WO2013078523A1 - Heterocyclic n-glycinyl-n-acyl hydrazone compounds, synthesis method, pharmaceutical compositions and treatment method - Google Patents

Abstract

New heterocyclic derivatives are described, having the N-glycinyl-N-acyl hydrazone subunit and acting as analgesic and anti-inflammatory agents with in vitro e in vivo inhibiting activity on pro-inflammatory TNF-α cytokine, and useful therefore for the treatment of pain and acute and chronic inflammation, including: relieving hyperalgesia associated with acute and chronic inflammation in mammals, preferably humans. Also described are pharmaceutical compositions containing these compounds and methods for preparing the same.

Description

 N-GLYCINYL-N-ACYLIDRAZONIC COMPOUNDS

 HYPEROCYCLICS, SYNTHESIS PROCESS, COMPOSITIONS

 PHARMACEUTICALS AND TREATMENT METHOD. Descriptive Report

Field of the Invention

 The present invention relates to heterocyclic derivatives having the β-glycinyl- / V-acylidrazone subunit. More particularly, the present invention relates to substituted (E) - / V '- (N-benzylidene) -2- (3-tert-butyl-1-phenyl-1H-pyrazol-5-amino) acetohydrazides or not, and their isomers, to a process for their preparation, to pharmaceutical compositions containing them and their use as anti-inflammatory and analgesic therapeutic agents inhibiting tumor necrosis factor alpha (TNF-α) production, particularly in the treatment of of chronic inflammatory diseases.

Background of the Invention

 Cytokines are hormone - // 7 soluble proteins that allow communication between cells and the external environment. It is a broad term that includes lymphokines, monokines, interleukins, colony stimulating factors, interferons (IFNs), tumor necrosis factor (TNF) and chemokines (Tayal, V.; Kalra, BS Eur. J. Pharmacol. 2008, 579, 1 .).

 In the primary immune response that occurs in response to an infection or injury, macrophages produce proinflammatory cytokines such as tumor necrosis factor alpha (TNF-), interleukin-6, and interferon γ, which play important roles in maintaining the inflammatory process. associated. Cytokines have many functions in the body, such as mediating and regulating immunity, the onset of the inflammatory response and hematopoiesis, but the largest group of cytokines is involved in cell proliferation and differentiation (McInnes, IB; Schett, G. Nat. Rev. Immunol. 2007, 7, 429.).

Cytokines are secreted by white blood cells and also by a variety of other body cells (fibroblasts, endothelial cells, epithelial cells, etc.) in response to inducing stimuli, and are not constitutively expressed. Proinflammatory cytokines are highly expressed in injured tissues and are induced when macrophages are exposed to gram negative bacterial endotoxins. Therefore, cytokine synthesis inhibitors prevent the immune response to invasive stimuli.

 The production of proinflammatory cytokines, e.g. TNF-α, IL-1β and IL-6, is key to chronic inflammatory diseases such as rheumatoid arthritis (RA), Crohn's disease, psoriasis and asthma. In addition to inflammatory diseases, there is evidence of cytokine involvement in other diseases, including heart failure, ischemic retinopathies, and the development of insulin resistance in diabetes (Moller, D.E. Trends Endocrinol. Metab. 2000, 11, 212.).

 Due to the numerous possible clinical applications for cytokine inhibitors such as inflammatory diseases, cancer, immunotherapy, bone diseases, metabolic diseases, wound healing, antiviral therapy, among others, many pharmaceutical industries have invested in the development of orally active molecules capable of act on molecular targets involved in the regulation of proinflammatory cytokine production.

The Alpha Tumor Necrosis Factor (TNF-g) The tumor necrosis factor family is primarily involved in regulating cell proliferation and apoptosis, but TNF-cc also has proinflammatory properties, orchestrating a number of events related to genesis and apoptosis. maintenance of the inflammatory response. The precursor form of TNF-α, cell membrane-bound TNF-a (TNFm or pro-TNF), is expressed as a 26 kD polypeptide on the cell surface of activated macrophages and lymphocytes, as well as other cell types (endothelium). . TNF-α gene transcription is complexly regulated by multiple transcription factors such as nuclear factor κΒ (NF-κΒ), AP-1, NFIL-6 and NFAT. The mitogen-activated protein kinase signaling pathway p38 (MAPK p38) also plays a critical role in the production of TNF-α (Campbell, I.K .; Roberts, L.J .; Wicks, I. P. Immunol. Celi Biol. 2003, 81, 354.).

TNF-α synthesis is regulated by messenger RNA stability control, allowing rapid responses to external stimuli such as lipopolysaccharide (LPS), a structural component of the wall of gram-negative bacteria. The presence of adenylate-uridylate-rich elements in the 3'-UTR region of TNF-α messenger RNA plays a major role in post-transcriptional repression and is a target for rapid degradation or inhibition of translation. The MAP38 p38 signaling pathway is the main signaling pathway that stabilizes TNF-α through adenylate uridylate-rich elements in the 3'-UTR region (Dumitru, CD, Ceci, JD, Tsatsanis, C, Kontoyiannis, D. Stamatakis, K., Lin, J.-H., Patriotis, C., Jenkins, NA, Copeland, NG, Kollias, G., et al., Celi 2000, 103, 1071.).

