Low-molecular weight trp-p8 activity modulators

FIELD: chemistry.

SUBSTANCE: invention relates to a compound of formula I

or a pharmaceutically acceptable salt thereof, where R1 is H or R1 and R2 together with a nitrogen group can form where A, B, C and D are independently selected from a group consisting of CR1a and N; where at least one of A, B, C and D is CR1a; where R1a is selected from a group consisting of H, -ORi, -SRii, -S(O)Riii, -C(O)NRvRvi and CF3, where Ri is selected from a group consisting of methyl, ethyl, propyl, hydroxyethyl, hydroxypropyl, 2-oxo-2-phenylethyl, butyl, acetonitrile and benzyl; Rii, Riii and Riv denote methyl; Rv and Rvi are independently selected from a group consisting of H, methyl, ethyl, hydroxyethyl, hydroxypropyl, diethyalminoethyl, phenyl, pyridinyl, methoxyethyl, hydroxyethoxyethyl, benzyl, phenylethyl, 2-hydroxy-1-hydroxymethyl-2-phenylethyl and carbomoylethyl, or Rv and RVi together form morpholine or ethyl ester of piperazine; R2 is selected from a group consisting of phenyl, naphthyl, pyrazolyl and C1-C8alkylene phenyl; R3 is C1-C8alkylene; R4 is selected from a group consisting of H, C1-C8alkyl and -C=NH(NH2). The invention also relates to compounds of formulae I-A

I-B I-C

I-D I-E

values of radicals of which are given in the claim; a method of treating said pathological conditions, a pharmaceutical composition based on said compounds, a method of identifying a Trp-p8 agonist and specific compounds.

EFFECT: obtaining compounds which are useful as Trp-p8 modulators.

25 cl, 19 dwg, 8 tbl, 17 ex

 

Cross-reference to related application

This application claims the benefit of priority of provisional patent application U.S. No. 60/773435, registered February 15, 2006, the contents of which are incorporated herein by reference.

The level of technology

The present invention relates generally to the fields of cell biology, biochemistry and organic chemistry. More specifically, the present invention provides small molecule modulators of the activity of Trp-p8, which include agonists Trp-p8 and antagonists Trp-p8, as well as compositions containing small molecule modulators of Trp-p8. Also provides methods of identification and characterization of new low molecular weight modulators of Trp-p8, as well as methods of modulating mediamoo Trp-p8 influx of cations and/or apoptosis in the cell and related methods of treatment of diseases associated with expressionism, by activating and/or transfer of signals Trp-p8. Examples of diseases that can be treated using compositions and methods of the present invention, include cancers, such as cancers of the lung, breast, colon cancer and/or prostate.

The level of technology

Carcinoma of the prostate is the most common cancer diagnosed is in men in the United States, and gives the second largest proportion of deaths from cancer, second only to adenocarcinoma of the lung. Parker et al., CA Cancer J. Clin. 46:5-27 (1996). Although it is possible for the effective treatment of prostate cancer, limited in this body, there are very limited treatment options for metastatic disease. Thus, it is very important to find new ways to diagnose the disease at a very early stage and to closely monitor both the development and treatment of the disease and to develop new therapeutic approaches. To achieve this, it is important to understand the molecular mechanisms of prostate cancer and to identify new biochemical markers for diagnosis and disease progression.

To date, there is very little available prostate-specific markers. The most well-known and well-characterized markers shown diagnostic value for prostate cancer are proteins, acid phosphatase prostate (PAP), prostate-specific antigen (PSA) and prostate-specific membrane antigen (PSMA). Each of these proteins has also become a target for new immunotherapy approaches for the treatment of diseases. Horoszewicz et al., Anticancer Res. 7:927-935 (1987); Barren et al., Prostate 30:65-68 (1997); Murphy et al., Prostate 33:281-285 (1997); Murphy et al., Prostate 26:164-168 (1995); Rochon et al., Prostate 25:219-223 (1995); Correale et al., J. Immunol 161:3186-3194 (1998); and Murphy et al., Prostate 38:73-78(1999).

2+the trp family that is highly expressed in brain tissues. Nagamine et al., Genomics 54:124-131 (1998). Trp-p8 also shows significant homology with melastatin person, other protein related to the Trp family, expressed in melanocytes and which, as expected, the genome of the tumor suppressor. Duncan et al., Cancer Res. 58:1515-1520(1998) and Hunter et al., Genomics 54:116-123 (1998). Perhaps of most interest is the observation that the gene Trp-p8 apparently is expressed in a wide range of neoplastic lesions, outside the prostate, in addition to the prostate. Tsavaler et al., Cancer Res. 69(9):3760-9 (2001).

The Trp superfamily contains more than 20 related proteins cationic channels that are involved in processes including sensory function, sensory physiology to vasorelaxation and fertility of men. Defects in Trp channels associated with the ISM is the changes in the control of growth and suppression of tumors. Although all Trp proteins are calcium channels, they significantly differ in their selectivity and method of activation. Members of the Trp superfamily share significant homology sequence and predicted structural similarity, such as the size of the predicted transmembrane segments.

Trp-p8 sverkhekspressiya in a number of cancers, including prostate, breast, lungs and large intestine, whereas in normal tissues it is mainly expressed in the prostate [Tsavaler et al., see above] and ganglia dorsal root (DRG), (Dendreon, unpublished observation). Fuessel et al. reported that Trp-p8 is a highly prostate-specific and associated with carcinoma of the prostate gene, thus qualifying it as a potential target for specific therapy. International J. of Oncology 23:221-228 (2003). Among other species orthologues Trp-p8 clearly expressed in a subset of neurons in DRG and trigeminal ganglion (TG) rats [McKemy et al., Nature 416(6876}:52-8 (2002)]and in mice [Peier et al., Cell 108(5):705-15 (2002)]. Thus, Trp-p8 is a marker expressed by many tumors, with significant potential for use in diagnosing disease and monitoring of disease progression during treatment, as well as a viable target for cancer therapy.

Communication Trp-p8 with Raco the diversified diseases of the prostate, lung, breast and colon cancer and the important role of different ion channels, they play in the vital functions of cells, suggests that the channel Trp-p8 can have a significant function in signal transmission and/or proliferation of cancer cells. Modulation of activity of Trp-p8, either through activation with agonist or inhibition using antagonist, at physiological temperature can be valuable as a therapeutic agent for manipulating cells expressing Trp-p8, a specific way. See, for example, an application for U.S. patent No. 10/923413.

Accordingly, in this area there remains a need for low molecular weight modulators of the activity of Trp-p8, in compositions containing one or more low molecular weight modulators of Trp-p8, and the methods of identification and use of small molecules to modulate the activity of Trp-p8 cells for the treatment of diseases associated with aberrant expressionism Trp-p8.

The invention

The present invention satisfies these and other related needs by creating a low-molecular modulators of the activity of Trp-p8, including agonists Trp-p8 and antagonists Trp-p8, as well as compositions containing such modulators of Trp-p8, and methods of identification and use of modulators of Trp-p8. In certain embodiments Khujand the exercise of compounds of the present invention bind and activate Trp-p8, and/or stimulate the influx of cations, including, but not limited to, the influx of calcium into the cell, where the influx of cations correlates with toxicity induced by modulators of Trp-p8. Thus, in these and other embodiments, implementation agonists Trp-p8 of the present invention are effective in the inhibition of growth and/or inducing apoptosis and/or necrosis in cells expressing Trp-p8. In alternative embodiments, the implementation provides antagonists Trp-p8 that are effective in reducing the basal activity of Trp-p8 in the cell, thus reducing the survival rate of cells expressing Trp-p8. Mainly for this reason, agonists and antagonists of the present invention can be used to treat diseases including, but not limited to, cancers of the breast, lung, colon and/or prostate that are associated with expressionism Trp-p8.

One or more modulators of Trp-p8 can be prepared as compositions, including pharmaceutical compositions, containing one or more pharmaceutically acceptable carriers or excipients and/or one or more additional therapeutic compounds. Such compositions find use in methods of treating one or more diseases associated with expressionism Trp-p8.

Thus, in one embodiment, implemented the program the present invention provides small molecule modulators of Trp-p8 and their derivatives, where small molecules include compounds of the following formula I:

where R1selected from H, alkyl, heteroalkyl, arylalkyl and aryl, or R1and R2together with a group of nitrogen may form a cyclic or heterocyclic group, most of the 25 atoms;

R2is selected from aryl and arylalkyl;

R3selected from alkyl, heteroalkyl and arylalkyl;

R4selected from H, alkyl, heteroalkyl and arylalkyl; and

R3and R4together with a group of nitrogen form an aliphatic amine.

In related embodiments, the implementation of the present invention provides small molecule modulators of Trp-p8 and their derivatives, where small molecules include compounds of the following formula I-A:

where A, B, C and D are independently selected from CR2and N; where at least one of A, B, C and D represents CR2; where R2represents a residue selected from H, alkyl, heteroalkyl, aryl, halogen and arylalkyl, R6O - and R6S-, where R6represents alkyl; where, when two adjacent A, B, C and D represent CR2two R2can be combined to form a unified aryl, cycloalkyl or geteroseksualnoe group; and

R1selected from H, alkyl, heteroalkyl, aryl and aryl is Qila;

R3selected from alkyl, heteroalkyl, aryl, arylalkyl, -NR7C(O)-, -C(O)NR7-, -O-, -S-, -S(O)-, -S(O)2- , and-NR7-, where R7represents a residue selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R4selected from-C(O)R8-, alkyl, arylalkyl and heteroaryl, where R8selected from alkyl and heteroalkyl;

R5selected from H, alkyl, heteroalkyl and arylalkyl; and

R4and R5together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-A R1represents H; R7represents H; R8contains 2, 3 or 4 carbon atoms; R4choose from propionyl, ethyl, butyryl, hydroxypropionic and 3-hydroxybutyryl; R5selected from H and methyl; R6contains 1, 2, 3, 4, 5 or 6 carbon atoms; and/or R2selected from methoxy, methylsulfonyl, phenyl and H.

Here are illustrated the compounds of formula I-A, including a group selected from 2-(2-aminodiphenylamine)-4-methoxyphenyl, N-(2-amino-ethyl)-2-amino-5-methylsulfinylphenyl, 1-(2-aminoethoxy)naphthalene-2-yl, 2-(2-aminoethylamino)-4-methylsulfinylphenyl, N-(2-amino-ethyl)-5-methoxybenzamide, 2-(2-aminoethylamino)-4-methoxyphenyl, 2-(2-amino-3-hydroxypropylamino)-4-methoxyphenyl, 3-(2-aminoethylamino)naphthalene-2-yl, N-(2-amino-ethyl)-2-aminobenzamide, 2-(2-amino-3-guide is oxypropylene)-4-methoxyphenyl, 2-(2-aminoethylamino)phenyl,2-(2-amino-3-hydroxyethylamino)-4-methoxybenzylamine and 2-(2-aminoethylamino)-4-methoxyphenyl.

In an alternative related embodiments, the implementation of the present invention provides small molecule modulators of Trp-p8 and their derivatives, where small molecules include compounds of the following formula I-B:

where R1selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R2selected from aryl, alkyl, heteroalkyl and arylalkyl;

R3selected from alkyl, heteroalkyl and arylalkyl;

R4selected from H, alkyl, heteroalkyl and arylalkyl; and

R3and R4together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-B R1represents H; R3selected from methylene, ethylene, propylene and butylene; R4selected from H and methyl; and/or R2selected from phenyl, furan, methylpyrrole, methylbenzoate, AMINOPHENYL, hydroxyphenyl, cyanophenyl and methoxyphenyl.

Here are illustrated the compounds of formula I-B, including a group selected from 2-(2-amino-ethyl)-5-furan-2-yl-2H-pyrazole-3-yl, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(1-methyl-1H-pyrrol-2-yl)-2H-pyrazole-3-yl, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-am is noetic)-5-(4-AMINOPHENYL)-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(4-hydroxyphenyl)-2H-pyrazole-3-yl, 2-(2-methylaminomethyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(3-cyanophenyl)-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(3-methoxyphenyl)-2H-pyrazole-3-yl, methyl ester 4-{1-(2-amino-ethyl)-1H-pyrazole-3-yl}-benzoic acid, 2-(2-amino-ethyl)-5-(3-AMINOPHENYL)-2H-pyrazole-3-yl and 2-(2-amino-ethyl)-5-(3-hydroxyphenyl)-2H-pyrazole-3-Il.

In other related embodiments, the implementation of the present invention provides small molecule modulators of Trp-p8 and their derivatives, where small molecules include compounds of the following formula I-C:

where R1selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R2is selected from aryl and arylalkyl;

R3selected from alkyl, heteroalkyl, arylalkyl, -NHC(O)R5-, -OR5and other5-, where R5represents an alkyl or heteroalkyl;

R4selected from H, alkyl, heteroalkyl and arylalkyl; and

R3and R4together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-C R1represents H; R2represents phenyl; R5selected from methylene, ethylene, propylene and butylene; R3choose from propionamido, ethoxy, propoxy, ethylamino; and/or R4selected from H and methyl.

the here illustrated the compounds of formula I-C, including a group selected from 2-(2-aminodiphenylamine)-2-phenylethyl, 2-(2-aminoethoxy)-2-phenylethyl, 2-(2-aminoethoxy)-2-phenylethyl, 2-(3-aminopropoxy)-2-phenylethyl, 2-(2-dimethylaminoethoxy)-2-phenylethyl and 2-(2-aminoethylamino)-2-phenylethyl.

