs in diseased tissue. Next, such systems are briefly described. The ADEPT system (Antibody Directed Enzyme Prodrug Therapy = therapy with a prodrug and an enzyme directed at antibodies, Bagshawe 1987, Br. J. Cancer 56: 531-532) is a two-stage system, in which, in In the first stage, a conjugate of an antibody and an enzyme (CAE) is injected intravenously (iv). CAE is retained in the tumor because of its selectivity for said tumor, but within 2-7 days it is separated from healthy tissues. The prodrug injected i. v. in the second stage (a non-toxic precursor of the drug) is activated in the tumor by the enzymatic activity of CAE to form the toxic drug. As a result of this activation of the prodrug, specific for the tumor, increased concentrations (in 5-50 times) of the drug in the tumor and lower concentrations of the drug in the healthy tissue are observed, in comparison with the classical therapy. This results in better compatibility as well as superior therapeutic effects in xenograft models in human tumors. { Sharma, S. K. et al. 1991, Disease Markers 9: 225-231).

In a similar way to the ADEPT system, the FMPA concept (from the English Fusion protein Mediated Prodrug Activation = activation by a prodrug, mediated by a fusion protein) works, in which, instead of the CAE, which is xenogenic and as a consequence of it immunogen, a non-immunogenic human fusion protein is used for the activation of a prodrug, selective for a tumor (Bosslet et al., 1994, Cancer Res. 54: 2151-2159). Also in the VDEPT system (from the English Vector Dependent Enzyme Prodrug Therapy = therapy with a prodrug and an enzyme, dependent on a vector, Trinh et al., Cancer Res. 55: 4808-4812), which is a gene therapeutic approach of two. In step, pro-drugs are activated selectively for a tumor after injection of a vector and expression of a structural gene, which encodes an enzyme. An endogenous activation of prodrugs (glucuronil-spacer-anthracycline, Jacquessy et al., 1991, PCT patent document WO 92/19639) in necrotic tumors and inflammatory processes, linked with intense anti-tumor and anti-inflammatory pharmacological effects, was described by Bosslet et al. . , 1994, Cancer Res. 54: 2151-2159 and 1995, Tumor Targeting 1. 45-50, for the first time as PMT (from English Prodrug MonoTheraphie = monotherapy with prodrug). In the pharmacological elaboration of the PMT system it was shown that both the chemistry of the spacer that is spontaneously removed as well as the hydrophilicity and the molar cytotoxicity of the drug component in the prodrug are of decisive importance for efficiency in vivo. A further increase in the efficiency of PMT was observed in combination with substances, which induce necrosis (European patent document EP 0696456 A2). Especially the use of conjugates with antibodies, which have specificity for the VEGF complex and the receptor VEGF, covalently linked with coagulatory proteins, such as for example shortened tissue factor, shows an especially good activity in pharmacological in vivo models, combined with appropriate prodrugs.

Surprisingly, it has been made possible for the authors of the present invention to synthesize prodrugs, which in vi ve, after a corresponding endogenous enzymatic activation, are still essentially more effective than the prodrugs described in EP 0511917 A1 and EP 0595133 A2 . This superior activity is conditioned, on the one hand, by the new and advantageous chemistry of the spacer, but, on the other hand, also by the high molar cytotoxicity of the drug component used. The new advantageous chemistry of the spacer is distinguished by the fact that, especially in the case of active substances, which are bound to the spacer via a hydroxyl group, the active substance is released after an enzymatic dissociation of the glycosyl radical by cyclization and separation of the spacer. This achieves an improved stability of the prodrugs, simultaneously with a good enzymatic dissociability. The prodrugs according to the invention are, moreover, more stable under physiological conditions than the known prodrugs, since they do not release the active substance so rapidly. The invention concerns prodrugs of the formula I Glycosyl-Y [-C (= Y) -X-] p-W (R) n-Z-C (= Y) - active substance (I) and / or physiologically compatible salts of the compound of the formula I, in which glycosyl represents an enzymatically dissociated poly-, oligo- or mono-saccharide, W represents 1) an aromatic radical of 5 to 14 members, 2) naphthyl, 3) indenyl, 4) anthryl, 5) phenanthryl, 6) a heterocyclic radical of 5 to 14 members with 1, 2, 3 or 4 heteroatoms taken from the group consisting of oxygen, nitrogen and sulfur, 7) alkyl (Ci-Cg) , 8) (C2-C6) alkenyl, 9) (C3-C6) cycloalkyl or 10) phenyl, represents 1) a hydrogen atom, 2) (C1-C4) alkyl, 3) phenyl, 4) methoxy, 5) carboxy, 6) methyloxycarbonyl, 7) -CN, 8) -OH, 9) -N02, 10) halogen, such as fluoro, chloro or bromo, 11) sulfonyl, 12) sulfonamido or 13) sulfon-alkyl (C ^ Cj) ) - amido, p represents zero or 1,. • n represents zero, l, 2 or 3, X represents 1) an oxygen atom, 2) -NH-, 3) methyleneoxy, 4) methyleneamino, 5) methylene-alkyl (C ^ C ^ -amino, 6) alkyl (C1-C4) -amino or 7) (C3-C6) -cycloalkyl-amino, Y represents an oxygen atom or -NH-, Z represents 1) alkyl (C1-C4) -amino, 2) N (CH3) - , 3) -C (CH3) 2-NH-, 4) -CH (CH3) -NH-, 5) -C (CH3) 2-N (R2) - wherein R2 means alkyl (Cj) or ) -NH-, when W represents alkyl (-C6), and active substance represents a compound with biological effect, which is linked through a radical with oxygen, a primary or secondary amino radical or an imino radical.