 Membrane-bound TNF is further cleaved by a metalloprotease, the TNF-α converting enzyme (TACE), which releases the soluble form of TNF-α, a 17 kD polypeptide. Inhibition of TACE as a means of reducing soluble TNF-α levels is a strategy that has been studied by various research groups for the treatment of inflammatory diseases and several compounds are in the preclinical and clinical studies phase ( Moss, M. L.; Sklair-Tavron, L.; Nudelman, R. Nat. Clin. Prac. Rheumatol. 2008, 4, 300.).

 Another approach that can be explored for TNF-α inhibition is the inhibition of biosynthesis of its precursor form, TNFm or pro-TNF, by compounds that inhibit the activation of nuclear factor κΒ, a transcription factor that leads to production. RNA for protein translation, which include many pro-inflammatory cytokines, such as ΙΚΚβ (1) and (2) inhibitors, phosphodiesterase (PDE) inhibitors, specifically PDE4 (rolipram (3) and roflumilast (4) inhibitors. ) which are able to decrease mRNA accumulation for TNF-α), p38 MAPK inhibitors (SB-203580 (5) and BIRB-796 (6)) and thalidomide analogues (7) such as lenalinomide (8). ) (Figure 1 ).

TNF-α biological responses are mediated by two structurally distinct receptors on the cell surface, type I (TNFR1 or p55 / p60) and type II (TNFR2 or p75 / p80). Secreted TNF-α binds to both type I receptor and type II receptor, whereas membrane-bound TNF-α binds mainly to type II receptor. The TNF-α converting enzyme (TACE) can also cleave both the type I receptor and the type II receptor generating soluble TNF-α receptors. So TNF-a receptors exist both as a membrane-bound form that modulates the pathophysiological effects of TNF-α, and as a soluble form that can bind and neutralize TNF-α actions (Campbell, IK; Roberts, LJ .; Wicks, IP Immunology and Celi Biology 2003, 81, 354-366.)

 Both TNF-α receptors, TNF-R1 and TNF-R2, can activate NF-κΒ transduction pathways by activating ΙκΒ kinase and protein kinase C (PKC) proteins, and MAPK's promoting a variety of cellular responses (Locksley, RM; Killeen, N.; Lenardo, MJ; Celi 2001, 104, 487.). For the TNF-R1 signaling complex, receptor activation leads to the recruitment of RIP (receptor integrating protein) and TRAF-2 (TNF receptor-associated factor) signaling proteins through TRADD (TNF receptor associated death domain protein) which ultimately leads to caspase activation and cell death. RIP is a serine / threonine kinase crucial for NF-κΒ activation while TRAF-2 activates MAP kinases (Declercq, W .; Vanden Berghe, T .; Vandenabeele, P .; Celi 2009, 138, 229-232. ; Chan, FKM; Siegel, RM; Lenardo, MJ; Immunity 2000, 13, 419-422.)

 High TNF-α production is associated with a number of pathological conditions of autoimmune and inflammatory origin such as Crohn's disease, psoriasis and rheumatoid arthritis (RA). In these diseases TNF-α modulates processes such as immune cell activation, proliferation, apoptosis and regulation of leukocyte migration. Specifically, TNF-α is involved in the mechanisms of inflammation and joint destruction in rheumatoid arthritis (RA). Large amounts of TNF-α are detected in the synovial membranes of RA patients in the acute and chronic stages of the disease (Feldmann, M. Nat. Rev. Immunol. 2002, 2, 364.).

Therapy with anti-TNF-ct biological agents has been instrumental in validating TNF-α as a therapeutic target for RA, as these agents have been shown to be effective in improving clinical manifestations and progression of damage in both RA patients. as in animal models of arthritis (Toussirot, E.; Wendling, D. Expert Opin. Pharmacother. 2004, 5, 581.). The main innovations for the treatment of arthritis are the use of drugs such as Enbrel® (etanercept), manufactured by Wyeth Laboratories; Remicade® (infliximab) from Schering-Plow laboratories; and more recently Abbott's Humira® (adalimumab), all inhibitors of TNF-α action. While infliximab and adalimumab are recombinant monoclonal antibodies that bind only to TNF-α, etanercept is a soluble TNF receptor fusion protein that binds to both TNF-α and TNF-β (Mclnnes, IB; Schett, G. Nat. Fiei (Immunol. 2007, 7, 429).

 All three anti-TNF agents are approved for the treatment of rheumatoid arthritis and Crohn's disease. However, only etanercept has been approved for juvenile chronic arthritis since TNF-β is elevated in inflamed tissues in this disease.

 Although all TNF-α inhibitors result in clinical improvement in about 70% of RA patients, including some patients who do not respond to conventional therapies, most patients have only limited improvement with the use of these agents. In addition there are common problems related to parenteral administration of these drugs, such as local reactions at the site of administration. In addition, the onset of serious side effects such as bacterial sepsis, tuberculosis and other opportunistic infections, multiple sclerosis and lupus are described (Campbell, I.K.; Roberts, L.J .; Wicks, I. P. Immunol. Celi Biol. 2003, 81, 354.). Thus, the major contraindications to the use of anti-TNF agents include untreated tuberculosis, pulmonary infections, and multiple sclerosis.