In other related embodiments, the implementation of the present invention provides small molecule modulators of Trp-p8 and their derivatives, where small molecules include compounds of the following formula I-D:

where R1selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R2is selected from aryl and arylalkyl;

R3selected from alkyl, heteroalkyl and arylalkyl;

R4selected from H, alkyl, heteroalkyl and arylalkyl; and

R3and R4together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-D R1represents H; R2selected from phenyl, phenylamino; R3selected from methylene, ethylene, propylene, butylene, methylamino, ethylamino, propylamino, butylamino and acetyl; and/or R4selected from H and methyl.

Here are illustrated the compounds of formula I-D, including a group selected from 2-[2-(2-aminoethylamino)phenyl]ethyl, 2-(2-aminomethylphenol)ethyl and 2-[(2-aminoacetyl)phenylamino]ethyl.

In other related embodiments, the implementation of the present izopet the tion provides small molecule modulators of Trp-p8 and their derivatives, where small molecules include compounds of the following formula I-E:

where A, B, C and D are independently selected from CR1and N; where at least one of A, B, C and D represents CR1; where R1selected from H, alkyl, heteroalkyl, aryl, arylalkyl and halogen; where, when two adjacent A, B, C and D represent CR1two R1can be combined to form a unified aryl, cycloalkyl or geteroseksualnoe group;

R2selected from alkyl, heteroalkyl and arylalkyl;

R3selected from H, alkyl, heteroalkyl and arylalkyl; and

R2and R3together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-E

(i) R1represents H or-ORiand Riselected from methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, acetonitrile, phenyl, phenylmethoxy, venlafaxi, phenylpropoxy, phenylmethoxy and benzyl;

(ii) R1represents-SRiiand Riiselected from methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, acetonitrile, phenyl, phenylmethoxy, venlafaxi, phenylpropoxy, phenylmethoxy and benzyl;

(iii) R1represents-S(O)Riiiand Riii

(iv) R1represents-S(O)2Rivand Rivselected from methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, acetonitrile, phenyl, phenylmethoxy, venlafaxi, phenylpropoxy, phenylmethoxy and benzyl;

(v) R1represents-C(O)NRvRviand Rvand Rviindependently selected from H, methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, diethylaminoethyl, phenyl, pyridinyl, methoxyethyl, hydroxyethoxyethyl, benzyl, methylphenyl, phenylethyl, hydroxyhydrocinnamate, carbamoylmethyl and hydroxymethyl of hydroxyethyl;

(vi) R1represents-C(O)NRvRviand Rvand Rvitogether form a morpholine, piperazine, complex ethyl ester piperazine;

(vii) R2selected from methylene, ethylene, propylene and butylene;

(viii) R2represents ethylene, and R3represents H; and

(ix) R1is a CF3or halogen.

Here are illustrated the compounds of formula I-E, including a group selected from 3-(2-amino-ethyl)-5-methods the si-1,3-dehydrobenzperidol-2-it, 3-(2-amino-ethyl)-5-(3-hydroxypropoxy)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-ethoxy-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-methanesulfonyl-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-(2-hydroxyethoxy)-1,3-dehydrobenzperidol-2-it, amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-5-methylsulfanyl-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-methanesulfonyl-1,3-dehydrobenzperidol-2-it, (2-diethylaminoethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-aminopropyl)-2,3-dehydrobenzperidol-2-it, [3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-yloxy]acetonitrile, ethylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, pyridine-3-ylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-methoxyethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 1-(2-amino-ethyl)-1,3-dehydrobenzperidol-2-it, 1-(2-amino-ethyl)-1,3-dihydronaphtho[2,3-d]imidazol-2-it, (2-hydroxyethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-5-propoxy-1,3-dehydrobenzperidol-2-it, 3-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-c]pyridine-2-it, (2-diethylaminoethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-4-carboxylic acid, pyridine-4-ylamide 3-(2-amino-ethyl)-2-oxo-2,3-di is Idro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-b]pyridine-2-it, 1-(3-aminopropyl)-1,3-dehydrobenzperidol-2-it, phenylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, [2-(2-hydroxyethoxy)ethyl]amide of 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 1-(2-amino-ethyl)-5-trifluoromethyl-1,3-dehydrobenzperidol-2-it, 1-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-c]pyridine-2-it, benzylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-5-(morpholine-4-carbonyl)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-(2-oxo-2-phenylethane)-1,3-dehydrobenzperidol-2-it 3-(2-methylaminomethyl)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-butoxy-1,3-dehydrobenzperidol-2-it, methylphenylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, ethyl ester 4-[3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carbonyl]piperazine-1-carboxylic acid diethylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, phenetylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-hydroxy-1-hydroxymethyl-2-phenylethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, carbamoylmethyl 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-hydroxy-1-hydroxymethylation)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihyd is about-1H-benzoimidazol-5-carboxylic acid, N-{2-[2-oxo-2,3-dihydrobenzoic-1-yl]ethyl}guanidine, 3-(2-amino-ethyl)-5-benzyloxy-1,3-dehydrobenzperidol-2-it 1-(4-aminobutyl)-1,3-dehydrobenzperidol-2-it. In one of these embodiments provides for the connection of 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-5-methoxy-1,3-dehydrobenzperidol-2-it.

Other aspects of the present invention provide compositions, including pharmaceutical compositions, containing one or more low molecular weight modulators of Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E in combination with a pharmaceutically acceptable excipient, carrier and/or diluent. Here the examples are specific agonists and antagonists Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E; methods of synthesis of exemplary agonists and antagonists Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E; and data EC50demonstrate efficiency in vitro and specific activity of each of these agonists and antagonists Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E.

In other aspects, the compositions of the present invention contain one or more compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E, prepared together with one who does multiple therapeutic agents against cancer. Alternative compositions of the present invention contain a compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E, independently prepared, together with one or more therapeutic agents against cancer. That is, one or more compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E and therapeutic agent against cancer, are prepared separately.

Appropriate therapeutic agents against cancer include, but are not limited to, antimitoticescoe agents, including, but not limited to, paclitaxel, vincristine and etoposide; alkylating agents, including, but not limited to, mechlorethamine, cyclophosphamide, carmustine; antimetabolites, including, but not limited to, methotrexate, gemcitabine, lometrexol, 5-fluorouracil and 6-mercaptopurine; cytotoxic antibiotics, including, but not limited to, doxorubicin, daunorubicin, bleomycin, mitomycin C and streptozocin; agents, platinum-based, including but not limited to this, cisplatin and carboplatin; hormonal agents, including, but not limited to, antiestrogens, such as tamoxifen and diethylstilbestrol and antiandrogens, such as flutamide; agents against angiogenesis and inhibitors farnesyltransferase.

In certain aspects of compounds of formula I, f is rmula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E are entered in combination with therapeutic agents against cancer, which themselves are ineffective in modulating the activity of Trp-p8 cells expressing Trp-p8. Unexpectedly, these types of combination therapy leads to improved efficiency compared to using only one of the compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E.

In other aspects, the compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E are entered in combination with one or more additional modulators (modulator) Trp-p8, including, but not limited to, compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E.

In some of these embodiments provides a low-molecular low molecular weight antagonists agonists Trp-p8 presented here. Thus, in certain embodiments of the implementation of the envisaged low-molecular-weight antagonists of Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E and their derivatives for one or more agonists Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E.

Additional embodiments of the present invention provide methods of reducing survive the t cells and/or inhibiting cell growth, methods of stimulating the flow of cations and methods of inducing apoptosis and/or necrosis in cells expressing Trp-p8. Examples of such methods include the stage of contacting cells with a compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E at a concentration and for a time necessary to reduce the survival rate of cells and/or inhibiting growth of cells to increase intracellular calcium levels and/or to induce apoptosis and/or necrosis of the cells.

In other embodiments, implementation of the present invention provides methods of treating diseases in mammals, most people, through the introduction of one or more compounds and/or compositions of the present invention. In certain aspects, the methods include the introduction of a composition containing a combination of compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E with one or more therapeutic agents against cancer, delivered by way with the simultaneous introduction of, for example, as a single drug. In some other aspects of the methods of the present invention include combination therapy, where the compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E is introduced first in one drug, and then therapeutic agent against the cancer in the individual drug. The methods also include a method, when the first therapeutic agent against cancer is delivered in one drug, and then the compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E in a separate drug.

therapeutic methods of the present invention are particularly effective in the treatment of cancer associated with the expression of Trp-p8, including, but not limited to, certain cancers of the colon, lung, breast and prostate.

The above and additional features of the present invention and the manner of obtaining them will become apparent, and the present invention will be better understood by reference to the following detailed description, taken in conjunction with the accompanying figures.

Brief description of figures

Figa-1B are graphs depicting the approximate survival analysis using ATP, suitable for the study and characterization of low molecular weight modulators of Trp-p8 of the present invention. In the preliminary analysis (tiga) compounds tested at 1 μm and a specific loss of CHO cells expressing Trp-p8 (CHO/Trp-p8), measured at 37°C. In the following analysis (pigv) compounds tested at various concentrations and the death of CHO cells expressing Trp-p8 (CHO/Trp-p8), measured at 37°C. the Value EC50get the C of the graph of the survival rate of cells as a function of concentration.

Figa-2C are graphs depicting induced modulators of Trp-p8 increase in the level of intracellular calcium, as determined using flow analysis of calcium carried out at 37°C. Figa is a positive control showing that CHO cells and CHO/Trp-p8 react in a similar manner to 2 µm ionomycin at 37°C, in the analysis of the flow of calcium. Figv represents the negative control, demonstrating that parentally CHO cells that do not Express endogenous or exogenous Trp-p8, do not respond to agonists Trp-p8 at a concentration of 10 μm. Pigs shows that agonist Trp-p8 induces a specific, concentration-dependent response in the cells of CHO/Trp-p8 at 37°C.

Figure 3 represents the data for a graph of flow cytometry showing that agonist Trp-p8 is able to induce specific apoptosis in expressing Trp-p8 CHO cells at 37°C dependent on dose.

Figure 4 is a graph showing exemplary results of the initial screening of antagonists of Trp-p8 using survival analysis using ATP as described here, cells CHO/Trp-p8 at 37°C. Cells CHO/Trp-p8 exhibit for compounds at different concentrations in 1% DMSO or 1% DMSO in combination with toxic concentration of agonist Trp-p8. Cell survival was measured after 24-26 hours at 37° C using ATP analysis. Compounds that protect cells from the toxic effect of agonist Trp-p8, are classified as antagonists Trp-p8 (connection A-B). Inactive compound (compound C) has no protective effects shown here are for illustration purpose analysis.

Figure 5 is a graph depicting the screening and characterization of antagonists of Trp-p8 through flow analysis of calcium at 37°C. Cells CHO/Trp-p8 load indicator dye for calcium Fura-2, and increased levels of intracellular calcium in response to the connection determined by the increase in fluorescence. Cells CHO/Trp-p8, loaded with dye Fura-2, exhibit 1% DMSO or antagonist, at different concentrations in 1% DMSO at 37°C. After three minutes to cells add agonist. When cells are exposed to effective concentrations of the antagonist, their ability to respond to agonist significantly reduced or completely disappears.

Figa-6B are graphs depicting the results for the sample of animal models suitable for the study and characterize small molecule modulators of Trp-p8 of the present invention. Mice inject subcutaneously with cells CHO/Trp-p8, which leads to the formation of solid tumors. Length (longest dimension) and width (dimension perpendicular thereto and in the same plane as the length) of each tumor are measured using a caliper gauge, and tumor volume are approximated using the formula for volume of an ellipsoid: 0,52*L*W2. When the average tumor volume reaches approximately 100 mm3mice are divided into groups randomly. On figa mouse injected a single dose or the compounds as drug water-based or carrier itself with oral gastric tube. On FIGU mice dosed several times or investigational compound as the drug is water-based, or the media itself with oral gastric tube. Then tumors consistently measured in these days. Data represent mean tumor volumes ± standard error.

7 is a graph depicting the plasma concentration of several compounds, as a function of time and dose for the mouse after administration of a single intraperitoneal injection. All compounds are dissolved in the product is water-based and administered at comparable levels of dosing. Blood is collected at the indicated time points and analyzed for the levels of drugs.

Figa-8B are graphs depicting the efficiency demonstrated by several compounds in mice with transplanted tumors expressing Trp-p8. Mice inject subcutaneously cells CHO/Trp-p8, resulting in the formation of solid tumors. Length is (L; the largest size) and width (W; dimension perpendicular to the length and located in the same plane) of each tumor was measured using a caliper gauge and tumor volume are approximated using the formula for volume of an ellipsoid: 0,52*L*W2. When the average tumor volume reaches approximately 100 mm3mouse randomly divided into groups and administered either compound as the drug is water-based, or the media itself via intraperitoneal injection in these days. All connections are introduced at comparable levels of dosing. Tumors consistently measured in these days. Data are presented as mean tumor volumes ± standard error.

