MXPA99008680A - Benzamidas sustitui - Google Patents

Abstract

The present invention relates to: Substituted benzamides and their use in medicaments, in particular as immunomodulators, are described

Description

SUBSTITUTE BENZAMIDES Description of the Invention The invention relates to substituted benzenes of the general formula I and its use in medicines. Autoimmune diseases arise due to a reactivity of the immune system against the body's own structures. As a result, the normally existing tolerance against the tissue of the body itself is lost. In the pathogenesis of the various autoimmune diseases, T lymphocytes and monocytes / macrophages, in addition to antibodies, have a determining role. Activated monocytes / macrophages secrete a multitude of different mediators that promote inflammation, which are directly or indirectly responsible for the destruction of tissues affected by the autoimmune disease. The activation of monocytes / macrophages takes place either in the interaction with T lymphocytes or through bacterial products such as lipopolysaccharide (LPS). The activation of monocytes / macrophages and granulocytes induced by various bacterial products is additionally characteristic of inflammatory reactions in general. The significance of the balance between inflammatory-promoting cytokines (for example interleukin IL-12) and inflammation-inhibiting cytokines (for example interleukin IL-10) for the development and course of inflammations or autoimmune diseases is clearly documented based on a multitude of experimental investigations with animals and important clinics. In various animal models of diseases, such as rheumatoid arthritis, multiple sclerosis, diabetes mellitus as well as cutaneous and mucosal inflammatory diseases, the pathophysiological significance of IL-12 is demonstrated (Immunol Today 16/8: 383-387, 1995; Im unol., 155: 4661-4668, 1995; J. Exp. Med. 182: 1281-1290, 1995; J. Exp, Med. 187/4: 537-546, 1998). Through the application of IL-12 it was possible to cause the respective disease, and after the neutralization of the endogenous IL-12 a weakening of the course of the disease was manifested until a cure of the animals.