The approval of these drugs from biotechnological processes proves the efficacy of therapeutic strategies based on TNF-α modulation. With this, there is a need to identify new mechanisms for TNF-α modulation, different from the aforementioned drugs, given the high cost of producing these drugs compared to micromolecules resulting from traditional medicinal chemistry strategies. In addition to the high cost, there are disadvantages such as the impossibility of oral administration, increased hypersensitivity reactions and compromised immune system through continued use of anti-TNF monoclonal antibodies licensed for use in humans (ie Enbrel®, Remicade®, Humira®) (Palladino, MA; Bahjat, FR; Theodorakis, EA; Moldawer, LL Nat. Rev. Drug Discov. 2003, 2, 736-746.).

Mitogen Activated Protein Kinase p38 (MAPK p38) MAPK p38 is a stress-activated protein kinase that modulates responses to cellular stressors such as ultraviolet light and osmotic shock. MAPK p38 is a 38 kD polypeptide (p38) that belongs to a family of four isoforms (α, β, γ, δ). This family can be divided into two groups based on the ability to respond to different stimuli, p38 α / β and ρ38γ / δ (Korb, A.; Tohidast-Akrad, M .; Cetin, E.; Axmann, R .; Smolen , J .; Schett, G. Arthr. Rheum. 2006, 54, 2745.) · All isoforms are activated by dual phosphorylation at residues Thr180 and Tyr182. This phosphorylation induces a conformational change in the protein, allowing ATP and substrate to bind. The protein kinase required for p38 MAPK phosphorylation depends on the stimulus and cell type. MKK3 and MKK6 phosphorylate p38 MAPK within minutes of exposure to various activating stimuli. The duration of phosphorylation is crucial in regulating cell fate, sustained phosphorylation is often associated with apoptosis induction, in contrast, transient phosphorylation may be associated with growth factor-induced cell survival. Signaling duration is controlled by phosphatases, including protein phosphatase 1, protein phosphatase 2A or MAPK phosphatases. These enzymes can be activated by phosphorylated p38 MAPK, creating negative feedback that regulates the actions of active p38 MAPK (Cuadrado, A.; Nebreda, A. R. Biochem. Journal 2010, 429, 403.).

The four isoforms of p38 MAP kinases are encoded by different genes and have different tissue expression patterns, with p38a being expressed at significant levels in most cell types, while the others appear to be expressed in a more tissue-specific manner, for example. for example, ρ38β in the brain, ρ38γ in skeletal muscle and p38õ in endocrine glands. Members of the p38 family overlap substrate specificities, although some differences have been reported, with particular substrates being better phosphorylated by p38a and ρ38β than by ρ38γ and ρ38δ or vice versa. Analysis of synovial tissue extracted from patients with RA suggests that α and γ isoforms are overproduced in inflamed tissue and may be the preferred targets for disease intervention (Korb, A.; Tohidast-Akrad, M .; Cetin, E .; Axmann, R.; Smolen J. Schett, G. Aiihr Rheum 2006, 54, 2745.)

 For more than 20 years the Smith Kline & French pharmaceutical laboratory has developed a new class of pyridinylimidazole derivatives, e.g. SKF-86002 (9), which have shown efficacy in animal models of chronic inflammatory disease. These compounds were called cytokine synthesis inhibiting anti-inflammatory drugs (CSAID's) (Boehm, J.C; Smietana, JM; Sorenson, ME; Garigipati, RS; Gallagher, TF; Sheldrake, P.L; Bradbeer, J.; Badger, AM; Laydon, JT; Lee, J.C; Hillegass, LM; Griswold, DE; Breton, JJ; ChabotFletcher, M.C; Adams, JLJ Med. Chem. 1996, 39, 3929.) In the 1990s, John Lee and colleagues found that SKF-86002 (9) was capable of suppressing the synthesis of IL-1 and TNF-α in monocytes (Lee, J. C; Laydon, JT; McDonnell, P. C; Gallagher, TF; Kumar, S.; Green, D.; McNulty, D.; Blumenthal, MJ; Heys, JR; Landvatter, SW; Strickler, JE; McLaughlin, MM; Siemens, IR; Fisher, SM; Livi, GP ; White, JR; Adams, J.L; Young, PR Nature 1994, 372, 739.). Subsequently, more potent inhibitors of TNF-α production of the same class of heterocyclic derivatives, e.g. SB-202190 (10), were described as being able to prevent translation of messenger RNA encoding TNF-α. A protein that specifically binds to these pyridinylimidazole compounds has been purified from cell extract and identified as a mitogen-activated protein kinase (MAPK) protein kinase. This protein was independently identified by different groups and crystallographically characterized and is commonly referred to as p38 MAPK (Wilson, KP; McCaffrey, PG; Hsiao, K.; Pazhanisamy, S.; Galullo, V .; Bemis, GW; Fitzgibbon, MJ; Garon, PR; Murcko, MA; Su, MSS Chem. Biol. 1997, 4, 423.).