Figa and 9B are graphs depicting the plasma concentration of several compounds, as a function of time and dose, in rats (figa) and dogs (pigv) after a single oral dose. All compounds are dissolved in the product is water-based and administered at a comparable dosage levels with oral gastric tube. Blood is collected at the indicated time points and analyzed for the levels of drugs.

Figa and 10B are graphs depicting the efficiency demonstrated multiple connections on the model with transplanted tumors in mice expressing Trp-p8. Mice make subcutaneous injection of cells CHO/Trp-p8 that p is igodit to the formation of solid tumors. The length (L; maximum size) and width (W; dimension perpendicular to the length and located in the same plane) of each tumor was measured using a caliper gauge and tumor volume are approximated using the formula for volume of an ellipsoid: 0,52*L*W2. When the average tumor volume reaches approximately 100 mm3mouse randomly divided into groups and administered either a single dose of a compound as a drug is water-based, or the media itself with oral gastric tube. Then tumors consistently measured in these days. Data represent mean tumor volumes ± standard error.

11 is a graph depicting the lack of efficacy demonstrated exemplary connection of the model with transplanted tumors in mice that lack the expression of Trp-p8. Mice make subcutaneous injection of cells CHO-K1, where there is no Trp-p8, which leads to the formation of solid tumors. The length (L; maximum size) and width (W; dimension perpendicular to the length and located in the same plane) of each tumor was measured using a caliper gauge, and tumor volume are approximated using the formula for volume of an ellipsoid: 0,52*L*W2. When the average tumor volume reaches approximately 50 mm3mouse randomly divided into groups and administered either a single dose of a compound as p is of Ephrata water-based, any media using oral gastric tube. Then tumors consistently measured in these days. Data presented as mean tumor volume ± standard error.

Fig is a graph depicting the efficacy demonstrated exemplary connection of the model with transplanted tumors in mice expressing Trp-p8. The tumor model LuCaP receive from Robert L. Vassella, Ph.D., Professor of Urology in the University of Washington's School of Medicine. The length (L; maximum size) and width (W; dimension perpendicular to the length and located in the same plane) of each tumor was measured using a caliper gauge and tumor volume are approximated using the formula for volume of an ellipsoid: 0,52*L*W2. When the average tumor volume reaches approximately 150 mm3the mice injected with the compound as a drug is water-based with intraperitoneal injection daily for 5 days. Then tumors consistently measured in these days. Data presented as mean tumor volume ± standard error.

SEQ ID NO: 1 is the nucleotide sequence of cDNA Trp-p8 human (GenBank Accession No. AY090109).

SEQ ID NO: 2 represents the amino acid sequence encoded by nucleotide sequence SEQ ID NO: 1 (GenBank Accession No. NP_076985).

Detailed description of the invention

The present invention is osnovyvaet is camping on the detected certain small molecule modulators of Trp-p8, including the agonist activity of Trp-p8, is able to inhibit the growth and/or induce apoptosis and/or necrosis in cells that Express Trp-p8. Having no desire to be limited to any particular mode of action, it is assumed that mediaremote agonist Trp-p8 activating receptor Trp-p8 significantly increases the influx of cations, which correlates with cellular toxicity. In addition, it is expected that antagonists of Trp-p8 can inhibit the basal level of activity and/or induced native ligands activity of endogenous Trp-p8, activation of which leads to reduced growth or death of cells expressing this protein cation channel.

Thus, the present invention provides small molecule modulators of Trp-p8, including agonists and antagonists of the activity of Trp-p8, as well as compositions, including pharmaceutical compositions, containing one or more low molecular weight modulators of Trp-p8 in combination with one or more pharmaceutically acceptable carriers and/or excipients. The present invention also provides a combined composition comprising one or more modulators of Trp-p8 and one or more additional therapeutic compounds, such as, for example, therapeutically the agent against cancer. Modulators of Trp-p8 and compositions containing modulators of Trp-p8, will find use in the methods of activation mediamoo Trp-p8 influx of cations into the cell, methods of inducing apoptosis and/or necrosis in a cell, and methods of treatment of diseases associated with expression of Trp-p8, including, but not limited to, cancers such as cancers of the breast, colon, lung and prostate.

Definition

The term "modulators of Trp-p8" refers generally to compounds which are low molecular weight agonists and antagonists that are associated with them and either increase or decrease, respectively, the activity of Trp-p8 in the cell. Agonists Trp-p8 include the compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E and their chemical derivatives. Antagonists Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E can easily be synthesized and characterised by specialists in this field through the use of the methodology clearly listed here, and/or such methodology, which is easily available in this area.

The phrase "activating Trp-p8" means mediaremote agonist activating the expression of Trp-p8 on the cell surface. For example, in certain embodiments of the implementation of the agonists of the present invention, when are contacted with the cell and/or injected in vivo to a subject to a mammal, activate Trp-p8, thereby facilitating the influx of cations, such as calcium ions, to such intracellular levels and/or until such duration that are sufficient to cause toxicity to cells, as indicated by the decrease in cell growth and/or the occurrence of necrotic and/or apoptotic cell death.

The term "aliphatic amine" means a substituted nitrogen atom, where any Deputy, other than H, is attached to the nitrogen through a saturated carbon atom.

The term "alkyl", by itself or as part of another substituent, means, unless approved otherwise by having a straight or branched chain or cyclic hydrocarbon group, or a combination thereof, which may be fully saturated, mono - or polyunsaturated and can include di - and multivalent group having the number of carbon atoms designated (i.e. C1-C10means one to ten carbon atoms). Examples of saturated hydrocarbon groups include groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, cyclohexyl, (cyclohexyl)ethyl, cyclopropylmethyl, homologs and isomers of, for example, n-pentyl, n-hexyl, n-heptyl, n-octyl and the like. Unsaturated alkyl group represents a group having one or more double bonds and is, and triple bonds. Examples of unsaturated alkyl groups include vinyl, 2-propenyl, crotyl, 2-isopentenyl, 2-(butadienyl), 2,4-pentadienyl, 3-(1,4-pentadienyl), ethinyl, 1 - and 3-PROPYNYL, 3-butynyl and higher homologs and isomers.

The term "alkenyl" denotes branched or unbranched hydrocarbon chain containing one or more double bonds in the carbon-carbon bonds.

The term "quinil" refers to a branched or unbranched hydrocarbon chain containing one or more triple relations of the carbon-carbon bonds.

The term "alkylene", by itself or as part of another substituent means a divalent group derived from an alkane, as illustrated by --CH2CH2CH2CH2--. As a rule, Allenova group will have from 1 to 24 carbon atoms, with the group having 10 or less carbon atoms are preferred in the present invention. "Lower alkyl" or "lower alkylene" represents having a shorter chain alkyl or alkilinity group, generally having eight or fewer carbon atoms.

The term "cycloalkyl", by itself or as part of another substituent means a divalent group derived from cycloalkane, as illustrated by cycloheximide. As a rule, cycloalkenes group will have from 5 to 8 carbon atoms, this group is, having 6 carbon atoms are preferred in the present invention.

The term "albaniles", by itself or as part of another substituent means a divalent group derived from alkenyl, as illustrated by-CH=CHCH2CH2-. As a rule, alkenylamine group will have from 2 to 24 carbon atoms, with the group having 10 or less carbon atoms are preferred in the present invention.

The terms "alkoxy", "alkylamino and alkylthio" refer to groups having an alkyl group attached to the rest of the molecule via an oxygen atom, nitrogen or sulfur, respectively. Similarly, the term "dialkylamino" is used in the ordinary sense to mention-R NR'r", where the groups R may be the same or different alkyl groups.

The term "heteroalkyl", by itself or in combination with another term, means, unless approved otherwise by having a stable straight or branched chain, or cyclic hydrocarbon, or combinations thereof, fully saturated or containing from 1 to 3 States of unsaturation, consisting of the specified number of carbon atoms and from one to three heteroatoms selected from the group consisting of O, N, Si and S, and the atoms of nitrogen and sulfur may optionally be oxidized, and the nitrogen heteroatom may optionally be kV is termizirovanny. Heteroatom (heteroatoms O, N and S may be placed at any interior position heteroalkyl group. Heteroatom Si may be placed at any position heteroalkyl groups, including the situation in which an alkyl group is attached to the remainder of the molecule. Examples include --CH2--CH2--O--CH3, --CH2--CH2--NH--CH3,--CH2--CH2-N(CH3-)--CH3, -CH2--S--CH2--CH3, --CH2--CH2--S(O)--CH3, --CH2--CH2--S(O)2--CH3, --CH=CH--O--CH3, --Si(CH3)3,--CH2-CH=N--OCH3- and-CH=CH--N(CH3)--CH3. At most two heteroatoms may be sequential, such as --CH2--NH--OCH3and --CH2--O--Si(CH3)3. Also included in the term "heteroalkyl" are those groups which are described in more detail below as "heteroseksualci". The term "heteroalkyl", by itself or as part of another substituent means a divalent group derived from heteroalkyl, as illustrated by --CH2--CH2--S--CH2CH2-- and --CH2--S--CH2CH2--NH--CH2--. For heteroarenes groups, heteroatoms can also occupy either or both end of the chain. In addition, for alkilinity and heteroarenes bridging groups implies the raised orientation bridging groups.

The term "acyl" refers to such groups derived from an organic acid by removal of the hydroxy portion of the acid. Accordingly, acyl, as implied, including, for example, acetyl, propionyl, butyryl, decanoyl, pivaloyl, benzoyl and the like.

"Activated carbonyl group" is a carbonyl group, whose ELECTROPILOT increases because of the groups attached to either side of the carbonyl. Examples of such activated carbonyl groups are (polyfluoroankyl)ketones, (polyfluoroankyl)aldehydes, complex alpha-keto esters, alpha-keto acids, alpha-keto amides, 1,2-diketones, 2-acility, 2-illimitably and the like.

The terms "cycloalkyl and heteroseksualci", by themselves or in combination with other terms, represent, unless approved otherwise, cyclic versions of "alkyl" and "heteroalkyl" respectively. In addition to geterotsiklicheskie heteroatom can occupy the position in which the heterocycle is attached to the remainder of the molecule. Examples of cycloalkyl include cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl and the like. Examples of geterotsiklicheskie include 1-(1,2,5,6-tetrahydropyridine), 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, 4-morpholinyl, 3-morpholinyl, tetrahydrofur the h-2-yl, tetrahydrofuran-3-yl, tetrahydrothieno-2-yl, tetrahydrothieno-3-yl, 1-piperazinil, 2-piperazinil and the like.

The terms "halo" or "halogen", alone or as part of another substituent, mean, unless approved otherwise, the atom of fluorine, chlorine, bromine or iodine. In addition, terms such as "foralkyl"is supposed to include monitorall and polyfluoroankyl.

The term "aryl"used alone or in combination with other terms (e.g., aryloxy, arylthiols, arylalkyl), means, unless approved otherwise, aromatic Deputy, which may be a single ring or multiple rings (up to three rings)which are fused together or linked covalently. The term "heteroaryl", is meant to include aryl rings that contain from zero to four heteroatoms, selected from N, O and S, where the nitrogen atoms and sulfur is optionally oxidized, and the atom (atoms) nitrogen is optional and stereoselectivity. "Heteroaryl" group can join the rest of the molecule through a heteroatom. Non-limiting examples of aryl and heteroaryl groups include phenyl, 1-naphthyl, 2-naphthyl, 4-biphenyl, 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, 3-pyrazolyl, 2-imidazolyl, 4-imidazolyl, pyrazinyl, 2-oxazolyl, 4-oxazolyl, 2-phenyl-4-oxazolyl, 5-oxazole is, 3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, 2-furyl, 3-furyl, 2-thienyl, 3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidyl, 4 pirimidil, 5-benzothiazolyl, purines, 2-benzimidazolyl, 5-indolyl, 1-ethanolic, 5-ethanolic, 2-honokalani, 5-honokalani, 3-chinolin and 6-chinolin. Substituents for each of the above aryl ring systems are selected from the group of acceptable substituents described below. The term "arylalkyl", is meant to include such groups in which the aryl or heteroaryl group attached to the alkyl group (e.g. benzyl, fenetre, pyridylmethyl and the like) or heteroalkyl group (for example, phenoxymethyl, 2-pyridylacetic, 3-(1-naphthyloxy)propylene and the like).

Each of the above terms (e.g., "alkyl," "heteroalkyl" and "aryl", as expected, includes both substituted and unsubstituted forms of the indicated groups. Preferred substituents for each type of groups are provided below.

Substituents for the alkyl and heteroalkyl groups (including those groups often referred to as alkylene, alkenyl, heteroalkyl, heteroalkyl, quinil, cycloalkyl, heteroseksualci, cycloalkenyl and geteroseksualen) can be a variety of groups selected from: --OR', =O, =NR', =N-OR', --NR'r R is the, -SR', -halogen, -SiR'R"R"', --OC(O)R', --C(O)R', --CO2R', CONR'R”, --OC(O)R NR'r”, --NR”C(O)R', --NR'--C(O)NR"R"', --NR”C(O)2R', --NH--C(NH2)=NH, --NR'r C(NH2)=NH, --NH--C(NH2)=NR', --S(O)R', --S(O)2R', --S(O)2NR'r R', --CN and --NO2in an amount in the range of zero to (2N+1), where N represents the total number of carbon atoms in such a group. R', R” and R'"each independently refer to hydrogen, unsubstituted(C1-C8)alkyl and heteroalkyl, the unsubstituted aryl, aryl substituted by 1-3 halogen atoms, the unsubstituted alkyl, alkoxy or dialkoxy groups, or aryl-(C1-C4)alkyl groups. When R' and R” are attached to the same nitrogen atom they may together with the nitrogen atom, with the formation of 5-, 6 - or 7-membered ring. For example, --NR'r R”, as implied, includes 1-pyrrolidinyl and 4-morpholinyl. From the above discussion of substituents specialists in this field will be clear that the term "alkyl", as implied, includes groups such as halogenated (for example, --CF3and --CH2CF3) and acyl (e.g., --C(O)CH3, --C(O)CF3, --C(O)CH2OCH3and the like).