In the case of inflammatory bowel diseases, both in sick animals and in patients with Crohn's disease there is a markedly increased reactivity of T cells in inflamed intestinal sections. This is characterized by increased expression of IL-12 and IFN. -? in the injuries. In contrast, the immunosuppressive cytokine IL-10 is clearly diminished in the lesions (Immunity 3: 171-174, 1995; J. Exp. Med. 182: 1281-1290, 1995; Eur. J. Immunol. 26: 1156- 1163; Eur. J. Immunol., 28: 379-389, 1998). The significance of the immunosuppressive cytokine IL-10 for the development of inflammatory bowel diseases is also demonstrated in that the IL-10 blocked mice developed a spontaneous colitis (Immunity 3: 171-174, 1995). The activation of the T cells that produce IFN-? in the lamina propria of the intestine is substantially based on the local formation of IL-12. In the animal model of an allergic-induced colitis, it could be shown that antibody therapy against IL-12 is possible in the case of existing severe colitis. The neutralization of IL-12 with antibodies led to a clinical and histopathological normalization of the diagnoses within a few days. In the case of the T cells of the lamina propria of the mice treated against IL-12, formation of IFN-α could not be confirmed. (J. Exp. Med. 182: 1281-1290). The first applications of recombinant IL-10 in humans confirm the inhibitory properties of inflammations. After administration of IL-10 to healthy subjects, the formation of TNF-a and IL-1 cytokines that promote inflammation by monocytes activated ex vivo with LPS is reduced from 65 to 95% (J. Immunol., 154: 5492-5499, 1995). ). The application of IL-10 to patients with Crohn's refractory steroid morbus results in an improvement of the clinical symptomatology (Gatroenterology, 113: 383-389). Recently, the subcutaneous application of IL-10 was also reported in 3 patients with psoriasis. There was an improvement in the symptomatology of the disease. In addition, the formation of IL-12 and TNF was reduced as well as the expression of surface molecules on monocytes (J. Clin.Invest.101: 783-794). The application of antibodies against IL-12 in humans is imminent. In summary it can be established that a defect of IL-10 or an excess of IL-12 conditions the pathophysiology of a multitude of inflammatory diseases. Therefore, the approaches to normalize the IL-10 / IL-12 balance have an enormous therapeutic potential. Therefore, the task on which the invention is based was to develop new immunomodulators that do not lead to general immunosuppression, and in doing so they produce a normalization of the IL-10 / IL-12 equilibrium. It was now discovered that certain substituted benzamides satisfy the requirements that were required of the substances to be developed. Accordingly, the substituted benzamides of the formula I are the subject of the invention wherein R1 represents a group of the formula COOR4, in which R4 represents a straight or branched chain alkyl radical with 1-6 carbon atoms, or represents a group of the formula C0NR5R6 in which R5 and R6 are the same or different and they mean an alkyl group with 1-6 carbon atoms (straight or branched chain), or together with the N atom means a pyrrolidine ring, of piperidine, hexamethyleneimino or morpholino, R2 means chloro, fluoro, CF3, a radical alkyl with 1-3 carbon atoms) or hydrogen, and R3 represents the hydroxy group, an alkyl or alkoxy radical with 1-6 carbon atoms (straight or branched chain and optionally substituted with OH-, an alkoxy, ester or an open chain or cyclic amido group with 1-6 carbon atoms), or a radical CH = - NR5R6 in which R5 and R6 are as defined above. The compounds according to the invention can be present both in the racemic form as well as in the enantiomeric form, or in the form of salts with pharmaceutically acceptable acids. Preferred are substituted benzamides in which the radical R 1 represents a group of the formula COOR 4, in which R 4 represents a straight or branched chain alkyl radical with 1-6 carbon atoms, or represents a group of the formula CONR 5 R 6 in the that R5 and R6 together with the N atom signify a morpholino ring, R2 is H, and R3 represents the hydroxy group, or a radical CH2-NR5R6 in which R5 and R6 together with the N atom signify a morpholino ring. Particularly preferred are 2- (morpholino-4-carbonyl) -N- (l-morpholin-4-ylmethyl-2,6-dioxo-piperidin-3-yl-benzamide and the methyl ester of N- (l-hydroxy) -2,6-dioxo-piperidin-3-yl) -phthalamide It is possible to obtain the compounds of the general formula I by first transforming a carboxylic acid of the formula Ilia or Ilb. ilb in a manner known per se in an ester (R1 = COOR) or in an amide (R1 = C0NR5R6). With this, compounds according to the invention are already obtained from carboxylic acid IIb. In those compounds correspondingly derived from the carboxylic acid Ia it is then possible to introduce in a known manner the radical R3 which is different from the hydroxy group, for example by the reaction of Mannich with paraformaldehyde and a secondary amine of the formula HNR5R6. Another object of the invention is the use of the substituted benzamides of the formula I in medicaments, in particular as immunomodulators. The substances according to the invention inhibit the formation of the cytokine IL-12 which promotes inflammation by human monocytes activated with LPS. On the other hand substances of this group increase the formation of the cytokine IL-10 which inhibits inflammation by human monocytes activated with LPS. This differentiates the new subtypes from the immunomodulators known as spheroids and phosphodiesterase inhibitors, which suppress both the synthesis of IL-12 and that of IL-10. By virtue of its characteristic unomodulatory activity (inhibition of IL-12, increase of IL-10), the substances according to the invention are suitable for the treatment and / or prophylaxis of inflammations, in particular cutaneous and mucosal inflammations. , vasales, as well as for the treatment and / or prophylaxis of autoimmune diseases. These include, among others, skin inflammations (eg, atopic dermatitis, psoriasis, exema), inflammations of the respiratory tract (eg bronchitis, pneumonia, bronchial asthma, ARDS). (syndrome of respiratory insufficiency in adults), sarcoidosis (sílicosa / fibrosa), inflammations of the gastrointestinal tract (for example gastroduodenal ulcer, morbus Crohn, ulcerative colitis), as well as diseases such as hepatitis, pancreatitis, appendicitis, peritonitis, nephritis, aphtosis, conjunctivitis, keratitis, uveitis, rhinitis. Autoimmune diseases include diseases of the arthritic morphological environment (for example rheumatoid arthritis, diseases associated with HLA-B27), as well as multiple sclerosis, juvenile diabetes or lupus erythematosus. Other indications are septis, bacterial meningitis, cachexia, rejection reactions to transplants, graft versus host reactions, as well as reperfusion syndrome and arteriosclerosis. In addition to at least one compound of the formula In accordance with the invention, the medicaments according to the invention contain vehicles, fillers, solvents, diluents, colorants and / or binders.