The finding that p38 MAPK played a key role in inflammatory processes, including chronic processes, made enzyme inhibition an attractive therapeutic strategy for the treatment of different pathological conditions of inflammatory origin. Currently several p38 MAPK inhibitors are in clinical study phases (Goldstein, DM; Kuglstatter, A.; Lou, Y .; Soth, MJJ Med. Chem. 2010, 53, 2345).

 Activation of p38 MAPK results in several adaptive responses through p38-dependent phosphorylation of serine and threonine residues on a wide variety of substrates, primarily kinases and transcription factors. P38 MAPK kinase substrates include activated MAPK protein kinases (MKs) such as MK2, MK3, MK5, as well as various other kinases. MK2 is believed to be among the most important substrates of p38 MAPK in mediating the inflammatory response to cell stress. In addition, activated p38 MAPK increases cytokine production by direct phosphorylation of transcription factors (eg ATF2, CREB) and direct or indirect stabilization (via lower kinases) or increased mRNA translation encoding proinflammatory cytokines ( Schett, G.; Zwerina, J.; Firestein, G. Ann. Rheum. Dis. 2008, 67, 909.).

 MAP38 p38 inhibitors have been shown to be prophylactically effective in reducing clinical severity (paw edema, inflammation, cartilage degradation, and bone erosion) in different models of arthritis in rats and mice. These disease suppressive effects could also be seen in therapeutically treated animals, suggesting that p38 MAPK inhibitors have therapeutic potential for patients with well-established disease (Thalhamer, T.; McGrath, MA; Harnett, MM Rheumatology 2008, 47, 409.).

 This validation of the therapeutic potential of p38 MAPK inhibitors in inflammatory diseases has led to the development of a large number of inhibitors such as Boehringer Ingelheim's BIRB-796 (6) (doramapimod®), which was one of the first MAPK p38 inhibitors to enter. in clinical trials (Regan, J.; Breitfelder, S.; Cirillo, P.; Gilmore, T.; Graham, AG; Hickey, E.; Klaus, B.; Madwed, J.; Moriak, M .; Moss, N.; Pargellis, C.; Pav, S.; Proto, A.; Swinamer, A.; Tong, L.; Torcellini, C. J. Med. Chem. 2002, 45, 2994.).

Numerous p38a MAPK inhibitors have advanced in clinical study steps (Figure 2). Some of them, such as Boehringer Ingelheim's BIRB-796 (6), Vertex's VX-702 (11), Roche's Paramapimod (12) and SCio-469 (13), had their studies discontinued due to lack of efficacy in the studies. Clinics For Crohn's Disease And Rheumatoid Arthritis Or Adverse Effects that seem to be related to the chemical structure of some particular agents and not to the mechanisms of action of these compounds. To date no toxic effects observed have been conclusively related to inhibition of p38 MAPK. Many candidates for p38 MAPK inhibitor drugs are in the clinical trial phase, for example: Bristol-Myers Squibb BMS-582949 (14), which is in phase 2 for rheumatoid arthritis; ARRY-797 (structure not disclosed) in phase 2 for acute inflammatory pain; ARRY-614 (undisclosed structure) in phase 1 for myelodysplastic syndrome; PH-797804 (15) in phase 2 for rheumatoid arthritis and neuropathic pain; GS-856553 (losmapimod (16)) in phase 2 for COPD, cardiovascular disease and depression; and SB-681323 (dilmapimod (17)) in phase 2 for neuropathic pain (Goldstein, MD; Kuglstatter, A.; Lou, Y .; Soth, MJ; J. Med. Chem. 53, 2345-2353.

Due to the importance of modulating proinflammatory cytokine biosynthesis as a therapeutic intervention strategy in the treatment of several inflammatory diseases, particularly TNF-, whose therapeutic potential is validated by the successful use of biotechnological drugs. And, due to the limitations arising from the chronic use of these drugs, such as the high cost of production and the occurrence of serious associated side effects, there is a growing need for the discovery of new micromolecules capable of modulating the biosynthesis of this cytokine. For years, the MAPK pathway has been emerging as a promising target for modulating pro-inflammatory cytokine production, and numerous pharmaceutical industries have been investing efforts to develop drug candidates capable of acting as MAP38 p38 inhibitors. The results from clinical studies with these inhibitors in development for diseases such as rheumatoid arthritis, neuropathic pain, among others, encourage further research to modulate this important pharmacological target.

 Structures of synthetic derivatives described in the literature as inhibitors of TNF-α biosynthesis that act on different therapeutic targets.

Examples of p38 'MAPK inhibitors that have entered the phase of clinical studies. Description of the Figures

Figure 1 - Effect of the compounds LASSBio-1504 and LASSBio-1506 (100 μΓΤΐο K / kg, orally) and thalidomide (100 μιτιοΙ / kg, intraperitoneal) on carrageenan-induced thermal hypernociception, n = 5-10 (rats), ^* p <0.05; ^** p <0.01; ^*** p <0.001 (Student's t-test).