Similarly, substituents for the aryl groups vary, and are selected from: -halogen, --OR', --OC(O)R', --NR'r R”, --SR', --R', --CN, --NO2, --CO2R', --CONR'R”, -C(O)R', --OC(O)R NR'r”, --NR”C(O)R', --NR”C(O)2R', --NR'-C(O)NR”R"', --NH--C(NH 2)=NH, --NR'r C(NH2)=NH, --NH--C(NH2)=NR', --S(O)R', --S(O)2R', --S(O)2NR'r R”, --NR--S(O)2-R', --N3, -CH(Ph)2, PERFLUORO(C1-C4)alkoxy, and PERFLUORO(C1-C4)alkyl, in an amount in the range from zero to the total number of open valences on aromatic ring system; and R', R” and R'" are independently selected from hydrogen, (C1-C8)alkyl and heteroalkyl, unsubstituted aryl, (unsubstituted aryl)-(C1-C4)alkyl, and (unsubstituted aryl)oxy-(C1-C4)alkyl.

Two of the substituents on adjacent atoms of the aryl ring may optionally be replaced by a Deputy of the formula --T--C(O)-(CH2)q-U--, where T and U independently represent-NH--, --O--, --CH2-- or single bond, and the subscript q is an integer from 0 to 2. Alternative two of the substituents on adjacent atoms of the aryl ring may optionally be replaced by a Deputy of the formula --A--(CH2)r--B--, where A and B independently represent --CH2--, --O--, --NH--, --S--, --S(O)--, --S(O)2--, -S(O)2NR'- or a single bond and r is an integer from 1 to 3. One of the single links of the new ring thus formed may optionally be replaced by a double bond. Alternative two of the substituents on adjacent atoms of the aryl ring can optional the part to be replaced by the Deputy of the formula --(CH 2)s--X--(CH2)t--, where s and t independently represent integers from 0 to 3 and X represents --O--, --NR'--, --S--, --S(O)--, --S(O)2or --S(O)2NR'--. The substituent R' in-NR'--, and --S(O)2NR'-- is selected from hydrogen or unsubstituted (C1-C6)alkyl.

As used here, the term "heteroatom"as implied, includes oxygen (O), nitrogen (N) and sulfur (S).

The term "pharmaceutically acceptable salt"is meant to include salts of the active compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E , which receive relatively nontoxic acids or bases, depending on the particular substituents found on the compounds described herein. Examples of pharmaceutically acceptable basic additive salts include, but are not limited to, sodium, potassium, calcium, ammonium, organic amino, or magnesium other similar salts. Examples of pharmaceutically acceptable acid additive salts include, but are not limited to, salts derived from inorganic acids like hydrochloric, Hydrobromic, nitric, carbonic acid, disubstituted salts of carbonic acid, salts of acids, like phosphoric acid, disubstituted salts of phosphoric acid, salts of acids, such dihydrophosphate, sulfuric acid, is aerovodochody, itestosterone or phosphorous acids and the like, as well as the salts derived from relatively nontoxic organic acids like acetic, propionic, isoalkanes, oxalic, maleic, malonic, benzoic, succinic, cork, fumaric, almond, phthalic, benzosulfimide, p-toluensulfonate, citric, tartaric, methanesulfonic and the like.

Small molecule modulators of the activity of Trp-p8

Small molecule modulators of Trp-p8, which are suitable for use in the compositions and methods of the present invention, are illustrated here by means of the compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E, described here, and their derivatives.

Thus, in one of the embodiments the present invention provides small molecule modulators of Trp-p8 and their derivatives, where low-molecular compounds include compounds of the following formula I:

where R1selected from H, alkyl, heteroalkyl, arylalkyl and aryl, or R1and R2together with a group of nitrogen may form a cyclic or heterocyclic group, most of the 25 atoms;

R2is selected from aryl and arylalkyl;

R3selected from alkyl, heteroalkyl and arylalkyl;

R4choose the from H, of alkyl, heteroalkyl and arylalkyl; and

R3and R4together with a group of nitrogen form an aliphatic amine.

In related embodiments, the implementation of the present invention provides small molecule modulators of Trp-p8 and their derivatives, where low-molecular compounds include compounds of the following formula I-A:

where A, B, C and D are independently selected from CR2and N; where at least one of A, B, C and D represents CR2; where R2represents a residue selected from H, alkyl, heteroalkyl, aryl, halogen, arylalkyl, R6O - and R6S-, where R6represents alkyl; thus, when two adjacent A, B, C and D represent CR2two R2can be combined to form a unified aryl, cycloalkyl or geteroseksualnoe group; and

R1selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R3selected from alkyl, heteroalkyl, aryl, arylalkyl, -NR7C(O)-, -C(O)NR7-, -O-, -S-, -S(O)-, -S(O)2- , and-NR7-, where R7represents a residue selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R4selected from-C(O)R8-, alkyl, arylalkyl and heteroaryl, where R8selected from alkyl and heteroalkyl;

R5selected from H, alkyl, heteroalkyl and arylalkyl; and

R 4and R5together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-A R1represents H; R7represents H; R8contains 2, 3 or 4 carbon atoms; R4choose from propionyl, ethyl, butyryl, hydroxypropionic and 3-hydroxybutyryl; R5selected from H and methyl; R6contains 1, 2, 3, 4, 5 or 6 carbon atoms; and/or R2selected from methoxy, methylsulfonyl, phenyl and H.

Here are illustrated the compounds of formula I-A, containing a group selected from 2-(2-aminodiphenylamine)-4-methoxyphenyl, N-(2-amino-ethyl)-2-amino-5-methylsulfinylphenyl, 1-(2-aminoethoxy)naphthalene-2-yl, 2-(2-aminoethylamino)-4-methylsulfinylphenyl, N-(2-amino-ethyl)-5-methoxybenzamide, 2-(2-aminoethylamino)-4-methoxyphenyl, 2-(2-amino-3-hydroxypropylamino)-4-methoxyphenyl, 3-(2-aminoethylamino)naphthalene-2-yl, N-(2-amino-ethyl)-2-aminobenzamide, 2-(2-amino-3-hydroxypropylamino)-4-methoxyphenyl, 2-(2-aminoethylamino)phenyl, 2-(2-amino-3-hydroxyethylamino)-4-methoxybenzylamine and 2-(2-aminoethylamino)-4-methoxyphenyl.

In an alternative related embodiments, the implementation of the present invention provides small molecule modulators of Trp-p8 and their derivatives, where low-molecular compounds include compounds of the following formula I-B:

where R1selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R2selected from aryl, alkyl, heteroalkyl and arylalkyl;

R3selected from alkyl, heteroalkyl and arylalkyl;

R4selected from H, alkyl, heteroalkyl and arylalkyl; and

R3and R4together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-B R1represents H; R3selected from methylene, ethylene, propylene and butylene; R4selected from H and methyl; and/or R2selected from phenyl, furan, methylpyrrole, methylbenzoate, AMINOPHENYL, hydroxyphenyl, cyanophenyl and methoxyphenyl.

Here are illustrated the compounds of formula 1-B, containing a group selected from 2-(2-amino-ethyl)-5-furan-2-yl-2H-pyrazole-3-yl, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(1-methyl-1H-pyrrol-2-yl)-2H-pyrazole-3-yl, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(4-AMINOPHENYL)-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(4-hydroxyphenyl)-2H-pyrazole-3-yl, 2-(2-methylaminomethyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(3-cyanophenyl)-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(3-methoxyphenyl)-2H-pyrazole-3-yl, methyl ester 4-{1-(2-amino-ethyl)-1H-pyrazole-3-yl}-benzoic acid, 2-(2-amino-ethyl)-5-(3-AMINOPHENYL)-2H-pyrazole-3-yl and 2-(2-aminoet is)-5-(3-hydroxyphenyl)-2H-pyrazole-3-Il.

In other related embodiments, the implementation of the present invention provides small molecule modulators of Trp-p8 and their derivatives, where low-molecular compounds include compounds of the following formula I-C:

where R1selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R2is selected from aryl and arylalkyl;

R3selected from alkyl, heteroalkyl, arylalkyl, -NHC(O)R5-, -OR5and other5-, where R5represents an alkyl or heteroalkyl;

R4selected from H, alkyl, heteroalkyl and arylalkyl; and

R3and R4together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-C R1represents H; R2represents phenyl; R5selected from methylene, ethylene, propylene and butylene; R3choose from propionamido, ethoxy, propoxy, ethylamino; and/or R4selected from H and methyl.

Here are illustrated the compounds of formula I-C, containing a group selected from 2-(2-aminodiphenylamine)-2-phenylethyl, 2-(2-aminoethoxy)-2-phenylethyl, 2-(2-aminoethoxy)-2-phenylethyl, 2-(3-aminopropoxy)-2-phenylethyl, 2-(2-dimethylaminoethoxy)-2-phenylethyl and 2-(2-aminoethylamino)-2-phenylethyl.

In other related embodiments, the implementation of the present invention predusmatriva is no small molecule modulators of Trp-p8 and their derivatives, where low-molecular compounds include compounds of the following formula I-D:

where R1selected from H, alkyl, heteroalkyl, aryl and arylalkyl;

R2is selected from aryl and arylalkyl;

R3selected from alkyl, heteroalkyl and arylalkyl;

R4selected from H, alkyl, heteroalkyl and arylalkyl; and

R3and R4together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-D R1represents H; R2selected from phenyl, phenylamino; R3selected from methylene, ethylene, propylene, butylene, methylamino, ethylamino, propylamino, butylamino and acetyl; and/or R4selected from H and methyl.

Here are illustrated the compounds of formula I-D containing group selected from 2-[2-(2-aminoethylamino)phenyl]ethyl, 2-(2-aminomethylphenol)ethyl and 2-[(2-aminoacetyl)phenylamino]ethyl.

In other related embodiments, the implementation of the present invention provides small molecule modulators of Trp-p8 and their derivatives, where low-molecular compounds include compounds of the following formula I-E:

where A, B, C and D are independently selected from CR1and N; where at least one of A, B, C and D represents CR1; where R1selected from H, alkyl, heteroalkyl, ar is La, arylalkyl and halogen; when two adjacent A, B, C and D represent CR1two R1can be combined to form a unified aryl, cycloalkyl or geteroseksualnoe group;

R2selected from alkyl, heteroalkyl and arylalkyl;

R3selected from H, alkyl, heteroalkyl and arylalkyl; and

R2and R3together with a group of nitrogen form an aliphatic amine.

In certain exemplary compounds of formula I-E

(i) R1represents H or-ORiand Riselected from methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, acetonitrile, phenyl, phenylmethoxy, venlafaxi, phenylpropoxy, phenylmethoxy and benzyl;

(ii) R1represents-SRiiand Riiselected from methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, acetonitrile, phenyl, phenylmethoxy, venlafaxi, phenylpropoxy, phenylmethoxy and benzyl;

(iii) R1represents-S(O)Riiiand Riiiselected from methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, acetonitrile, phenyl, phenylmethoxy, venlafaxi, phenylpropoxy, phenylmethoxy and benzyl;

(iv) R1represents-S(O)2Rivand R ivselected from methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, acetonitrile, phenyl, phenylmethoxy, venlafaxi, phenylpropoxy, phenylmethoxy and benzyl;

(v) R1represents-C(O)NRvRviwhere Rvand Rviindependently selected from H, methyl, hydroxymethyl, ethyl, hydroxyethyl, propyl, hydroxypropyl, butyl, hydroxybutyl, diethylaminoethyl, phenyl, pyridinyl, methoxyethyl, hydroxyethoxyethyl, benzyl, methylphenyl, phenylethyl, hydroxyhydrocinnamate, carbamoylmethyl and hydroxymethyl of hydroxyethyl;

(vi) R1represents-C(O)NRvRviwhere Rvand Rvitogether form a morpholine, piperazine, complex ethyl ester piperazine;

(vii) R2selected from methylene, ethylene, propylene and butylene;

(viii) R2represents ethylene, and R3represents H;

(ix) R1is a CF3or halogen.