The choice of these auxiliary substances as well as the amounts to be used thereof will depend on the form of administration of the drug, whether oral, intravenous, intraperitoneal, intradermal, intramuscular, intranasal, buccal or local. For the oral application, preparations in the form of tablets, chewable tablets, dragees, capsules, granules, drops, juices or syrups are suitable for parenteral, topical and inhalation application solutions, suspensions, dry preparations of easy reconstitution, as well as aspersions. The compounds according to the invention in dissolved form in a tank, in a carrier sheet or a plaster, optionally with the addition of subcutaneous penetration promoting agents, are suitable examples of percutaneous application forms. The compounds according to the invention can be released in delayed form from oral and percutaneous application preparations. The amount of active substance that should be administered to patients varies according to the weight of the patient, the form of application, the indications and the severity of the disease. In general, from 1 to 150 mg / kg of at least one compound of the formula I according to the invention are applied.

Examples Table 1 The substances in Table 1 were characterized spectroscopically according to ""? -RMN (Apparatus: DPX 300 Advance of Bruker company, 300 MHz; Solvent: DMSO-d6; Indication of chemical deviation in ppm).

Example 1: 1, 97-2.16 (m, 2H, CH2); 2.52 - 2.86 (m, 2H, CH2); 3.20-3.75 (m, 8H, CH2); 4.50 - 4.65 (, 2H, NCH2N); 4.64 - 4.82 (m, 1H, CH); 7.49-7.66 (m, 3H, aro at); 7.79 - 7.85 (d, 1H, aromat); 8.60 - 8.72 (d, 1H, CONH). Example 2: 1.94-2.20 (m, 2H, CH2); 2.68-2.95 (m, 2H, CH2); 3.28 - 3.70 (m, 8H, CH2); 4.52 - 4.65 (m, 2H, NCH2N); 4.83-4.96 (m, 1H, CH); 7.42-7.58 (m, 3H, aromat); 7.85-7.92 (m, 2H, aromat); 8.85 - 8.89 (d, 1H, CONH). Example 3: 1.93-2.24 (m, 2H, CH2); 2.69 - 3.02 (m, 2H, CH2); 3.12 (S, 2H, CH2); 3.35 - 3.77 (m, 12H, CH2); 4.52-4.68 (m, 2H, NCH2N); 4.74-4.98 (m, 1H, CH); 7.28 - 7.32 (d, 1H, aromat); 7.45-7.59 (, 2H, aromat); 7.74 - 7.80 (d, IH, aromat); 8.78 - 8.88 (d, IH, CONH). Example 4: 2.01 - 2.18 (m, 2H, CH2); 2.58-2.88 (m, 2H, CH2); 3.79 (s, 3H, COOCH3); 4.70-4.80 (m, 1H, CH); 7.54 -7.72 (m, 3H, aromat); 7.77 - 7.81 (d, 1H, aromat); 8.77 -8.82 (d, 1H, CONH); 10.21 Cs, 1H, NOH). Example 5: 1.38-1.46 (m, 6H, CH2); 1.96 -2.16 (m, 2H, CH2); 2.20-2.32 (m, 4H, CH2); 2.60-2.92 (m, 2H, CH2); 3.26 - 3.75 (m, 8H, CH2); 4.57-4.70 (m, 2H, NCH2N); 4.72 - 4.88 (, 1H, CH); 7.28-7.30 (d, 1H, aromat); 7.43-7.57 (m, 2H, aromat); 7.70-7.74 (m, 1H, aromat); 8.75 - 8.78 (d, 1H, CONH). Example 6: 1.18-1.26 (t, 3H, CH3); 1.36-1.48 (, 6H, CH2); 1.95 - 2.16 (m, 2H, CH2); 2.22-2.34 (m, 4H, CH2); 2.60 - 2.90 (, 2H, CH2); 4.08-4.18 (q, 2H, OCH2); 4.55-4.68 (m, 2H, NCH2N); 4.70-4.80 (m, 1H, CH); 7.54-7.72 (m, 3H, aromat); 7.78 - 7.83 (d, 1H, aromat); 8.78 - 8.82 (d, 1H, CONH). Investigation of immunomodulatory activity From human peripheral blood cells (PBMC), human monocytes were isolated and obtained by Ficoll density gradient centrifugation of heparinized plethora. For this the PBMC were incubated with a monoclonal antibody that is directed against the monocyte-specific surface molecule CD14 and to which superparamagnetic micro-lobes are coupled (Miltenyi Biotech, Bergisch Gladbach). For a positive selection of the marked monocytes from the PBMC cell mixture, the total cell suspension was placed on a column with a ferro agnostic carrier matrix, and this was placed in a magnetic field. Through this, the cells loaded with the microbeads were ligated to the carrier matrix, the unlabeled cells passed the column and were discarded. After removing the matrix from the magnetic field, the cells loaded with antibodies were eluted by washing the demagnetized column with a regulator. The purity of this population of CD14 positive monocytes obtained in this manner was about 95 to 98%. These monocytes were incubated for one hour at 37 ° C and 5% C02 in a culture medium (RPMI, supplemented with 10% serum from fetal calves) with a density of 106 cells / ml with the test substances dissolved in DMSO . Then 20 μg / ml of E. coli LPS was added. After 24 hours cell-free culture supernatants were taken and assayed for the cytokine content of IL-12 and IL-10. The concentration of IL-12 and IL-10 in cell culture supernatants was determined by sandwich ELISA using two anti-IL-12 and anti-IL-10 monoclonal antibodies (Biosource Europe, Fleurus, Belgium). A standard reference curve with IL-12 and human IL-10 was included. The detection limit of the ELISA assay for IL-12 was 10 pg / ml, that of the ELISA assay for IL-10 of 15 pg / ml. Table 2: Influence of the test substances on the production of IL-12 and IL-10 of monocytes activated with LPS.