Figure 2 - Effect of LASSBio-1504 and LASSBio-1506 (100 μΐτιοΙ / Kg, v.o) on TNF-α levels in carrageenan-injected paws. Rats received LASSBio-1504, LASSBio-1506 or saline and intraplantar injection of carrageenan or saline 1 h later. Four hours after intraplantar carrageenan injection, the paws were homogenized and TNF-α in the supernatant was determined by ELISA.

Figure 3 - Effect of LASSBio-1504, LASSBio-1506 and thalidomide compounds (100 μιηοΙ / kg, vo, once daily, 1 h before mechanical sensitivity assessment) in the mBSA-induced arthritis model, n = 5-7 (mice), ** p <0.01; ^*** p <0.001 (Student's t-test).

Summary of the Invention

The main object of the invention are compounds of formula (la), (Ib), (Ma) and (Nb) belonging to the family A-glycinyl-A / -acylidrazone, as well as their derivatives, isomers, salts and racemates. capable of being used as anti-inflammatory and / or analgesic acting on the biosynthesis of proinflammatory cytokines.

The second object of the present invention is the process of synthesizing the compounds of formula (la), (Ib), (IIa) and (IIb), as well as their derivatives, isomers, salts and racemates thereof.

A pharmaceutical composition containing the compounds of formula (Ia), (Ib), (IIa) and (Mb) as well as their derivatives, isomers, salts and racemates thereof is also object of the present invention. Another object of the present invention is a pharmaceutical composition containing the compounds of formula (la), (Ib), (Ma) and (Nb), as well as their derivatives, isomers and racemates thereof, being employed as anti-inflammatory and / or action-related analgesics on the biosynthesis of proinflammatory cytokines.

The last object of the present invention is the method of treating diseases or disorders related to the biosynthesis of proinflammatory cytokines. Detailed Description

The compounds of formula (Ia), (Ib), (Na) and (Mb) of the N-glycinyl- / V-acylidrazone family, as well as their derivatives, isomers, salts and racemates are capable of being used as anti -inflammatory or analgesic drugs, also having an action on the biosynthesis of proinflammatory cytokines.

These compounds, besides having the N-glycinyl-N-acylhydrazone subunit, which give it the characterization of the chemical family they are inserted, may have distinct substituents on their radicals, where R ₁ , and R ₂ may be: hydrogen, alkyl, cycloalkyl , aryl, heterocycles, orthoalkylphenyl, orphoxyalkylphenyl, orfoalkylsulfonylphenyl, ortho-thioalkylphenyl, orthoalkylsulfoxylphenyl, ortho-phenylsulfonamides, o-aminophenyl, ortho-amidophenyl-carbophenyl-ortho-phenylphenyl , ortho-cyanophenyl, ortho-nitrophenyl, mefa-alkylphenyl, meta-alkoxyphenyl, meta-alkylsulfonylphenyl, methathioalkylphenyl, meta-alkylsulfoxylphenyl, meta-phenylsulfonamides, meta-aminophenyl, meta-amidophenyl, meta-amidophenyl, meta-alkylphenyl - carboalkoxyphenyl, meta-cyanophenyl, meta-nitrophenyl, para-alkylphenyl, para-cycloalkylphenyl, para-alkoxyphenyl, para-alkylsulfonylphenyl, para-thioalkylphenyl, para-alkylsulfoxylphenyl, para-phenylsulfonates, para-phenyls ulfonamides, para-aminophenyl, para-amidophenyl, para-halophenyl, para-carboalkoxyphenyl, para-cyanophenyl or para-nitrophenyl;

W is hydrogen, orthoalkyl, ortho-cycloalkyl, ortho-alkoxy, ortho-cycloalkoxy, ortho-thioxyl, ortho-aroxy-oxide, ortho-sulfones, ortho-sulfides, ortho-sulfonates, orthos-sulfonamides, ortho-sulfonamides. amino, ortho-starch, ortho-halides, ortho-carboalkoxy, ortho-carbothioalkyl, ortho-trihaloalkane, ortho-cyano, ortho-nitro, meta-alkyl, meta-cycloalkyl, meta-alkoxy, meta-cycloalkoxy, mefa-thioxyl, mefa-arioxyl, mefa-sulfones, mefa-sulfides, mefa-sulfoxides, mefa-sulfonates, mefa-sulphonamides, meia-amino, mefa-starch, mefa-carbalcoxyl, mefa-carbothioalcoxyl, mefa- trihaloalkane, mefa-cyano, half-nitro, para-alkyl, para-cycloalkyl, para-alkoxy, para-cycloalkoxy, para-oxyoxy, para-aroxy, para-sulfones, para-amphides, para-sulfoxides, para-sulfonates, -sulfonamides, para-amino, para-starch, para-halides, para-carboalkoxy, para-carbothioalkyl, para-trihaloalkane, para-cyano or para-nitro.

 The compounds of formula (Ia), (Ib), (IIa) and (IIb), as well as their derivatives, isomers, salts and racemates are capable of being used as anti-inflammatory and / or analgesics in disorders or diseases. related to the biosynthesis of proinflammatory cytokines in human and non-human mammalian animals.