Here are illustrated the compounds of formula I-E, including a group selected from 3-(2-amino-ethyl)-5-methoxy-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-(3-hydroxypropoxy)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-ethoxy-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-methanesulfonyl-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-(2-hydroxyethoxy)-1,3-dihydrobenzo Gasol-2-it, amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-5-methylsulfanyl-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-methanesulfonyl-1,3-dehydrobenzperidol-2-it, (2-diethylaminoethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-aminopropyl)-2,3-dehydrobenzperidol-2-it, [3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-yloxy]acetonitrile, ethylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, pyridine-3-ylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-methoxyethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 1-(2-amino-ethyl)-1,3-dehydrobenzperidol-2-it, 1-(2-amino-ethyl)-1,3-dihydronaphtho[2,3-d]imidazol-2-it, (2-hydroxyethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-5-propoxy-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-c]pyridine-2-it, (2-diethylaminoethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-4-carboxylic acid, pyridine-4-ylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-aminoethyl)-1,3-dihydroimidazo[4,5-b]pyridine-2-it, 1-(3-aminopropyl)-1,3-dehydrobenzperidol-2-it, phenylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, [2-(2-hydroxyethoxy)this is]amide of 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 1-(2-amino-ethyl)-5-trifluoromethyl-1,3-dehydrobenzperidol-2-it, 1-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-c]pyridine-2-it, benzylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-5-(morpholine-4-carbonyl)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-(2-oxo-2-phenylethane)-1,3-dehydrobenzperidol-2-it 3-(2-methylaminomethyl)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-butoxy-1,3-dehydrobenzperidol-2-it, methylphenylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, ethyl ester 4-[3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carbonyl]piperazine-1-carboxylic acid diethylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, phenetylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-hydroxy-1-hydroxymethyl-2-phenylethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, carbamoylmethyl 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-hydroxy-1-hydroxymethylation)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, N-{2-[2-oxo-2,3-dihydrobenzoic-1-yl]ethyl}guanidine, 3-(2-amino-ethyl)-5-benzyloxy-1,3-dehydrobenzperidol-2-it 1-(4-aminobutyl)-1,3-dehydrobenzperidol-2-it. In one of these embodiments provides for the connection of 3-(2-and inatel)-1-(2-isopropyl-5-methylcyclohexanone)-5-methoxy-1,3-dehydrobenzperidol-2-it.

Synthesis of low molecular weight modulators of Trp-p8

As noted above, the compounds of the present invention include compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E. In certain aspects, the compounds can be obtained using commercially available starting materials by using the methodology of synthesis are readily available in this area. The compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E can be selected using the typical methods of isolation and purification known in this field, including, for example, chromatographic methods and ways of recrystallization.

The person skilled in the art will easily see that connection, which is convenient to include in the compositions and methods of the present invention can exist as CIS - and TRANS-isomers, E/Z forms, diastereoisomers and optical isomers. Thus, the compounds used in the compositions and methods of the present invention include all such combinations and variations.

In the compounds of formula I, formula I-A, formula I-B, formula I-C, formula 1-D, and formula I-E the carbon atoms to which are attached four non-identical Deputy, are asymmetric. Accordingly, the compounds of formula I, formula I-A, formula I-B, formula IC, formula I-D, and formula I-E can exist as enantiomers, diastereomers or their mixture. Enantiomers and diastereomers can be separated by chromatographic methods or methods of crystallization, or using other methods known in this field. Asymmetric carbon atom can be in one of two configurations, R or S, both of which are within the present invention. The presence of a small amount of the opposite enantiomer or diastereoisomer in the final purified product does not affect therapeutic use of such compounds.

The compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula 1-E can further be processed with the formation of pharmaceutically acceptable salts. Treatment of the compounds of the present invention with an acid or a base can give respectively pharmaceutically acceptable acid additive salt and pharmaceutically acceptable basic additive salt, each of them is such as defined above. Various inorganic and organic acids and bases known in this area, including those described herein above may be used to effect conversion into salt.

The present invention also relates to pharmaceutically acceptable isomers, hydrates and Solvay the am compounds of the formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E. the Compounds of these formulas may also exist in various isomeric and tautomeric forms, including pharmaceutically acceptable salt, hydrate and solvate of such isomers and tautomers.

The present invention also encompasses derivatives, prodrugs of compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E. the Term "prodrug" refers to a derivative of the original molecules of drugs that require biological conversion in the body, either spontaneous or enzymatic, for the release of the original medicinal product. Prodrugs are variations or derivatives of compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E of the present invention, which has groups otsepleniya with metabolic conditions. Prodrugs become compounds of the present invention which are pharmaceutically active in vivo, when they undergo solvolysis under physiological conditions or undergo enzymatic degradation. Exemplary technology prodrugs, which may be suitable for use with the compounds of the present invention, is a technology protease-activated cancer treatment (PACT), the description is nnow in detail in the patent application U.S. No. 10/156214 and Published PCT applications for international patent # WO 02/095007, both of them are included in the present description as a reference.

The synthesis of compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E can be achieved by interaction of the acid chloride obtained by the interaction of p-methane-3-carboxylic acid with thionyl chloride with the appropriate amine. As noted below, as a rule, the interaction is carried out in solution at room temperature in the presence of receptor hydrogen chloride (e.g., sodium hydroxide).

The basic structure of a p-methane is kresloobraznye molecule that can exist in CIS - or TRANS-form. Replacement of the carboxyl or amide group at the 3-position gives rise to the four configurations or geometrical isomers, depending on whether the substitution axial or Equatorial, CIS - or TRANS-isomer, four isomers are related to each other as menthol with neomenthol, isomenthol and neotantra.

In exemplary embodiments, the implementation of the compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E are synthesized with the specific stereochemistry, where the relative stereochemistry around methane ring is the same as menthol, and/or where the absolute stereochemistry around methane ring is the same as (-)-menthol.

The synthesis methods for obtaining limernyh low molecular weight modulators of Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E of the present invention here presented in examples 1-9.

Compositions containing small molecule modulators of Trp-p8

As discussed above, the present invention is directed to small molecule modulators of Trp-p8, including agonists Trp-p8 and antagonists Trp-p8 that are associated with Trp-p8 and alter its activity. In certain embodiments of the implementation of the modulators of Trp-p8 represent agonists, which are in some cases capable of stimulating the influx of cations and toxicity to cells expressing the protein channels Trp-p8. In alternative embodiments, the implementation of the modulators of Trp-p8 are antagonists of the activity of Trp-p8, which can reduce the activity of ekspressirovali Trp-p8 cells. Thus, modulators of Trp-p8 of the present invention can find use in the compositions containing pharmaceutical compositions which are suitable for treatment of diseases associated with expressionism Trp-p8. The corresponding compositions in accordance with the present invention contain one or more agonists Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E and/or one or more antagonists of Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E, as anywayse above, in combination with one or more pharmaceutically acceptable carriers or excipients.

In one of the embodiments the present invention provides small molecule modulators of Trp-p8 in combination with a pharmaceutically acceptable excipient such as sterile saline solution or other environment, water, gelatin, oil and the like, with the formation of pharmaceutically acceptable compositions. The composition and/or agonists can be administered by themselves or in conjunction with any conventional carrier, diluent and the like, and such administration can be provided in one or in multiple doses. Suitable carriers include, but are not limited to, solid, semi-solid or liquid medium comprising water and non-toxic organic solvents.

The pharmaceutical compositions of the present invention can be obtained by mixing one or more agonists Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E with a pharmaceutically acceptable carrier or agent. Alternative pharmaceutical compositions can be obtained by mixing one or more antagonists of Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E with a pharmaceutically acceptable carrier or agent. the addition of the pharmaceutical composition can optionally include fillers, stabilizers, diluents and the like, and may include drugs with a slow release or with distributed time-release. Acceptable carriers, agents, fillers, stabilizers, diluents and the like for therapeutic use are well known in the pharmaceutical field and are described, for example, in "Remington''s Pharmaceutical Sciences", (Mack Publishing Co., ed. A.R. Gennaro, 1985), which is included in the present description by reference. Such materials are nontoxic to the patient when used dose and concentration, and include buffers such as phosphate, citrate, acetate and other organic acid salts, antioxidants such as ascorbic acid, low molecular weight peptides, such as polyalanine, proteins such as serum albumin, gelatin or immunoglobulins, hydrophilic polymers, such as serum albumin, gelatin or immunoglobulins, hydrophilic polymers such as polyvinylpyrrolidone, amino acids such as glycine, glutamic acid, aspartic acid, or arginine, monosaccharides, disaccharides, and other carbohydrates including cellulose or its derivatives, glucose, mannose or dextrin, chelating agents such as EDTA, sugar alcohols such as mannitol or sorbitol, counterions such as sodium; and/or nonionic surfactants which, such as TWEEN or polyethylene glycol.

In other aspects, the compositions of the present invention contain a compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E, prepared together with one or more therapeutic agents against cancer. Alternative compositions of the present invention contain a compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E, independently prepared with one or more therapeutic agents against cancer. That is, the compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E and therapeutic agent against cancer, are prepared separately.

Suitable for use with therapeutic agents against cancer include, but are not limited to, antimitoticescoe agents, including, but not limited to, paclitaxel, vincristine and etoposide; alkylating agents, including, but not limited to, mechlorethamine, cyclophosphamide, carmustine; antimetabolites, including, but not limited to, methotrexate, gemcitabine, lometrexol, 5-fluorouracil and 6-mercaptopurine; cytotoxic antibiotics, including, but not limited to, doxorubicin, daunorubicin, bleomycin, mitomycin C and streptozocin; agents based on platinum, which includes, but is not limited to IVaS this, cisplatin and carboplatin; hormonal agents, including, but not limited to, antiestrogens, such as tamoxifen and diethylstilbestrol, and antiandrogens, such as flutamide; agents against angiogenesis and inhibitors farnesyltransferase.

In certain aspects of compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E are entered in combination with a therapeutic agent against cancer, which is ineffective at stimulating mediamoo Trp-p8 influx of cations.

In other aspects, the compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E are entered in combination with one or more modulators of Trp-p8, including, but not limited to, the compound of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E.

Depending on the specific proposed treatment the pharmaceutical composition of the present invention may be introduced parenterally, local way, orally or locally. In certain aspects, the pharmaceutical compositions are injected parenterally, for example intravenously, subcutaneously, intradermally or intramuscularly. In one of the embodiments the present invention provides compositions for parenteral administration which contain the connection p of the present invention, dissolved or suspended in a carrier such as the carrier is water-based.

For solid preparations of the compounds may be mixed with conventional non-toxic solid carriers such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, saccharin sodium, talc, cellulose, glucose, sucrose, magnesium carbonate and the like.

For aerosol injection of the compounds of the present invention can be supplied in fine form along with a non-toxic surface-active substances and propellants. Examples of such agents are esters or partial esters of fatty acids containing from 6 to 22 carbon atoms, such as Caproic, octanoic, lauric, palmitic, stearic, linoleic, holesterinova and oleic acid.

The compositions of the present invention can be administered by injection, that is, intravenously, intramuscularly, transdermally, subcutaneously, intraduodenally or intraperitoneally. Alternative compositions can be administered by inhalation, for example, intranasal and can be entered transdermal, for example, using an adhesive or the like.

It will be understood that actually the preferred drug compositions containing the pharmaceutical composition will vary according what about the way of introduction, and with the specific disease that should be treated. Optimal drugs and routes of administration will be routine to be defined for this specific disease and the patient's specialist in this field.

Method identification and characterization of in vitro and in vivo efficacy of low molecular weight modulators of Trp-p8

As discussed above, the present invention is directed to small molecule modulators of Trp-p8, including agonists and antagonists of the activity of Trp-p8. These substances are modulators of Trp-p8, illustrated by compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E, described here above. The present invention, furthermore, suggests that additional modulators of Trp-p8 may also suitably be used in the compositions and methods of the present invention.

Additional or alternative agonists and antagonists Trp-p8 can be identified using the methodology described in the accompanying examples. For example, agonists Trp-p8, having efficacy in the treatment of diseases (diseases)associated with the expression of Trp-p8, include small molecules, which lead to one or more of the following results: (1) inhibition of growth or decrease in the survival of cells expressing Trp-p8; (2) the incentive is a key tributary of the calcium and/or other cations in cells expressing Trp-p8; (3) induction of apoptosis and/or necrosis in cells expressing Trp-p8; and/or (4) the efficiency for one or more systems of animal models of human disease. Antagonists Trp-p8, having efficacy in the treatment of diseases (diseases)associated with the expression of Trp-p8, include small molecules, which lead to one or more of the following results: (1) to protect expressing Trp-p8 cells from the toxic effect of agonists in the system models in vitro; (2) inhibition of growth and/or death in the line of cancer cells with endogenous expression of Trp-p8; (3) they are effective for one or more systems of animal models of human disease.

Thus, in certain embodiments of the implementation of the present invention provides methods of identifying agonists Trp-p8, which includes a stage of contacting cells expressing Trp-p8, with candidates agonists Trp-p8, within the time and in a quantity sufficient to inhibit the growth and/or decrease the survival expressing Trp-p8 cells, where the growth inhibition and/or reduction of the survival rate shows that the candidate agonists Trp-p8 is able to activate expressiona Trp-p8 cells.

Other embodiments of provide methods of identifying agonists Trp-p8 with adiu contacting cells expressing Trp-p8, with candidates agonists Trp-p8, within the time and in sufficient quantity to induce the influx of calcium and/or other cations in the cell, which increases influx of cations correlates with increased cellular toxicity.