When n represents the integer 2 or 3, then the radicals R, independently of one another, represent the meanings mentioned in Rl) to R13). By the term "alkyl or alkenyl" is meant radicals whose carbon chain may be linear, branched or cyclic; double bonds can occur multiple times. The cyclic alkyl radicals are, for example, 3 to 6 membered monocycles, such as cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl. To the concept "heterocyclic radical of 5 to 14 members with 1, 2, 3 or 4 heteroatoms, taken from the group consisting of oxygen, nitrogen and sulfur" belong p. ex. radicals that are derived from pyrrole, azepine, pyrroline, pyridine, imidazole, pyrimidine, triazine, furan, 1,2-diazepine, oxazole, pyrazine, isoxazole, isoxazoline, thiazole, isothiazole, isothiazolidine, indole, quinoline, isoquinoline, benzimidazole, indazole , purine, pteridine, thiopyran or thiophene. Suitable physiologically compatible salts of the compound of the formula I are, for example, alkali metal, alkaline earth metal and ammonium salts, including those of organic ammonium bases. The monosaccharide term means radicals such as D-glucuronyl, L-iduronyl, D-glucopyranosyl, D-galactopyranosyl, N-acetyl-D-glucosaminyl, N-acetyl-D-galactosaminyl., D-mannopyranosyl or L-fucopyranosyl. The oligo- or poly-saccharides consist of 2 to 20 of the monosaccharides mentioned above, which are linked together by alpha- or beta-O-glycosidic bonds. The link between the monosaccharide and the radical Y is alpha- or beta-glycosidic. Appropriate enzymes, which produce the dissociation of the glycosyl radical with respect to the radical Y, are iduronidase, glucosidase, galactosidase, N-acetyl-D-glucosaminidase, N-acetyl-D-galactosaminidase, mannosidase, fucosidase or glucuro-nidase, preferably 0- glucuronidase. Suitable active substances are compounds such as anthracycline, preferably doxorubicin, 4'-epi-doxorubicin, 4- or 4'-deoxy-doxorubicin or a compound preferably selected from the group of etoposides, N-bis (2-chloro-ethyl). -4-hydroxy-acyltin, 4-hydroxy-cyclophosphamide, vindesine, vinblastine, vincristine, terfenadine, terbutaline, fenoterol, salbutamol, muscarine, oxyphenbutazole, salicylic acid, p-amino-salicylic acid, 5-fluoro-uracil, 5 -fluoro-cytidine, 5-fluoro-uridine, methotrexate, diclofenac, fluphena quinoline, 4-methylaminophenazone, theophylline, nifedipine, mitomycin C, mitoxantrone, camptothecin, m-AMSA, taxol, nocodazole, colchicine, cyclophosphamide, raquelmycin, cisplatin, melphalan, belomycin, nitrogen-gas mustard, phosphoramide-mustard gas, quercetin, genistein, erbstatin, tyrphostin, a derivative of rohituquine ((-) - cis-5, 7-dihydroxy-2- (2-chloro- phenyl) -8- [4- (3-hydroxy-l-methyl) -piperidinyl] -4H-l-benzopyran-4-one; EP 0366 061), retinolic acid, butyric acid, phorbol esters, aclaci-nomicin, progesterone, buserelin, tamoxifen, mifepriston, onapristone, N- (4-amino-butyl) -5-chloro-2-naphthalene-sulphonamides -da, pyridinyl-oxazol-2-one, quinolyl-oxazolon-2-one, isoquinolyl-oxazolon-2-one, staurosporine, verapamil, fors olin, l, 9-dideoxy-forskolin, quinine, quinidine, reserpine, 18- 0- (3, 5-dimethoxy-4-hydroxy-benzoyl) -reserpate, lonidamine, butionin-sulphoximine, a diethyldithiocarbamate, cyclosporin A, azathioprine, chlorambucil, N- (4-trifluoromethyl) -phenyl-2-cyanoamide -3-hydroxy-crotonic (PCT patent document WO 91/17748), 15-deoxy-spergualin, FK 506, ibuprofen, indomethacin, aspirin, sulfasalazine, penicillamine, chloroquine, dexamethasone, prednisolone, lidocaine, propafenone, procaine, mefenamine acid , paracetamol, 4-amino-phenazone, muskosin, orciprenaline, isoprenaline, amiloride, p-nitro-phenyl guanidinobenzoate or its substituted derivatives s additionally with one or more hydroxy, amino or imino groups. Preferred are prodrugs, in which the active substance is a cytostatic agent, or is an antimetabolite, in which the