 The proinflammatory cytokines referred to in this invention are all those which act on the proliferation process, cell differentiation and the inflammatory process. Even more specific, the proinflammatory cytokines referred to in this invention are Interleukins (IL), Tumor Necrosis Factors (TNF) and Interferon (INF), further comprising all the isoforms that these proinflammatory cytokines may present.

 The compounds of formula (Ia), (Ib), (IIa) and (IIb), as well as their derivatives, isomers, salts and racemates, have the ability to inhibit protein kinase (MAPK) signaling cascades leading to the production of proinflammatory cytokines.

 Protein kinases are signaling molecules that are characterized as factors in signal translation and propagation of the immune response. And just like cytokines, protein kinases have isoforms that induce the cytokine biosynthesis process. Therefore, protein kinases isoforms are also targets for the compounds of formula (la), (Ib), (Ma) and (llb).

 The process of producing the compounds of formula (la), (Ib), (lia) and

(11b), as well as their derivatives, isomers, salts and racemates, were obtained in good chemical yields using the synthetic methodology described herein, starting from commercially available compounds, which qualifies this synthetic methodology for industrial use. The synthesis of the compounds of this invention comprises the following steps:

 A) carbonyl condensation;

 B) nucleophilic substitution to carbonyl;

 c) N-alkylation;

 D) cyclocondensation;

 E) multicomponent coupling reaction

A pharmaceutical composition may be prepared having as pharmaceutically active compounds the compounds of formula (Ia), (Ib), (IIa), (IIb), as well as their derivatives, isomers, salts and racemates. Such pharmaceutical compositions are used in the treatment of diseases and disorders in human and non-human mammalian animals.

 The present pharmaceutical composition, in addition to the pharmaceutically active compounds described above, comprises non-active pharmaceutically active compounds such as diluents, dispersants, sweeteners, antioxidants, among others known to one skilled in the art.

 Diseases and / or disorders in this invention are understood to be those related to inflammatory processes. Such diseases and / or disorders are more specifically related to the biosynthesis process of proinflammatory cytokines such as Interleukins (IL), Tumor Necrosis Factors (TNF) and Interferon (INF), further comprising all the isoforms that these pro cytokines produce. -inflammatory drugs may present.

 The diseases and / or disorders of this invention still have a relationship to processes modulated by mitogen-activated protein kinase (MAPK) and its isoforms, which have a variety of functions, such as inducing biosynthesis of proinflammatory cytokines. .

 The last object of this invention is the method of treating diseases and / or disorders by administering the compounds of formula (Ia), (Ib), (Na), (IIb) as well as derivatives, isomers, salts thereof. and racemates.

Diseases and / or disorders are related to inflammatory processes, being more specific to the pro-inflammatory cytokine biosynthesis process. In addition, the treatment method comprises diseases and / or disorders related to processes modulated by mitogen activated protein kinase (MAPK).

The following examples illustrate the process of synthesis and pharmacological action in vitro and in vivo of some of the derivatives of the compounds of formula (Ia), (Ib), (IIa) and (IIb), which are objects of this invention. The following examples should not be used to limit the scope of protection of this invention.

EXAMPLES

Example 1 Preparation of 3-Tere-Butyl-1-phenyl-5-aminopyrazole derivative

 A solution of phenylhydrazine (0.83 mL, 8.39 mmol) and 4,4-dimethyl-3-oxopentanenitrile (2.0 g, 8.0 mmol) in toluene (3 mL) was heated to reflux. The reaction is monitored by thin layer chromatography. When the total consumption of phenylhydrazine is observed, toluene is evaporated and the crude oil obtained is recrystallized from hexane and the product is obtained as a white solid in 76% yield. Mp = 50-52 ° C.

Example 1.1 Preparation of ethyl 2- (3-tert-Butyl-1-phenyl-5-aminopyrazol) -acetate derivative

To a mixture of 3-tert-butyl-1-phenyl-5-aminopyrazole (100 mg, 0.465 mmol) in toluene (3.0 ml) and triethylamine (0.1 ml) is added ethyl 2-bromoacetate (0 0.06 ml). The mixture is stirred and refluxed for four hours. When the reaction is over the medium is partitioned between water and ethyl acetate, the organic phase is evaporated and the product is purified by column chromatography (hexane-ethyl acetate; concentration gradient). The product is obtained as a yellow oil in 60% yield. Example 1.2

Preparation of 2- (3-tert-Butyl-1-phenyl-5-aminopyrazol) Acetohydrazide derivative

 To a 50 ml flask was added 600 mg (2 mmol) of ethyl 2- (3-tert-butyl-1-phenyl-5-aminopyrazol) acetate, 20 eq of 100% hydrazine hydrate and 5 ml of ethanol. The mixture is stirred and refluxed for 2h when TLC indicates total consumption of the starting ester. The mixture is poured onto ice and the product extracted with dichloromethane and obtained as a yellow oil in 80% yield.