Other embodiments of provide methods of identifying agonists Trp-p8, which includes the stage of introduction of candidates agonists Trp-p8 animals with one or more neoplastic cells that Express Trp-p8, within the time and in a quantity sufficient to inhibit the growth and/or induction of apoptosis and/or necrosis in cells, thereby increasing the survival of animals, where any one or more processes from the inhibition of cell growth, induction of apoptosis, induction of necrosis and/or increase the survival of animals show the effectiveness of agonist Trp-p8.

The present invention provides methods of identifying antagonists of Trp-p8, in addition to antagonists Trp-p8 described here. This method includes (1) analysis of the system in vitro to detect the protection of expressing Trp-p8 cells from toxicity induced by agonists Trp-p8; (2) analysis of the in vitro and in vivo detection of inhibiting the growth of cancer cells and/or lines of cancer cells, endogenously expressing Trp-p8; (3) systems animal models in vivo, the de one or more candidates in antagonists Trp-p8 entered the animal, with one or more neoplastic cells that Express Trp-p8, within the time and in a quantity sufficient to inhibit the growth and/or induction of apoptosis and/or necrosis in cells, thereby increasing the survival of animals.

Uses of modulators of Trp-p8

Small molecule modulators of Trp-p8 of the present invention can be suitably used in the methods of modification (i.e. activate or decrease) mediamoo Trp-p8 influx of calcium into cells and therapeutic methods of treating one or more diseases associated with expressionism Trp-p8. For example, and as noted above, it is observed that abnormal expression of Trp-p8 is associated with neoplastic phenotype in different cancer tissues, including breast tissue, colon, lung and prostate. Tsavaler et al., Cancer Research, see above.

Thus, in certain embodiments of the implementation provides methods of activating mediamoo Trp-p8 influx of calcium into cells, such methods include the stage of contacting cells expressing Trp-p8, with some agonist Trp-p8 in a period of time sufficient to inhibit the growth of cells and/or to induce necrosis and/or apoptosis in cells. Examples of methods of activation of Trp-p8 are given in the examples, the sight of the purposes here.

Other embodiments of the present invention provide therapeutic methods for treating diseases associated with expressionism Trp-p8, such methods include the stage of introduction of the mammal, typically a human, a therapeutically effective amount of a composition containing an agonist Trp-p8, a period of time sufficient to inhibit the growth of cells and/or to induce necrosis and/or apoptosis in cells. As used here, the phrase "therapeutically effective amount" refers to the amount of compound that, when introduced to a mammal for treating a disease, is sufficient to effect such treatment for the disease. "Therapeutically effective amount" will vary depending on the compound, the disease and its severity and the age, weight and the like of a mammal, which should be treated.

As used here, the terms "treat", "treated" and "treatment" include: (1) preventing the disease, i.e. preventing the development of clinical symptoms of the disease in a mammal which may be predisposed to the disease but does not feel until any symptoms of the disease; (2) inhibiting the disease, i.e. suspension or delay development of the disease or its clinical symptoms, or (3) facilitation of the disease, that is, the implementation of regression of the disease or its clinical symptoms.

Although the frequency and dosage regimens will vary depending on factors such as the disease and the treatment of the patient, the composition comprising one or more compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E, as a rule, are entered in the range from about 0.001 mg compound/kg body weight to about 1000 mg/kg As a rule, treatment is initiated with smaller dosages that are less than the optimum dose of the compound. After that, the dose can be increased until then, until you reach optimal efficiency.

In most cases, the introduction of the compositions (compositions) of the present invention is carried out by any method that provides a systematic exposure to the compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E. Thus, the compositions can be administered orally, parenterally, intraduodenal and intranasal. Typically, such compositions contain one or more of such compounds in combination with one or more pharmaceutically acceptable carriers or diluents, as described here in more detail.

Other embodiments of the present invention provide for combination therapy, where one or the number of compounds of the formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E are entered in combination with one or more therapeutic agents against cancer, as here described in more detail above, such as antimitoticescoe agent, an alkylating agent, an antimetabolite, a cytotoxic antibiotic, an agent platinum-based hormonal agent and/or antiandrogen. In addition, embodiments of the present invention provide for combination therapy, where two or more compounds of formula I, formula I-A, formula I-B, formula I-C, formula I-D and/or formula I-E are either simultaneously or sequentially to achieve the desired therapeutic result.

Thus, as used here, the term "combination" means that at least two compounds can be delivered simultaneously in a combined therapy, where you first entered the first connection, and then a second connection, and where first delivered to the second connection, and then the first connection. The desired result can be either subjective weakening of the symptom (symptoms)or an objectively identifiable improvement in a patient, which produce dosing.

The following examples are offered as illustrative and not as limitations.

EXAMPLES

Example 1

The synthesis of compounds of meta is-3-carboxamide

This example describes a methodology for the synthesis of compounds methane-3-carboxamide.

Methane-3-carboxylic acid

Water (300 ml) was placed in 2-l Erlenmeyer flask with a stirrer in the form of a wide bar. Carefully add sulfuric acid (500 ml) under stirring. Solution allow to cool to 75°C and add N-ethyl-p-methane-3-carboxamide (62.5 g). The temperature of the support at 75°C using a heated table and carefully add sodium nitrite (31 g). Add two 31-gram portions NaNO2at 1-hour intervals and the mixture is stirred overnight at 75°C.

The mixture is cooled to room temperature, diluted ~1 l of ice water and extracted with ~500 ml of a simple ester. The ether layer is separated, washed with water and extracted with 2×350 ml 1M NaOH. The aqueous layer was acidified with concentrated HCl and extracted with simple ether. The ether layer is dried over MgSO4and evaporated to obtain methane-3-carboxylic acid (33,2 g, 61%) as a crystalline solid, DDQ=-50,3 deg. (c=1, CHCl3, 25°C).

Methane-3-carbonylchloride

Methane-3-carboxylic acid (54,35 g) is heated under reflux with 80 ml of thionyl chloride for 3 hours. SOCl2removed by distillation and the acid chloride distil at 114-115°C (8 Torr). (Because, literature, 84-85°C at 3.5 Torr). Yield: 50 g (84%).

The General procedure is prigotovleniya methane-3-carboxamide

To a mixed solution of 0.2 mmol of amine in 1 ml of acetonitrile or NMP and 0.4 mmol DIPEA add to 0.022 ml of methane-3-carbonylchloride. The reaction mixture is shaken for 3 hours. For the less active chemically amines the mixture is heated (60°C) and shaken for 24 hours. The product is recovered from the crude reaction mixture by HPLC (gradient 40-95% within 10 minutes with the use of 0.05% TFA in CH3CN and 0.05% TFA in H2O) and evaporated to dryness.

Example 2

The synthesis of compounds of dihydrobromide formula 1-E

This example describes the synthesis methodology dehydrobenzperidol modulators of Trp-p8 formula 1-E.

4-Methoxy-2-fluoro-1-nitrobenzene

2-l round bottom flask, equipped with a bar stirrer and reflux condenser, load acetonitrile (1 l), K2CO3(263 g, 1.9 mol) and 4-hydroxy-2-fluoro-1-nitrobenzene (100 g, 0.64 mol). To the reaction mixture add methyliodide (271 g, 1.9 mol) and heated at the temperature of reflux distilled with vigorous stirring for 5 hours. The acetonitrile is removed and add ethyl acetate (1 l) and H2O (700 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×200 ml). The combined organic phases are washed with H2O (2×500 ml), saturated salt solution (500 ml), with the shat over MgSO 4filter and concentrate under reduced pressure to give the desired product as a slightly yellowish solid (107 g, 98%).

Tert-butyl ether [2-(5-methoxy-2-nitrophenylamino)ethyl]carbamino acid

2-l flask equipped with bar stirrers, download DMSO (800 ml), K2CO3(161 g, 1.6 mol) and 4-methoxy-2-fluoro-1-nitrobenzene (100 g, of 0.58 mol). To the reaction mixture of mono-N-Boc-1,2-diaminoethane (94 g, 0.55 mol) and stirred for 12 hours at 60°C. the Reaction mixture fray with cold water with ice (1.2 l) and the resulting yellow precipitate is collected by vacuum filtration. The precipitate is washed several times with water (5×1 l) and dried under high vacuum for 48 hours to give the desired product as a bright yellow solid (178 g, 98%.)

Tert-butyl ether [2-(5-methoxy-2-aminophenylamino)ethyl]carbamino acid

2-l round bottom flask, equipped with a bar stirrers, load the suspension of 20% Pd(OH)2(5 g) and 1,4-dioxane (800 ml). To the suspension is added tert-butyl ether [2-(5-methoxy-2-nitrophenylamino)ethyl]carbamino acid (100 g, 0.32 mol). The reaction mixture hydronaut (balloon) for 48 hours (up until the source material is not used), after adding to the mixture K 2CO3(100 g) and stirred for an additional 12 hours to remove microscopic amounts of water. The suspension was filtered to remove Pd(OH)2and K2CO3. The filtrate is used in the next stage without further purification (yield not determined).

Tert-butyl ether [2-(6-methoxy-2-oxo-2,3-dihydrobenzoic-1-yl)ethyl]carbamino acid

The above solution is treated with excess carbonyldiimidazole (104 g, 0.64 mol) and heated at 90°C for 4 hours. 1,4-dioxane is removed and the residue fray with water (1.5 l). The resulting precipitate is collected by vacuum filtration and washed several times with water (5×500 ml). The crude product is dried at 70°C under high vacuum for 12 hours and used without further purification (66 g, 67% yield for 2 stages).

Tert-butyl ether {2-[3-(2-isopropyl-5-methylcyclohexanone)-6-methoxy-2-oxo-2,3-dihydrobenzoic-1-yl]ethyl}carbamino acid

2-l flask equipped with bar stirrers, download tert-butyl ether [2-(2-oxo-2,3-dihydrobenzoic-1-yl)ethyl]carbamino acid (40 g, 0.20 mol), DMAP (48 g, to 0.39 mol) and CH2Cl2(500 ml). Mentorplace (40 g, 0.20 mol) is added dropwise over 15 minutes and stirred at ambient temperature for 4 the aces. The reaction mixture was quenched with 1 N. HCl and stirred for additional 20 minutes Heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with CH2Cl2(2×200 ml). The combined organic phases are washed with 1 N. HCl (2×300 ml), H2O (300 ml), saturated NaHCO3(aq.) (2×300 ml), saturated salt solution (300 ml), dried over MgSO4filter and concentrate under reduced pressure. The crude product is dissolved in a minimum amount of CH2Cl2and elute through a layer of silica gel (10% hexane/ethyl acetate for elution) to give the desired product as a colourless solid (93 g, 96%).

The TFA salt of 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-5-methoxy-1,3-dehydrobenzperidol-2-she (compound No. 36)

500-ml round-bottom flask is charged with tert-butyl ether {2-[3-(2-isopropyl-5-methylcyclohexanone)-2-oxo-2,3-dihydrobenzoic-1-yl]ethyl}carbamino acid (90 g, to 0.19 mol) and 95% TFA/H2O (200 ml). The reaction mixture is stirred for 2 hours and TFA is removed under reduced pressure to get crude product as a thick oil which solidifies with the formation of brittle foam when standing in a vacuum). The crude product is dissolved in 30% mixture of acetonitrile/H2O and purified using preparative the Oh HPLC (Ultro 120 (10 μm) C18Q) using 40-60% gradient acetonitrile/H 2O (1% TFA). Pure fractions unite, concentrate, and lyophilizers with getting slightly loose colorless solid (79 g, 94%). MS (ESI) m/z 374(M++1).

The TFA salt of 3-(2-amino-ethyl)-5-ethoxy-1-(2-isopropyl-5-methylcyclohexanone)-1,3-dehydrobenzperidol-2-it(compound No. 38)

In a procedure similar to the synthesis of compound No. 36, compound 38 was obtained from 4-ethoxy-1-2-fluoro-1-nitrobenzene (obtained from ethylbromide and 4-hydroxy-2-fluoro-1-nitrobenzene). MS (ESI) m/z 387 (M++1).

The TFA salt of 1-(2-amino-ethyl)-3-(2-isopropyl-5-methylcyclohexanone)-1,3-dehydrobenzperidol-2-she (compound No. 50)

In a procedure similar to the synthesis of compound No. 36, compound No. 50 is obtained from 2-fluoro-1-nitrobenzene. MS (ESI) m/z 344 (M++1).

The TFA salt of 3-(2-amino-ethyl)-5-(3-hydroxypropoxy)-1-(2-isopropyl-5-methylcyclohexanone)-1,3-dehydrobenzperidol-2-she (compound No. 37)

In a procedure similar to the synthesis of compound No. 36, compound No. 37 is obtained from 4-(2-tert-butoxypropyl)-2-fluoro-1-nitrobenzene (obtained from 1-bromo-3-tert-butoxypropan and 4-hydroxy-2-fluoro-1-nitrobenzene). MS (ESI) m/z 418 (M++1).

The TFA salt of 3-(2-amino-ethyl)-5-(2-hydroxyethoxy)-1-(2-isopropyl-5-methylcyclohexanone)-1,3-digitale tinidazol-2-she (compound No. 40)

In a procedure similar to the synthesis of compound No. 36, compound No. 40 is obtained from 4-(2-tert-butoxyethoxy)-2-fluoro-1-nitrobenzene (obtained from 1-bromo-3-tert-butoxyethanol and 4-hydroxy-2-fluoro-1-nitrobenzene). MS (ESI) m/z 404 (M++1).