active substance is 5-fluoro-uracil, 5-fluoro-cytidine, 5-fluoro-uridine, cytosine-arabinoside or methotrexate, in which the active substance is a substance that is intercalated into DNA, in which the active substance is doxorubicin, daunomycin, idarubicin, epirubicin or mitoxantrone, in which the active substance inhibits topoisomerases I and II, in those which the active substance is camptothecin, an etoposide or m-AMSA, in which the active substance is a tubulin inhibitor, in which the active substance is vincristine, vinblastine, vindesine, taxol, nocodazole or colchicine, in which active substance is an alkylating agent, in which the active substance is cyclo-phosphamide, mitomycin C, rachelomycin, • cisplatin, phosphoramido-gas mustard, melphalan, bleomycin, nitrogen-gas mustard or N-bis (2-chloro-ethyl) -4-hydroxy-aniline, in which the sus Active substance is neocarcinostatin, calicheamicin, dynemycin, or esperamycin, in which the active substance is a compound that inactivates ribosomes, in which the active substance is verrucarin A, in which the active substance is a tyrosine inhibitor. -phosphokinase, in which the active substance is quercetin, genistein, erbstatin, tyrphostin or a rohituquine, in which the active substance is an inducer of differentiation, in which the active substance is retinolic acid, butyric acid, an ester of phorbol or aclacinomycin, in which the active substance is a hormone, an agonist of a hormone or an antagonist of a hormone, in which the active substance is progesterone, buserelin, tamoxifen, mifepristron or onapristone, in which the active substance is a substance that modifies the pyelotropic resistance against cytostatic agents, in which the active substance is a calmodulin inhibitor, in which the active substance is an inhibitor of protein kinase C, in which the active substance is an inhibitor of P-glycoproteins, in which the substance is a modulator of hexokinase bound to mitochondria, in which the active substance is an inhibitor of the β-glutamyl cysteine synthetase or glutathione-S-transferase, in which the active substance is an inhibitor of superoxide dismutase, in which the active substance is an inhibitor of the protein associated with proliferation, defined by the monoclonal antibody MAC Ki67, in the cell nucleus of a dividing cell, in which the active substance is a substance that exerts immunosuppressive effects, in which the active substance is a classic immunosuppressive agent, in which the active substance is a macrolide, in which the active substance is cyclosporin A, rapamycin, FK 506, in which the active substance is azathioprine, methotrexate, cyclophosphamide or chlorambucil, in which the active substance is a substance which has an anti-inflammatory effect River, in which the 'active substance is a non-steroidal anti-inflammatory substance, in which the active substance is a slow-acting anti-rheumatic drug, in which the active substance constitutes a steroid, in which the substance active substance is a substance having an antiphlogistic, analgesic or antipyretic effect, in which the active substance is a derivative of an organic acid, in which the active substance constitutes an analgesic / antiphlogistic agent of non-acidic nature, in which the substance active is oxyphenbutazone, in which the active substance is a local anesthetic, in which the active substance is an antiarrhythmic agent, in which the active substance is a Ca ++ antagonist, in which the active substance is an antihistamine agent, in those in which the active substance is a sympathomimetic agent, or in which the active substance is a substance with an inhibitory effect on human urokinase; and derivatives of the above-mentioned active substances, the active substance being linked via a radical with oxygen, an -NH- radical or an imino radical, to the radical Y of the compound of the formula I.