Table 2 The results presented in Table 2 demonstrate that the immunomodulators known as dexamethasone, pentoxifylline and rolipram inhibit both the formation of IL-12 and also that of IL-10 in monocytes activated with LPS. In comparison, benzamides of similar structure substituted with carboxyl groups (Example 1) only show negligible activity at high dosages. Surprisingly, the esters according to the invention (examples 4 and 6) and the amides according to the invention (examples 3 and 5) of the substituted benzamides have immunomodulatory activity in the model examined. Such compounds potently inhibit the synthesis of IL-12 by monocytes activated with LPS at concentrations of 10, 6 and 2 μg / ml. In contrast to the known immunomodulators, however, they also increase the synthesis of IL-10. This characteristic model (evident inhibition of IL-12, increased IL-10 in the same concentration range of substances) distinguishes a novel type of immunomodulator.

Claims (1)

1. CLAIMS Substituted benzamides of the formula I wherein R1 represents a group of the formula COOR4, in which R4 represents a straight or branched chain alkyl radical with 1-6 carbon atoms, or represents a group of the formula C0NR5R6 in which R5 and Rs are the same or different and meaning an alkyl group with 1-6 carbon atoms (straight or branched chain), or together with the N atom means a ring of pyrrolidine, of piperidine, hexamethyleneimino or morpholino, R2 means chloro, fluoro, CF3, a alkyl radical with 1-3 carbon atoms) or hydrogen, and R 3 represents the hydroxy group, an alkyl or alkoxy radical with 1-6 carbon atoms (straight or branched chain and optionally substituted with OH-, an alkoxy group, ester or an open chain or cyclic amido group with 1-6 carbon atoms), or a radical CH2_NR5R6 wherein R5 and R6 are as defined above. Substituted benzamide of the formula I according to claim 1, characterized in that R1 represents a group of the formula COOR4, in which R4 represents a straight or branched chain alkyl radical with 1-6 carbon atoms, or represents a group of the formula CONR5R6 wherein R5 and R6 together with the N atom signify a morpholino ring, R2 is H, and R3 represents the hydroxy group, or a radical CH2_ NR5R6 in which R5 and R6 together with the N atom signify a morpholino ring. Use of a substituted benzamide of the formula I according to claim 1 or 2 as active substance in a medicament. Use according to claim 3, characterized in that the medicament is an immunomodulator.