The TFA salt of 1-(2-amino-2-(R)-methylethyl)-3-(2-isopropyl-5-methylcyclohexanone)-1,3-dehydrobenzperidol-2-she (compound No. 45)

In a procedure similar to the synthesis of compound No. 36, compound No. 45 is obtained from 2-fluoro-1-nitrobenzene and tert-butyl methyl ether (2-amino-1-(R)-ethyl) - carbamino acid. MS (ESI) m/z 358 (M++1).

Example 3

Synthesis of additional compounds of dihydrobromide formula I-E

This example describes the synthesis methodology dehydrobenzperidol modulators of Trp-p8 formula I-E.

Methyl ester of 3-Fluoro-4-nitrobenzoic acid

1-l round bottom flask, equipped with a bar stirrer and reflux condenser, download H2SO4(4 ml), methanol (400 ml) and 3-fluoro-4-nitrobenzoic acid (10 g). The reaction mixture is heated at the temperature of reflux distilled with vigorous stirring for 18 hours. The methanol is removed and the crude residue fray with hexane and concentrated to obtain a colorless solid matter (9,79 g), which is used without updat the additional cleanup.

Methyl ester 3-(2-tert-butoxycarbonylmethylene)-4-nitrobenzoic acid

2-l flask equipped with bar stirrer, charged with 1,4-dioxane (300 ml), DMF (40 ml), K2CO3(10 g) and 3-fluoro-4-nitrobenzoic acid (9.7 g). To the reaction mixture of mono-N-Boc-1,2-diaminoethane (8.6 g) and stirred for 12 hours at 60°C. the Reaction mixture was concentrated and the residue dissolved in CH2Cl2(400 ml) and H2O (500 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with CH2Cl2(2×100 ml). The combined organic phases are washed with H2O (5×100 ml), dried over Na2SO4filter and concentrate under reduced pressure to obtain specified in the title compound as an orange solid (14 g, 84%).

Methyl ester of 4-amino-3-(2-tert-butoxycarbonylmethylene)benzoic acid

2-l round bottom flask, equipped with a bar stirrers, load the suspension of 20% Pd(OH)2and 1,4-dioxane (400 ml). To the suspension is added methyl ether 4-amino-3-(2-tert-butoxycarbonylmethylene)benzoic acid (14 g). The reaction mixture hydronaut (balloon) for 48 hours (up until the source material is not used), after adding to the mixture of K CO3(100 g) and stirred for an additional 12 hours to remove microscopic amounts of water. The suspension was filtered to remove Pd(OH)2and K2CO3.The filtrate is used in the next stage without additional purification.

Methyl ester 3-(2-tert-butoxycarbonylamino)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid

The above solution is treated with excess carbonyldiimidazole (26,8 g, 4 EQ.) and heated at 90°C for 4 hours. 1,4-dioxane is removed and the residue fray with water (1.5 l). The resulting precipitate is collected by vacuum filtration and washed several times with water (5×500 ml). The crude product is dissolved in a minimum amount of CH2Cl2and purified using flash chromatography on silica gel, methanol/CH2Cl2(10% for elution) to give the desired product in the form of a whitish solid (11.8 g, 85%).

3-(2-tert-Butoxycarbonylamino)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid

2-l flask equipped with bar stirrer, charged with 1,4-dioxane (70 ml), methyl ether 3-(2-tert-butoxycarbonylamino)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (10.4 g) and LiOH (3.7 g)dissolved in H2O (300 ml). The reaction solution premesis the Ute for 6 hours at 65°C. The mixture is concentrated and the crude residue is dissolved in H2O. the Solution is neutralized with concentrated HCl (aq.) and the resulting precipitate collected by vacuum filtration. The solid product is washed several times H2O and dried under high vacuum overnight to give the desired product as a white solid (8,66 g, 87%).

The TFA salt of 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid

In a 100-ml reaction vessel equipped with a bar stirrers, download THF (20 ml), DMAP (1.8 g) and 3-(2-tert-butoxycarbonylamino)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (4 g). The reaction mixture was cooled to 0°C and treated with motorglider (2.9 g). The reaction mixture was allowed to warm to ambient temperature and concentrate. Add 1 N. HCl (aq.) (50 ml) and CH2Cl2(50 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×100 ml). The combined organic phases are washed with 1 N. HCl (2×50 ml), H2O (50 ml), saturated salt solution (100 ml), dried over MgSO4filter and concentrate under reduced pressure. The residue is purified using flash chromatography on silica gel (4:1 CH2Cl2/THF for suirou the deposits) to obtain specified in the title compound as a colourless solid (3.2 g, 52%).

The TFA salt of amide 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (compound No. 41)

A 10-ml reaction vessel equipped with a bar stirrers, download DMF (5 ml), 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (1.5 g, 3.9 mmol), EDC (824 mg, 4.3 mmol), HOBt (581 mg, 4.3 mmol), DIEA (1,11 g, 8.6 mmol) and NH4Cl (230 mg, 4.3 mmol). The reaction mixture is heated using a microwave oven at 60°C for 10 min and poured into a mixture of ethyl acetate (50 ml) and 1 N. HCl (50 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×50 ml). The combined organic phases are washed with 1 N. HCl (100 ml), H2O (2×100 ml), saturated NaHCO3(3×100 ml), saturated salt solution (100 ml), dried over MgSO4filter and concentrate under reduced pressure. The residue is dissolved in 95% TFA/H2O, stirred for 2 hours and concentrated. The crude product is dissolved in 30% mixture of acetonitrile/H2O and purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 10-60% acetonitrile/H2O (0.1% TFA). Pure fractions unite, concentrate, and lyophilizers with getting slightly loose colorless solid (910 mg, 1%). MS (ESI) m/z 387 (M++1).

The TFA salt of (2-diethylaminoethyl)amide 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (compound No. 44)

In a procedure similar to the synthesis of compound No. 41, compound No. 44 is obtained from N',N'-diethylether-1,2-diamine. MS (ESI) m/z 486 (M++1).

The TFA salt of ethylamide 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (compound No. 47)

In a procedure similar to the synthesis of compound No. 41, compound No. 47 get from ethylamine. MS (ESI) m/z 415 (M++1).

The TFA salt of pyridine-3-yl amide 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (compound No. 48)

In a procedure similar to the synthesis of compound No. 41, compound No. 48 is obtained from pyridine-3-ylamine. MS (ESI) m/z 464 (M++1).

Example 4

Synthesis of additional compounds of dihydrobromide formula I-E

This example describes the synthesis methodology dehydrobenzperidol modulators of Trp-p8 formula I-E.

Tert-butyl ether [2-(5-methylsulfanyl-2-nitrophenylamino)ethyl]carbamino acid

In a 1-liter round-bottom to the forehead, equipped with a bar stirrers, download DMSO (200 ml), K2CO3(13 g, 0.10 mol) and 2,4-debtor-1-nitrobenzene (5 g, 0.03 mol). The reaction mixture is treated with mono-N-Boc-1,2-diaminoethane (5 g, 0.32 mol) and stirred at ambient temperature for 18 hours. Sodium thiomethoxam (2.24 g, 0.03 mol) is added to the reaction mixture and stirred for 12 hours at 60°C. the Reaction mixture was cooled to 0°C and fray with water (800 ml), the yellow precipitate which forms is collected by vacuum filtration. The precipitate is washed several times with water (5×500 ml) and dried under high vacuum for 48 hours to give the desired product as a bright yellow solid (8.7 g, 71%).

Tert-butyl ether [2-(2-amino-5-methylsulfonylamino)ethyl]carbamino acid

In a 500-ml round-bottom flask equipped with bar stirrers, download MeOH (200 ml), tert-butyl ether [2-(5-methylsulfanyl-2-nitrophenylamino)ethyl]carbamino acid (5 g, 0.02 mol) and NiCl2(19 g, 0.05 mol) and cooled to 0°C. Add NaBH4(1.7 g, 0.05 mol) in four equal portions to the reaction mixture over a period of 1 hour. After complete addition, the reaction mixture is stirred for additional 2 hours. To the reaction mixture is added a saturated salt solution (100 ml) and ethyl acetate (200 ml). Heterogen the th mixture is transferred into a separating funnel, where the aqueous phase is separated and re-extracted with ethyl acetate (2×100 ml). The combined organic phases are washed with H2O (3×100 ml), saturated salt solution (100 ml), dried over MgSO4filter and concentrate under reduced pressure to obtain a residue of black color. The crude product is dissolved in 100 ml of CH2Cl2,divided into two parts 100-ml portion (50 ml each) and both concentrated under reduced pressure and used without further purification.

3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-5-methylsulfanyl-1,3-dehydrobenzperidol-2-on (compound No. 42)

In a procedure similar to the synthesis of compound No. 36, compound No. 42 is obtained from the crude tert-butyl ether [2-(2-amino-5-methylsulfonylamino)ethyl]carbamino acid. MS (ESI) m/z 390,1 (M++1).

3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-5-methylsulfanyl-1,3-dehydrobenzperidol-2-on (compound No. 43)

10-ml reaction flask is charged with 3-(2-amino-ethyl)-l-(2-isopropyl-5-methylcyclohexanone)-5-methylsulfanyl-1,3-dehydrobenzperidol-2-one (compound No. 42, 300 mg) and 1% TFA/DMSO (1 ml). Oxygen is bubbled through the reaction mixture for 20 min and sealed. The reaction mixture is stirred for 18 hours and the crude product is purified is by using preparative HPLC (Ultro 120 (10 μm) C18Q) using 40-60% gradient acetonitrile/H 2O (1% TFA). Pure fractions unite, concentrate, and lyophilizers with getting slightly loose colorless solid (296 mg, 94%). MS (ESI) m/z 406 (M++1).

3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-5-methylsulphonyl-1,3-dehydrobenzperidol-2-on (compound No. 39)

10-ml reaction vessel equipped with a bar stirrers, download 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-5-methylsulfanyl-1,3-dehydrobenzperidol-2-one (compound No. 42), Oksana (1 g) and 20% aqueous solution of methanol (5 ml). The reaction mixture is titrated with a saturated NaHCO3(aq.) to pH~5. The reaction mixture is stirred for 1 hour. The reaction mixture is filtered and concentrated. The crude product is dissolved in 30% mixture of acetonitrile/H2O and purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 15-50% acetonitrile/H2O (0.1% TFA). Pure fractions unite, concentrate, and lyophilizers with getting loose colorless solid (79 g, 94%). MS (ESI) m/z 422 (M++1).

Example 5

The synthesis of compounds of formula I-B

This example describes the synthesis methodology dehydrobenzperidol modulators of Trp-p8 formula I-A.

(4-Methoxy-2-nitrophenyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid

2Cl2(100 ml) and 1 N. HCl (100 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with CH2Cl2(2×100 ml). The combined organic phases are washed with 1 N. HCl (8×100 ml), H2O (1×100 ml), 1 N. NaOH (2×100 ml), saturated salt solution (100 ml), dried over MgSO4,filter and concentrate under reduced pressure. The residue is purified using flash chromatography on silica gel (20-50% mixture of ethyl acetate/hexane for elution) to obtain the specified title compound as a colourless solid (4.9 g, 83%).

(2-Amino-4-methoxyphenyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid

(4-Methoxy-2-nitrophenyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid (4.9 g) was dissolved in a suspension of 10% Pd-C (5 g) and THF (150 ml). The reaction mixture hydronaut over 20% Pd(OH)2within 48 hours with balloon. The reaction mixture was filtered and concentrated to give the desired compound of sufficient purity to use in the next reaction without further purification.

Tert-butyl methyl ether (1-{2-[2-isopropyl-5-methylcyclohexanone)amino]-5-methoxyphenylacetyl}ethyl) - carbamino acid

(2-Amino-4-methoxyphenyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid (5 g, to 0.016 mol), EDC (4,2 g of 0.022 mol), HOBt (2,97 g of 0.022 mol) and DIEA (8.53 g, of 0.066 mol) is dissolved in DMF (50 ml) and stirred at 45°C for 6 hours. The reaction mixture is cooled to room temperature and poured into a mixture of ethyl acetate and 1 N. HCl (100 ml). A heterogeneous mixture is transferred into a separating funnel and the phases are separated. The aqueous phase is re-extracted with ethyl acetate () and the combined organic phases, washed with 1 N. HCl (5×100 ml), H2O (100 ml), saturated 1 N. NaOH (2×100 ml), saturated salt solution (100 ml), dried (MgSO4), filtered and concentrated to obtain slightly yellow solid (7.5 g). Part of the crude product (1.5 g) is purified using flash chromatography on silica gel (SiO230% of a mixture of ethyl acetate/hexane for elution) to give the desired product as a colourless solid (1.6 g).

The TFA salt of [2-(2-aminodiphenylamine)-4-methoxyphenyl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 1)

Tert-butyl methyl ether (1-{2-[2-isopropyl-5-methylcyclohexanone)amino]-5-methoxyphenylacetyl}ethyl) - carbamino acid (1 g) of restore the t in 95% TFA/H 2O and stirred for 1 hour. The reaction mixture was concentrated and the crude product is dissolved in 30% mixture of acetonitrile/H2O and purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 40-60% acetonitrile/H2O (0.1% TFA). Pure fractions unite, concentrate, and lyophilizers with getting slightly loose colorless solid (880 mg). MS (ESI) m/z 376 (M++1).