The active substance is also preferably the nitrogen-mustard gas compound mentioned in the examples, quinine or dipyridamole.

Preferred are prodrugs in which glycosyl represents a glucuronic acid separable enzymatically, W represents phenyl, R represents a hydrogen atom, CN, nitro, fluoro, chlorine or bromine, P is zero, n represents an integer, zero or 2, Y represents an oxygen atom, Z represents -N (CH3) -, -C (CH3) 2-NH, -CH (CH3) -NH-, -C (CH3) 2-N- (C-alkyl? -C4), -CH (CH3) -N- (C?-C4 alkyl) and active substance is a compound with biological effect bound through a hydroxy, amino or imino group. Especially preferred are the compounds 2- [N-methyl-N- [(4- (N, N'-bis- (2-chloro-ethyl) amino) -phenyloxycarbonyl] aminoj-4-nitro-phenyl-3-D acid -glucuronic acid, 2- [N-methyl-N- [(4- (N, N'-bis- (2-iodo-ethyl) amino) -phenyloxycarbonyl] amino] -4-nitro-phenyl-β-D- glucuronic The invention also relates to a process for the preparation of the prodrug of the formula I, which is characterized in that a compound of the formula II is reacted, glycosyl-Y [-C (= Y) -X-] pW (R) nZ (II), in which the radicals Glycosyl, Y, X, p, W, R, n and Z have the meanings mentioned in formula I, with an active substance, having an activated carboxyl, ammonium or imino radical, effected the reaction in the presence of a solvent taken from the group consisting of acetonitrile, dioxane, tetrahydrofuran, dichloromethane, dimethylformamide and acetone, and then the protecting groups are removed by hydrolysis. Activation of the active substance is effected, for example, according to H. J. Marley, Chem. Soc. Chem. Communication (1987) pages 112-113 or according to H. Hagemann Angew. Chem. 93 (1981) pages 813-814. The separation of the protecting groups is effected, for example, with an alkali metal hydroxide, alkali metal carbonate, an alkali metal cyanide, barium oxide, piperidine or morpholine, in the presence of methanol, ethanol or water. The invention also concerns medicaments, which are characterized by an effective content of at least one compound of the formula I and / or a physiologically compatible salt of the compound of the formula I, the radicals being glycosyl, Y, X, p , w, R, n, Z and active substance defined as above, in common with a pharmaceutically appropriate and physiologically compatible carrier material, an additive substance and / or other active substances and adjuvants. By virtue of the pharmacological properties, the compounds according to the invention are excellently suitable for the prophylaxis and therapy of all those diseases or disorders in which intracellular enzymes can be overexpressed and / or released or made accessible by cell destruction. they can dissociate the glycosyl radical. These are, above all, diseases such as cancer, autoimmune diseases or arthritis and acute and chronic arthropathies, conditioned by inflammatory, immunological or metabolic causes, especially tumor diseases and rheumatoid arthritis. The invention also concerns the use of the compound of the formula I for the preparation of medicaments for the prophylaxis and therapy of cancer diseases, autoimmune diseases and chronic inflammatory diseases, such as rheumatoid arthritis. The invention also relates to a process for the preparation of a medicament, which is characterized in that an at least one compound of the formula I is transported to an appropriate presentation form with a pharmacologically appropriate and physiologically compatible vehicle, and optionally other active substances. , additives or appropriate adjuvants. Suitable preparation forms are, for example, injectable solutions, in the preparation of which the usual adjuvants, such as vehicle materials, binding agents, swelling agents or lubricants and solubilizers, are used. Magnesium carbonate will be mentioned as frequently used adjuvants, titanium dioxide, lactose, mannitol and other sugars, talcum, milk albumin, gelatin, starch, cellulose and its derivatives, animal and vegetable oils such as cod liver oil, sunflower oil, peanut or sesame, poly (ethylene glycol) and solvents such as, for example, sterile water and uni- or pluri-valent alcohols such as glycerol. Liposomes or human proteins can also be used as carriers. Preferably, the pharmaceutical preparations are produced and administered within dosage units, each unit containing as active constituent a certain dose of the compound of the formula I according to the invention. In the case of injectable solutions, this dose may be up to about 10 g, but preferably from about 3 g to 5 g for an adult patient who weighs about 70 kg. In certain circumstances, higher or lower daily doses may also be appropriate. The administration of the daily dose can be effected either by a single dose in the form of a single dosage unit or also by several smaller dosage units, as well as by multiple intake of subdivided doses at determined intervals. The prodrugs according to the invention can also be used in all non-oncological diseases in which macrophages, granulocytes and thrombocytes occur, especially in the activated state. In the activated state, the aforementioned cells predominantly secrete intracellular enzymes, which makes possible a specific activation of a site of the prodrugs according to the invention. In the case of the oncological indication, the activation of the prodrugs according to the invention is effected by intracellular enzymes released from moribund tumor cells. This phenomenon occurs mainly in the case of larger tumors (diameter greater than 0.3 cm), but also after damage to the tumor by treatment with immunotoxins, cytostatic agents, irradiation, fusion proteins or conjugates of antibodies and enzymes. Since the glycosyl portion of the prodrugs according to the invention has been chosen in such a way that it can be separated only by enzymes released locally under pathophysiological conditions, the lipophilic drug can also be released only next to the target tissue and can develop there its cytotoxic effect. The superior effect of a prodrug according to the invention with a cytotoxic drug component can be increased by combining it with prodrugs according to the invention having another cytotoxic drug component. In this case, combinations of prodrugs are advantageous, in which cytotoxic components with different activity mechanisms are used, corresponding to the poly-chemotherapy. The use of active substances that very efficiently cause breaks of single chains and double chains in DNA, such as calicheamicin, is particularly appropriate. However, combinations of prodrugs according to the invention, in which one of the drugs has a cytotoxic potential and the other blocks multidrug resistance, are especially advantageous. In addition, the invention concerns pharmaceutical compositions, which contain a compound of the formula I and conjugates of an antibody and an enzyme. As conjugates of an anti-body and an enzyme are meant compounds which are fixed through the antibody part specifically to the tumor tissue or inflamed tissue, and which have a part of the enzyme which can dissociate the glycosyl radical of the compound of the formula I. Examples of such compounds are described in EP 0,501,215, EP 0,390,530 or EP 0,623,352.

Example i: Prodrug of a nitrogen-mustard gas derivative (F 373; Compound 11) was synthesized in the following manner: The starting material for the synthesis was 2-amino-4-nitro-phenol (Compound 1). Compound 1 was first monomethylated with the aid of methyl iodide (Compound 2) and the amino function was protected as a BOC derivative (Compound 3). The protected glucuronic acid was introduced by coupling with silver oxide of Compound 3 and bromide (Compound 4) to obtain Compound 5. After the BOC protecting group was separated with HCl, the amine was obtained (Compound 6). Compound 7 was reacted to form the chloroformate (Compound 8) and condensed with Compound 6 to form Compound 9. After dissociation or cleavage of the glucuronic acid part of Compound 3 in two steps (MeONa / MeOH , then aqueous NaOH) was obtained through Compound 10 the prodrug (Compound 11).