Example 1.3

Preparation of phenylpyrazolic A-glycinyl-A-acylhydrazones

To a 50 ml flask containing 1.6 mmol of 2- (3-tert-butyl-1-phenyl-5-aminopyrazolyl) acetohydrazide is added 10 ml of ethanol, catalytic concentrated hydrochloric acid and 1.68 mmol (1.05 eq) of the aldehyde. The mixture is stirred for approximately 2h at room temperature. After completion of the reaction the volume of ethanol is reduced, saturated sodium bicarbonate solution and crushed ice are added to the reaction. The formed precipitate is filtered off, or the mixture is extracted with ethyl acetate (or dichloromethane).

(EJ-AT 4 -chlorobenzylidene-1 H -S-tert-butyl-1-phenyl-S-aminopyrazole)

acetohydrazide (LASSBio-1506)

White solid obtained by recrystallization from ethanol (R = 70%) (mp = 193 ° C). H NMR (ppm, DMSO-d _6, 200 MHz) 1 1 62 and 1 1, 60 (2s, 1H, NHCO); 8.27 and 8.01 (2s, 1H, N = CH); 7.75-7.30 (m, 9H, H-Ar); 5.53 and 5.46 (2s, 1H, CH-pyrazole); 4.21 and 3.78 (2s, 2H, CH ₂ ); 1.23 (s, 9H, (CH ₃ ) ₃ ).

³ C NMR (δ ppm; DMSO-d ₆ , 50 MHz): 171.6 (CO); 161.5 (C3-pyrazole); 148.9 (C5-pyrazole); 143.1 (H = N); 139.1 (C1'-phenylpyrazole); 134.9 (C-CI); 133.5 (C1 - phenyl); 129.8 (2H-Ar); 129.4 (2H-Ar); 129.2 (2x1 H-Ar); 127.0 (CH-Ar); 123.9 (2x CH-Ar); 85.3 (CH-pyrazole); 46.7 (CH ₂ ); 32.5 (C (CH ₃ ) ₃ ); 30.7 ((CH ₃ ) ₃ ).

IR (KBr) (cm ^-1 ): 3362 (NH), 2952 (CH), 1686 (C = O).

Elemental Analysis calculated (C 22 H 24 N 5 OCl): C: 64.46; H: 5.90; N: 17.09.

Elemental Analysis Found: C: 64.24; H: 5.83; N: 16.75.

(E) -W ^, - ((4- (2-morpholininooxy) naphthyl) methylene) -2- (3-tert-butyl-1-phenyl-5-aminopyrazole) acetohydrazide (LASSBio-1504)

 White solid recrystallized from acetone (R = 75%) (mp = 180 ° C).

¹ H NMR (δ ppm; DMSO-d ₆ , 300 MHz): 11, 37 and 1.28 (2s, 1H, NHCO); 8.91 and

8.72 (2d, 1H, J = 9 Hz, CH-naphthyl); 8.72 and 8.53 (2s, 1H, N = CH); 8.26 (m, 1H,

CH-naphthyl); 7.77 (m, 1H, CH-naphthyl); 7.40-7.60 (m, 6H, ArH); 7.33 (m, 1H, ArH);

7.04 (d, 1H, J = 9Hz, ArH); 5.64 and 5.35 (2t, 1H, J = 5.7 Hz, NH); 5.45 (s, CH-pyrazole); 4.33 (t, 2H, J = 4.6 Hz, -OCH ₂ -CH ₂ -); 4.24 and 3.78 (2d, 2H, J = 5.7 Hz, CH ₂ ); 3.56 (t, 4H, J = 4.7 Hz, 2xCH ₂ -morpholine); 2.88 (t, 2H, J = 4.7 Hz, -OCH ₂ -

CHb-); 2.56 (t, 4H, J = 4.7 Hz, 2xCH ₂ -morpholine); 1.23 (s, 9H, (CH ₃ ) ₃ ).

³ C NMR (δ ppm; DMSO-d ₆₎ 75 MHz): 170.6 and 166.1 (C = 0); 161.0 (C3-pyrazole);

155.7 (C4-naphthyl); 148.2 (C5-pyrazole); 147.7 and 144.3 (N = CH); 139.2 (C1'-phenyl);

131.0 (C8a-naphthyl); 129.2 (ArCH); 127.8 (ArCH); 126.2 (ArCH); 125.7 (ArCH); 125.2 (ArCH); 124.6 (ArCH); 124.0 (ArCH); 123.1 (C1-naphthyl); 122.3 (ArCH);

121.7 (C4a-naphthyl); 105.3 (ArCH); 84.5 (CH-pyrazole); 66.4 (OCH ₂ -ethoxy); 66.22

(CH ₂ -morpholine); 56.9 (CH ₂ -ethoxy); 53.6 (CH ₂ -morpholine); 48.0 and 46.5 (CH ₂ );

31, 9 _{((CH3) 3);} 30.2 (3x H ₃ ).

IR (KBr) (cm ^-1 ): 3468 (NH), 2964 (CH), 1655 (C = O), 1572 (CN).

Elemental Analysis calculated (C 32 H 38 6 O 3 H 2 O): C: 67.11; H: 7.04; N: 14.67. Elemental analysis determined: C: 67.16; H: 7.05; N: 14.74.

98% purity determined by HPLC (mobile phase: acetonitrile-phosphate buffer 60:40 (pH 7.4). Example 2

The examples described here illustrate the tests performed with 2 compounds obtained, LASSBio-1504 and LASSBio-1506, however, the test results should not be used to limit the scope of protection.