Compound 2: 2- (N-methylamino) -4-nitro-phenol (2) To a solution of 2-amino-4-nitro-phenol (1) (1.54 g, 10 mmol) and methyl iodide (1 mi, 16 mmol) in methanol (10 mL) was added triethylamine (2 mL, 14.4 mmol). After 1 hour at 40 ° C additional amounts of methyl iodide (1 ml) and triethylamine (1 ml) were added and stirred at 40 ° C for a further 2 hours. The reaction mixture was concentrated to dryness in vacuo, added to a 2N aqueous solution of sodium acetate and extracted with ethyl acetate. The organic phase was dried with sodium sulfate and chromatographed on silica gel (eluent: dichloromethane / methanol 95/5). Yield: 880 mg (52%).

C7H8N203 Calculated C: 50.02 H: 4.76 N: 16.73 Found C: 50.00 H: 4.80 N: 16.66 Melting point: 148 ° C (toluene) X H NMR (250 MHz, DMSO): d 7.44 (dd, Jorto = 9 Hz, Jmeta = 3.0 Hz, 1H), 7.13 (d, J = 3 Hz, 1H), 6.77 (d , J = 9 Hz, 1H), 2.76 (s, 3H).

IR (KBr): see "1) 3363 (OH), 1538, 1338 (N02).

MS (DC1, NH3): m / z [M + H] +: 169 Compound 3: 2- (N-Boc, N-methylamino) -4-nitro-phenol (3) To a solution of 2-N-methylamino-4-nitro-phenol (2) (4.18 g, 24.9 mmol) in tetrahydrofuran (50 ml) was added di-tert. dicarbonate. -butyl (14 g, 64.15 mmol), potassium carbonate (17 g, 123 mmol) and water (50 mL) and stirred at room temperature overnight. The reaction mixture was acidified with a saturated aqueous solution of ammonium chloride, extracted with ethyl acetate, dried with sodium sulfate and concentrated in vacuo. The crude product was stirred for two hours in methanol (100 ml) with potassium carbonate (17 g, 123 mmol), acidified with a saturated aqueous solution of ammonium chloride, extracted with ethyl acetate, dried with sodium sulfate and concentrated in vacuum The product was chromatographed on silica gel (eluent: dichloromethane / methanol 97.5 / 2.5). Yield: 6.2 g (93%).

C12H16N205: Calculated Cr 53.72 H: 6.01 N: 10.44 Found C: 53.64 H: 6.20 N: 10.36 Melting point: 197 ° C (toluene / petroleum ether) 1 H NMR (250 MHz, DMSO): d 8.10-8.00 (2H), 7.03 (d, J = 8.5 Hz, 1H), 3.04 (s, 3H), 1.40-1.30 (d. 9H).

IR (KBr): v (cm_1) 3129 (OH), 1672 (CO), 1529, 1339 (NO- MS (DC1, NH3): m / z [M + H] +: 269, [M + H-C4H8] +: 213, [M + H- C4H8OCO] +: 169 Compound 5: methyl ester of 2- ( N-BOC, N-methylamino) -4-nitro-f-enyl-2,3,4-tri-0-acetyl- / 8-D-glucuronic acid (5) To a solution of methyl ester bromide of acid 2, 3 , 4-tri-0-acetyl-aD-glucuronic acid (4) (126 mg, 0.317 mmol) in acetonitrile (5 mL) was added with silver oxide (0.23 g, 0.92 mmol) and 2- (N -BOC, N-methylamino) -4-nitro-phenol (3). The reaction mixture was stirred for 1 hour at room temperature, filtered on Celite and concentrated in vacuo. The product was chromatographed on silica gel (eluant dichloromethane / methanol 97.5 / 2.5). Yield 165 mg (89%).

C25 32N2 ° 14: Calculated C: 51.37 H: 5.52 N: 4.79 Found C: 51.79 H: 5.72 N: 4.66 Melting point 80 ° C (toluene / petroleum ether) [Q!] D20 = -39 ° (c = l, 02 in CHC13) 1 H NMR (250 MHz, CDC13): d 8.14 (dd, Jorto = 9 Hz, Jmeta = 2.5 Hz, 1H), 8.15-8.05 (1H), 7.30-7.20 ( 1H), 5.45-5.30 (4H), 4.25 (d, J = 9 Hz, 1H), 3.73 (s, 3H), 3.13 (s, 3H), 2.15- 2.05 (9H), 1.65-1.40 (9H).

IR (CDC13): vícpT1) 1760 (CO, ester), 1699 (CO) carbamate), 1529, 1349 (N02).

MS (DC1, NH3): m / z [M + NH4] +: 602 Compound 6: Methyl ester of (N-methylamino) -4-nitro-phenyl-2,3,4-tri-0-acetyl- acid -D-glucuronic (6) A solution of 2- (N-BOC, N-methylamino) -4-nitro-phenyl-2,3,4-tri-0-acetyl-3-D-glucuronic acid methyl ester ( 5) (3 g, 5.13 mmol) in 2.12 M hydrochloric acid in ethyl acetate (60 ml) was stirred for 1 hour at room temperature. The solution was poured into a saturated aqueous solution, in excess, of sodium bicarbonate, and extracted with ethyl acetate. The organic phase was dried with sodium sulfate and concentrated. The product was chromatographed on silica gel (eluent: dichloromethane / methanol 97.5 / 2.5). Yield 2.14 g (86%) solid yellow material.

C20H24N2O12: Calculated C: 49.59 H: 4.99 N: 5.78 Found C: 49.81 H: 5.12 N: 4.80 Melting point: 120 ° C (in toluene) [a] D20 = -58 ° (c = 1.04 in CHC13) * H NMR (300 MHz, CDC13): d 7.53 (dd, Jorto-8.5 Hz, Jmßta, 2.5 Hz, 1H), 7.37 (d, J = 2.5 Hz, 1H), 6.91 (d, J = 8.5 Hz, 1H), 5.45-5.30 (1H), 5.16 (d, J = 7 Hz, b, 1H), 4.50 (d, J = 5 Hz, 1H), 4.24 (d, J = 9 Hz, 1H), 3.75 (s, 3H), 2.90 (d, J = 5 Hz, 1H), 2.10-2, 05 (9H).

IR (CDC13): vfcrrf1) 3443 (NH), 1758 (CO), 1553, 1346 (N02).

MS (DC1, NH3): m / z [M + H] +: 485.

Compound 8 Chloroformate of 4- [N, N-bis- (2-chloro-ethyl) amino] phenyl (8) Phosgene in toluene (1.93 M) (8 mL, 15.4 mmol) was added to a suspension of 4- [N, N-bis (2-chloro-ethyl) amino] phenol hydrochloride (7) (1, 85 mmol) in tetrahydrofuran (30 ml) and stirred for 30 minutes at 0 ° C. After adding triethylamine (1.0 mL, 7.17 mmol), it was stirred at 0 ° C for another hour. Then, the suspension was filtered and concentrated in vacuo at room temperature. Flash chromatography on silica gel with dichloromethane as eluent provided the product as a colorless liquid, which was used immediately in the next reaction. Yield 70%.

XH NMR (250 MHz, CDC13): d 7.10 (d, J = 9 Hz, 2H), 6.66 (d, J = 9 Hz, 2H), 3.73 (t, J = 6.5 Hz , 4H), 3.63 (t, J = 6.5 Hz, 4H).

IR (CDC13): víff1) 1779 (CO), 1514 (aromatic).

MS (DC1, NH3): m / z [M + H] +: 296, [M + 2 + H] *: 298.

Compound 9: 2- (N-methyl-N- [4- (N, N'-bis- (2-chloro-ethyl) amino] phenyloxycarbonyl] amino] -4-nitro-phenyl-2,3-methyl ester , 4-tri-0-acetyl-jS-D-glucuronic acid (9) To a solution of chloroformate of 4- [N, N-bis (2-chloro-ethyl) amino) phenyl (8) (0.31 g, 1.04 mmol) in tetrahydrofuran (15 mL), diisopropyl-ethylamine (0.25 mL, 1.44 mmol) was added 2 (N-methylamino) -4-nitro-phenyl-2,3-methyl ester, 4-tri-0-acetyl-β-D-glucuronic acid (6) (0.50 g, 1.03 mmol) and boiled under reflux for 2 hours. After cooling to room temperature, it was concentrated. The product was chromatographed on silica gel (eluant dichloromethane / methanol 97.5 / 2.5). Yield: 487 mg (64%) C3iH35N3014Cl2 Calculated C: 50.00 H: 4.74 N: 5.64 Cl: 9.52 Found C: 49.90 H: 4.82 N: 5.62 Cl: 9.78 Melting point: 101 ° C (methanol) [] O? = -47 ° (c = 1.10 in CHC13) XH NMR (300 MHz, CDC13): d 8.25-8.15 (2H), 7.45-7.35 (1H), 7.25-7.05 (1H), 6.95-6.85 (1H), 6.70-6.55 (2H), 5.45-5.25 (4H), 4.28 (d, J = 9 Hz, 1H), 3.80-3.55 (11H) , 3.38 (s) 2 diastereomeric carbamates (40/60) 3H, 3.27 (s), 2.20-2.00 (9H).

IR (CDC13): vícpT1) 1760 (CO, ester), 1722 (CO, carbamate), 1530, 1350 (NO,).

MS (DC1, NH,) m / z [M + H] +: 744; [M + 2 + H] +: 746, [M + Na] +: 766; [M + 2 + Na] +: 768 Compound 10: 2- [N-methyl-N- [(4- (N, N'-bis- (2-chloro-ethyl) amino) phenyloxycarbonyl] amino acid methyl ester ] -4-Nitro-phenyl-jβ-D-glucuronic (10) To a suspension of 2- [N-methyl-N- [4- (N) methyl ester., N '-bis- (2-chloro-ethyl) amino) phenyloxy] amino] -4-nitro-phenyl-2, 3,4-tri-0-acetyl- / 3-D-glucuronic (9) (68 mg , 0.0915 mmol) in methanol (5 mL) was added at -15 ° C sodium methylate (2 mg, 0.037 mmol) and stirred for 6 hours at -15 ° C. After neutralization with an ion exchanger (Amberlite IRC-50 S) and filtration, the solution was concentrated and chromatographed on silica gel with ethyl acetate as eluent. Yield 50 mg (89%).

C25H29N2011C12 XH NMR (300 MHz, CDC13): d 8.22 (if, 1H), 8.16 (d, J = 8.5 Hz, 1H), 7.25 (d, 1H), 7.15-6, 90 (2H), 6.80-6.45 (2H), 5.06 (1H), 4.09 (1H), 4.20-3.15 (17H).