The new phenylpyrazole N-glycinyl- / V-acylhydrazonic derivatives were subjected to in vitro pharmacological screening in the model of inhibition of TNF-α production by culture of peritoneal macrophages from LPS-stimulated mice (100 ng / ml). The most active compounds selected for in vitro pharmacological screening had their anti-inflammatory and antinociceptive pharmacological profile evaluated in in vivo models of acute and chronic inflammation.

 The results showed significant inhibitory activity on TNF-α production in vitro and in vivo, as well as important antihypernociceptive activity in models of acute and chronic inflammation when administered orally, particularly the LASSBio-1504 derivative.

EXAMPLE 2.1 Effect of Compounds on Carraqenin-Induced Thermal Hypernociception

The most potent phenylpyrazole A / -glycinyl-A / -acylhydrazonic derivatives in the model of inhibition of TNF-α production in vitro were chosen to evaluate their oral antiinflammatory and antinociceptive profiles. The compounds LASSBio-504 and LASSBio-1506 were tested in the carrageenan-induced thermal hypernociception model. The compounds were administered orally at a dose of 100 μηηοΙ / kg. Thalidomide (100 μηιοΙ / kg, ip) was used as a positive control because it is an anti-TNF-α drug. Analysis of the results shows that the compounds LASSBio-1504 and LASSBio-1506 were effective as antihypernociceptive agents. Although LASSBio-1504 and LASSBio-1506 showed similar capabilities to inhibit in vitro α-TFN production (IC ₅ o = 3.6 μΜ and IC ₅ o = 1,6 μΜ, respectively), the LASSBio-1504 derivative was more active in vivo, being able to completely inhibit hypernociceptive stimulation while LASSBio-1506 was able to partially inhibit. This difference in pharmacological profile in vivo may suggest different bioavailability profiles, with LASSBio-1504 having a better pharmacokinetic profile than LASSBio-1506. This difference may be explained by the presence of the ethoxymorpholine subunit in the LASSBio-1504 derivative, which may improve the pharmacokinetic properties of this substance as it does for the p38 MAPK inhibitor BIRB-796 (Figure 1).

EXAMPLE 2.2 Effect of Compounds on TNF-α Production In Vivo

 It was also investigated how much LASSBio-1504 and LASSBio-1506 inhibit carrageenan-induced thermal hypernociception by inhibiting TNF-α production in vivo. Four hours after intraplantar injection of carrageenan, rat paws were homogenized and TNF-α in the supernatant was determined by ELISA. The results shown in Figure 2 reveal that four hours after carrageenan injection the TNF-α levels in the paw are higher than double compared to the paw in which only saline was injected. Previous oral treatment with LASSBio-1504 and LASSBio-1506 (100 μιηοΙ / kg) inhibited the elevation of TNF-tissue levels, corroborating the results in vitro. These results clearly demonstrate that the new phenylpyrazole A / -glycinyl- / V-acylhydrazonic derivatives LASSBio-1504 and LASSBio-1506 significantly inhibit TNF-α production in vitro and in vivo. EXAMPLE 2.3

Effect of LASSBio-1504 on Antigen Induced Arthritis (mBSA) - methylated bovine serum albumin

Due to the in vitro and in vivo pharmacological profile evidenced for the phenylpyrazolic A / -glycinyl-A / -acylhydrazonic derivative LASSBio-1504 in inhibiting TNF-α production and carrageenan-induced thermal hypernociception, this compound was selected for evaluation of its activity. in the antigen-induced arthritis (mBSA) model, a important model of chronic inflammation. The challenge with mBSA in the joint of previously immunized mice is an experimental model that has several features of human arthritis and involves the initial release of TNF-α which triggers subsequent release of IL-1β and chemokines.

 The results are illustrated in Figure 3 and show that thalidomide was not effective in mBSA-induced arthritis at the tested dose, although it was able to inhibit carrageenan-induced thermal hypernociception. Phenylpyrazole A / -glycinyl- / V-acylhydrazonic derivative LASSBio-1504 was effective from the fourth day of treatment with 100 μιηοΙ / kg orally (once daily treatment, one hour before mechanical sensitivity assessment). LASSBio-1504 inhibited mechanical hypernociception by 51% and 45% on the fourth and fifth day of treatment, respectively.

Claims

Claims

 1) Heterocyclic N-glycynyl-N-acylidrazone compounds, as well as their derivatives, isomers, salts and racemates, characterized in that they have the general formula (la), (Ib), (llia) and (llb).

2) Synthesis process of compounds of general formula

as their derivatives, their isomers, salts and racemates characterized by the following steps:

A) carbonyl condensation;

B) nucleophilic substitution to carbonyl; c) V-alkylation;

D) cyclocondensation;

E) multicomponent coupling reaction

3) Pharmaceutical composition characterized in that it has the compounds of general formula

 as well as their derivatives, isomers, salts and racemates.

4) Method of treating diseases and / or disorders characterized by the administration of compounds of general formula

as well as their derivatives, isomers, salts and racemates.