IR (CDC13): vícpT1) 3601, 3448 (OH), 1714 (CO), 1528, 1349 (N02).

MS (ES): m / z [M + Na] +: 640; [M + 2 + Na] +: 642.

Compound 11 2- [N-Methyl-N- [(4- (N, N'-bis- (2-chloro-ethyl) amino) -phenyloxycarbonyl] amino] -4-nitro-phenyl-jS-D-glucuronic acid (eleven) To a solution of Compound 10 (50 mg, 0.0809 mmol) in acetone (4 mL) was added at -15 ° C 0.3 mL of an aqueous solution i N of sodium hydroxide. The mixture was stirred for 2 hours at -15 ° C, neutralized with 1 N aqueous hydrochloric acid and concentrated in vacuo (T <40 ° C). The product was chromatographed on silica gel with a mixture of acetonitrile and water (9/1). The yield was 45 mg (89%). (^ 24H27N30llC -! - 2 XH NMR (250 MHz, CD3OD): d 8.30 (yes, 1H), 8.24 (dd, Jorto = 9 Hz, Jmeta = 2.5 Hz, 1H), 7.52 (d, J = 9Hz, 1H), 6.99 (d, J = 9 Hz, 2H), 6.69 (d, J = 9Hz, 2H), 5.23 (1H), 3.89 (d, J = 9Hz, 1H), 3.80-3.33 (1H).

IR (KR): victimTT1) 3418 (OH), 1705 (CO), 1516, 1349 (NO,) MS (ES): m / z [M + H] +: 603; [M + 2-H] +: 605 Example 2 2- [N-Methyl-N- [(4- (N, N'-bis- (2-iodo-ethyl) amino) phenyloxycarbonyl] amino] -4-nitro-phenyl-8-D-glucuronic acid ( 12) (Compound 12): The synthesis was carried out analogously to Example l Example 3 (Compound 13): Quinine prodrug (F 391) The synthesis was carried out analogously to Example 1 Example 4 (Compound 14): Prodrug of dipyridamole (F 392) The synthesis was carried out analogously to Example 1 Example 5: Enzymatic Dissociation of F 373 The prodrug F373 (Compound 11), when incubated with β-glucuronidase, is enzymatically dissociated to form a nitrogen-aromatic mustard gas derivative (4- [N, N-bis- (2-chloro -ethyl) -amino] phenol) (Compound 7), glucuronic acid and the spacer.

F 373 Glucuronic acid Compound 7 Spacer (Compound 15) The prodrug F 373 is stable in solution in anhydrous DMSO. To investigate the dissociability, 10 μl of a solution of F 373 (5 mg / ml in DMSO) was mixed with 180 μl of 0.02 M phosphate buffer pH 7.2 and 10 μl of E. coli β-glucuronidase. (Sigma) (330 μg / ml) and incubated at 37 ° C. A 25 μl batch was diluted with 225 μl of a mixture of 0.1 M phosphate buffer pH 3.0 (85%) and acetonitrile (15%) and analyzed immediately with the aid of the following HPLC system.

HPLC system: The system used for HPLC (high pressure liquid chromatography) consisted of a gradient pump (Gynko-tek, model 480), a sampler (Abimed, model 231/401), a UV detector (Beckman, model 166, detection wavelength 212 nm) and an evaluation unit (Beckman, Gold system). The separations were carried out on a RP column (Zorbax SB-C 18.5 μm, 125 * 4.6 mm). The mobile phase was formed based on two components according to the following scheme: A - acetonitrile B - 0.02 M phosphate buffer of pH 3.0. 0 min: 15% of A, 85% of B, 15 min: 75% of A, 25% of B, 25 min: 75% of A, 25% of B, 27 min: 15% of A, 85% of B, 35 min: 15% A, 85% B.

The following peak surfaces were found: Time F 373 Compound 7 Compound spacer 15 min RT = 1 2.6 RT = 10.7 RT = 14, 1 0 1 9.41 0 0 1 10.46 3.32 1, 92 5 0.35 7.43 4 , 75 7 0 7,33 5,37 10 0 6,79 5,43 1 5 0 5,58 5,43 25 0 4,00 5,70 60 0 0,88 6,53 Example 6: Cytotoxicity of F 373, * 391 and 392 on tumor cells in the presence and absence of 0-glucuronidase. In a 96-well microtiter plate, 2 x 103 LoVo cells per well were seeded in 100 μl of MEM + 10% FKS. After 24 h the substances under test were added to 100 μl of medium in the desired concentration and optionally 0-glucuronidase (final concentration 50 μg / ml, Sigma G 7896). Each group consisted of 4 wells, the witness was incubated only with the medium. After 65 h, 50 μl of MTT (2 ml) was added thereto., 5 mg / ml in PBS) and after 3 h the supernatant was removed. The dye formed by the living cells was dissolved by the addition of 100 μl of DMSO / well. The extinction was measured for each well with the help of a Multiscan 340 CC photometer (Flow entity) at 492 nm. The values of the 4 wells per group were averaged and from them the dose and effect curve were calculated as well as the 50% inhibitory concentration, IC50 with Software GraFit 3.0.

Substance ssiinn ?? - GGlluucc .. with? -Gluc. (prodrug) C Cll5500 eenn /// mmooll Cl50 in // mol F 373 > > 550000 6.3 F 391 > > 440000 113 F 392 > > 440000 49.5 The toxic compound in the prodrug F 373 is Compound 7 (Example 1, page 14) and has, alone tested, an IC 50 of 5 μmol. The toxic compound present in the prodrug F 391 is quinine and has alone tested an IC 50 of 103 μmol. The toxic compound in the prodrug F 393 is dipyridamole and has, alone tested, an IC 50 of 43 μmol.

Claims (8)

1. CLAIMS 1.- Compound of formula I
2. Glycosyl-Y [-C (= Y) -X-] p-W (R) n-Z-C (= Y) - active substance (I) and / or physiologically compatible salts of the compound of the formula I, in which glycosyl represents an enzymatically dissociable poly-, oligo- or mono-saccharide, W represents 1) an aromatic radical of 5 to 14 members, 2) naphthyl, 3) indenyl, 4) anthryl, 5) phenanthryl, 6) a heterocyclic radical of 5 to 14 members with 1, 2, 3 or 4 heteroatoms taken from the group consisting of oxygen, nitrogen and sulfur, 7) alkyl (C ^ Cg), ) (C2-C6) alkenyl, 9) (C3-C6) cycloalkyl or 10) phenyl, R represents 1) a hydrogen atom ,. 2) alkyl (C? -C4), 3) phenyl, 4) methoxy, 5) carboxy, 6) methyloxycarbonyl, 7) -CN, 8) -OH, 9) -N02, 10) halogen, such as fluoro, chloro or bromine, 11) sulfonyl, 12) sulfonamido or 13) sulfon-alkyl (C 1 -C 4) -amido, p represents zero or 1, n represents zero, 1, 2 or 3, X represents 1) an oxygen atom, 2 ) -NH-, 3) methyleneoxy, 4) methyleneamino, 5) methylene-alkyl (Cx-C4) -amino, 6) alkyl (Cx-C4) -amino or 7) (C3-C6) -aminocycloalkyl-amino, Y represents an oxygen atom or -NH-, Z represents 1) alkyl (CL-CJ) -amino, 2) -N (CH3) -, 3) -C (CH3) 2-NH-, 4) -CH (CH3) -NH-, 5) -C (CH3) 2-N (R2) - wherein R2 means (Cx-C4) alkyl or 6) -NH-, when W represents alkyl (Ci-Cg), and active substance represents a compound with biological effect, which is linked through a radical with oxygen, a primary or secondary amino radical or an imino radical. 2. Compound of the formula I according to claim 1, wherein Glycosyl represents a glucuronic acid separable by enzymatic route, WW rreepprreesseennttaa phenyl, R represents a hydrogen atom, CN, nitro, fluoro, chlorine or bromine, P is zero , n represents an integer, zero 1 or 2, YY rreepprreesseennttaa an oxygen atom, Z represents -N (CH3) -, -C (CH3) 2-NH, -CH (CH3) -NH-, -C (CH3 ) 2-N- (Cx-C4 alkyl), -CH (CH3) -N- (C -C alkyl.)
3. 3. Compound of the formula I according to claim 1 or 2, characterized in that the compound of the formula I is 2- [N-methyl-N- [(4- (N, N'-bis- (2-chloro-ethyl) amino) -phenyloxycarbonyl] amino] -4-nitro-phenyl-β-D-glucuronic acid or acid 2- [N-methyl-N- [(4- (N, N '-bis- (2-iodo-ethyl) amino) -phenyloxycarbonyl] amino] -4-nitro-phenyl- / 3-D-glucuronic,
4. 4. - Process for the preparation of a compound of the formula I, characterized in that a compound of the formula II is reacted, glycosyl-Y [-C (= Y) -X-] pW (R) nZ (II), wherein the glycosyl radicals Y, X, p, W, R, n and Z have the meanings mentioned in formula I according to claim 1, with an active substance having an activated carboxyl, ammonium or imino radical, the reaction being carried out in the presence of of a solvent taken from the group consisting of acetonitrile, dioxane, tetrahydrofuran, dichloromethane, dimethylformamide and acetone, and then the protecting groups are removed by hydrolysis.
5. 5. Medicaments, characterized by an active content of at least one compound of the formula I according to one or more of claims 1 to 3, and / or of a physiologically compatible salt of the compound of the formula I, the glycosyl radicals being defined, Y, X, p, W, R, n, Z and active substance as in formula I according to claim 1, in common with a pharmaceutically appropriate and physiologically compatible carrier material, an additive material and / or other active substances and adjuvants
6. 6. Use of the compound of formula I according to claim 1, for the preparation of medicaments for the prophylaxis and therapy of cancerous diseases, autoimmune diseases and chronic inflammatory diseases such as rheumatoid arthritis.
7. 7. Process for the preparation of a medicament, characterized in that an at least one compound of the formula I according to claim 1 is brought into an appropriate form of presentation, together with a pharmaceutically appropriate and physiologically compatible vehicle and optionally with other active substances , additives or appropriate adjuvants.
8. 8. - Pharmaceutical preparation, which contains a compound of formula I according to one or more of claims 1 to 3, and a conjugate of an antibody and an enzyme, which can enzymatically dissociate the glycosyl radical in the formula.