The TFA salt of [2-(2-aminoethylamino)-4-methylsulfinylphenyl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 4)

A 100-ml round-bottom flask equipped with bar stirrer, containing the crude tert-butyl ether [2-(2-amino-5-methylsulfonylamino)ethyl]carbamino acid load THF (50 g) and DMAP (1.8 g, 0.02 mol). The reaction mixture was cooled to 0°C and added dropwise to mentorplace (1.5 g, 0,008 mol) over a period of 5 minutes the Reaction mixture was allowed to warm to ambient temperature and stirred for an additional 30 minutes the Crude product is dissolved in a minimum amount of CH2Cl2and purified using flash chromatography on silica gel (10% hexane/ethyl acetate for elution), which gives a slightly yellow solid (1,76 g, 61%). The purified material was dissolved in 20 ml of 95% TFA/H2O and stirred for 1 hour, and concentrated. Crude about the SPS was dissolved in 30% mixture of acetonitrile/H 2O and purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 40-60% acetonitrile/H2O (0.1% TFA). Pure fractions unite, concentrate, and lyophilizers with getting slightly loose colorless solid (1,41 g). MS (ESI) m/z 364 (M++1).

Tert-butyl ether [2-(4-Fluoro-2-nitrobenzylamine)ethyl]carbamino acid

In a 100-ml round-bottom flask equipped with bar stirrers, load acetonitrile (40 ml), EDC (1.12 g, 5.9 mmol), HOBt (coefficient was 0.796 g, 5.9 mmol), DIEA (3,76 ml, 21.6 mmol) and mono-N-Boc-1,2-diaminoethane (0,865 g, 5.4 mmol). The reaction mixture is stirred for 18 hours and concentrated. The residue is dissolved in a mixture of ethyl acetate (50 ml) and 1 N. HCl (50 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×100 ml). The combined organic phases are washed with 1 N. HCl (2×50 ml), H2O (1×50 ml), saturated NaHCO3(3×50 ml), saturated salt solution (100 ml), dried over MgSO4filter and concentrate under reduced pressure. The residue is purified using flash chromatography on silica gel (30-50% ethyl acetate/hexane for elution) to obtain the specified title compound as a slightly purple solid (1.12 g, 63%).

Tert-butyl ether [2-(5-methylsulfanyl-2-NIT is openline)ethyl]carbamino acid

A 10-ml reaction vessel equipped with a bar stirrers, download DMF (5 ml), NaSMe (rate £ 0.162 g, 2.3 mmol) and tert-butyl methyl ether [2-(4-fluoro-2-nitrobenzylamine)ethyl]carbamino acid (0,757 g, 2.3 mmol). The reaction mixture was stirred at ambient temperature for 2 hours and poured into a mixture of ethyl acetate (20 ml) and H2O (25 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×10 ml). The combined organic phases are washed with 1 N. HCl (2×10 ml), H2O (1×10 ml), saturated. NaHCO3(2×10 ml), saturated salt solution (10 ml), dried over MgSO4filter and concentrate under reduced pressure. The residue is purified using flash chromatography on silica gel (30-50% ethyl acetate/hexane for elution) to obtain the specified title compound as a slightly yellow solid (500 mg, 61%).

N-(2-amino-ethyl-2-[(2-isopropyl]-5-methylcyclohexanone)amino-4-methylsulfonylbenzoyl (compound No. 2)

In a procedure similar to the synthesis of compound No. 42, compound No. 2 obtained from tert-butyl ether [2-(5-methylsulfanyl-2-nitrophenylamino)ethyl]carbamino acid. MS (ESI) m/z 392 (M++1).

Example 6

The synthesis of compounds of formula I-B

This example describes the synthesis methodology dihydrobenzo the azole modulators of Trp-p8 formula I-B.

Scheme 5

2-(5-Amino-3-phenylpyrazol-1-yl)ethanol

Benzoylacetonitrile (25 g, to 0.17 mol) is suspended in a mixture of 125 ml of anhydrous alcohol chemical quality and 20 ml of glacial acetic acid. 2-Hydroxyethylhydrazine (14.4 g, 1.1 equiv.) dissolved in 35 ml of alcohol, add at one time. The mixture is heated under reflux for 4 hours, cooled, water is added to obtain the total volume of 500 ml, and the solution is cooled in the refrigerator overnight. The crystals are filtered off cold with a Buechner funnel, washed with cold water and dried under high vacuum to give the desired product (27,2 g, 79%).

[2-(2-Hydroxyethyl)-5-phenyl-2H-pyrazole-3-]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid

2-(5-amino-3-phenylpyrazol-1-yl)ethanol (87,3 g, 0.43 mol) is suspended in a mixture of dichloromethane (500 ml) and pyridine (40 ml) and cooled in a bath with ice. Mentorplace (100 g, 1.15 EQ.) dissolved in dichloromethane (200 ml) and added dropwise from the filling funnel protected with a drying tube containing CaCl2. After 45 min, required to complete the addition, the ice bath is removed and stirring is continued for 3 hours. 1M HCl (aq., 200 ml) is added again and the phases are separated. The organic phase is re-extracted with 1M HCl (aq., 10 ml). 1M HCl added again and the dichloromethane removed under reduced pressure, resulting in heavy precipitation. The precipitate is collected by vacuum filtration and the solids washed with water several times. The solid residue fray with 400 ml of a mixture of 1:1 simple ether/hexane (rapid stirring for 2 hours). The solid product is filtered on a Buechner funnel and washed with hexane. After air drying overnight, additional drying is carried out at high vacuum for 24 hours to obtain a colorless solid matter (144,4 g).

2-{5-[2-Isopropyl-5-methylcyclohexanecarboxylic]-3-phenylpyrazol-1-yl}ethyl ester methanesulfonic acid

[2-(2-Hydroxyethyl)-5-phenyl-2H-pyrazole-3-yl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (140 g, 0.38 mol) is suspended in CH2Cl2(500 ml) and pyridine (47 ml, 1.5 EQ.), then add methanesulfonanilide (44 ml, 1.5 EQ.) when the temperature of the ice bath. Solution allow to warm to room temperature and stirred for additional 12 hours. Add water (500 ml) and the mixture is stirred for 0.5 hours. Dichloromethane is removed by evaporation, leaving a residue of light yellow pieces in the form of granules. After desantirovaniya handle additional 500 ml of water and again decanted. Konecne-ml amount of water is used to transfer solid product in a Buechner funnel, where it is dried by pumping (yield not determined).

[2-Azidoethyl]-5-phenyl-2H-pyrazole-3-yl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid

Raw mesilate (0.38 mol) is dissolved in DMSO (500 ml) with sodium azide (37 g, 1.5 EQ.). The mixture is heated to 70°C for 6 hours. When cooled add water (1 l) and ethyl acetate (500 ml) and the mixture is shaken in a separating funnel. The layers are separated and the organic layer washed sequentially with 200 ml of water, saturated NaHCO3and a saturated solution of salt. The organic layer is dried over Na2SO4, decanted and the solvent is removed on a rotary evaporator. The output is not defined, because the product contains a certain amount of solvent before moving to the next stage.

(2-amino-ethyl)-5-phenyl-2H-pyrazole-3-alamid 2-isopropyl-5-methylcyclohexanecarboxylic acid

Raw connection azide dissolved in 500 ml of absolute alcohol chemical quality and process 5 g of activated charcoal. All this is stirred for several hours and filtered through celite. Approximately 300 ml of the solvent is removed on a rotary evaporator and replaced with fresh solvent. Add 10% Pd-C (4.8 g, ~50%-wt H2O) and maintain a constant flow of hydrogen over the reaction mixture with subsequent stirring for 24 hours. The disconnect hydrogen and m is Glenna add concentrated HCl (32 ml). After filtration through celite, the filtrate concentrated on a rotary evaporator, resulting in heavy precipitation. While it is still damp, to the residue add a simple diisopropyl ether and the suspension is rapidly stirred for 0.5 hours. The solid is filtered on a Buechner funnel and washed with simple diethyl ether. Get air-dried white powder.

Output: 108,6 g (71% for three steps).

The TFA salt of [2-amino-ethyl)-5-phenyl-2H-pyrazole-3-yl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 16)

Conversion to trifenatate salt: solid neutralize and distribute in a separating funnel by shaking with 500 ml of simple ether and 150 ml of 2 N. NaOH. When the solid is completely dissolved, the layers separated and the organic phase dried over Na2CO3. Decanted and mixed with 23 ml triperoxonane acid, followed by evaporation of the solvent and drying under high vacuum. The foam is crushed and fray with 300 ml of hexane (repeated stirring for 3 hours), when filtering gives a white powder, which contain much less simple ether. The solvent is finally removed completely by heating in a round bottom flask at 80°C for 6 hours.

The TFA salt of [2-(2-what aminoethyl)-5-furan-2-yl-2H-pyrazole-3-yl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 14)

In a procedure similar to the synthesis of compound No. 16, compound No. 14 was obtained from 2-proletaria and 2-hydroxyethylhydrazine. This material is purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 40-60% acetonitrile/H2O (0.1% TFA). MS (ESI) m/z 344 (M++1).

The TFA salt of [2-(2-amino-ethyl)-5-(1-methyl-1H-pyrrol-2-yl)-2H-pyrazole-3-yl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 17)

In a procedure similar to the synthesis of compound No. 16, compound No. 17 is obtained from 1-methyl-1H-pyrrole-2-carbaldehyde and 2-hydroxyethylhydrazine. This material is purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 40-60% acetonitrile/H2O (0.1% TFA). MS (ESI) m/z 372 (M++1).

The TFA salt of [2-(2-amino-ethyl)-5-(1-methyl-1H-pyrrol-2-yl)-2H-pyrazole-3-yl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 15)

In a procedure similar to the synthesis of compound No. 16, compound No. 15 is obtained from 2-benzoylacetonitrile and tert-butyl methyl ether (2-deadenylation)carbamino acid. This material is purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 40-60% acetonitrile/H2O (0.1% TFA). MS (ESI) m/z 383 (M++1).

The TFA salt of [2-(2-amino-ethyl)-5-(1-methyl-1H-pyrrol-2-yl)-2H-Piras the l-3-yl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 18)

In a procedure similar to the synthesis of compound No. 16, compound No. 18 is obtained from 2-benzoylacetonitrile and tert-butyl methyl ether (3-diazenyl-propyl)carbamino acid. This material is purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 40-60% acetonitrile/H2O (0.1% TFA). MS (ESI) m/z 383 (M++1).

Example 7

The synthesis of compounds of formula I-C

This example describes the synthesis methodology dehydrobenzperidol modulators of Trp-p8 formula I-C.

Scheme 6

(2-Hydroxy-2-phenylethyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid

500-ml round-bottom flask equipped with bar stirrers, download CH2Cl2(200 ml), DIEA (28 g, 0,219 mol) and 2-amino-1-phenylethanol (10 g, 0,073 mol) and cooled to 0°C. is Added dropwise mentorplace (14.8 g, 0,073 mol) over a period of 15 minutes After complete addition, the reaction allow to warm to ambient temperature and stirred for 2 hours. To the reaction mixture are added-CH2Cl2(100 ml) and 1 N. HCl (100 ml) and stirred for additional 20 minutes Heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with CH2Cl2(2×100 ml). The combined organic phases are washed with 1 N. HCl (8×100 ml), H2O (1×100 is l), 1 N. NaOH (2×100 ml), saturated salt solution (100 ml), dried over MgSO4filter and concentrate under reduced pressure. The remainder elute through a layer of silica gel (50% mixture of ethyl acetate/hexane for elution) to obtain the specified title compound as a colourless solid (18,8 g, 85%)

The TFA salt of [2-(2-amino-4-ethoxy)-2-phenylethyl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 30)

500-ml round-bottom flask equipped with bar stirrers, download anhydrous THF (200 ml) and (2-hydroxy-2-phenylethyl)amidon 2-isopropyl-5-methylcyclohexanecarboxylic acid (10 g, 0.03 mol). NaH (0.87 g, 0.04 mol) is added at once and stirred for 10 min (as long as you do not stop the formation of H2). To the reaction mixture was added 1-bromacil-2-amine hydrobromide (6,74 g 0,033 mol) and NaH (0.87 g, being 0.036 mol) and stirred for 2 hours. An additional equivalent of NaH (0.87 g, being 0.036 mol) is added and stirred for additional 2 hours. Excess NaH extinguish, pouring the reaction mixture on ice. Add ethyl acetate (200 ml) and H2O and stirred for 20 minutes Heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×100 ml). The combined organic phases are washed with H2O (1×100 ml), a saturated solution is Oli (100 ml), dried over MgSO4filter and concentrate under reduced pressure. The crude product is dissolved in 30% mixture of acetonitrile/H2O and purified using preparative HPLC (Ultra 120 (10 μm) C18Q) using a gradient of 30-60% acetonitrile/H2O (0.1% TFA). Pure fractions unite, concentrate, and lyophilizers with getting slightly loose colorless solid (9.4 g, 62%). MS (ESI) m/z 341 (M++1).

The TFA salt of [2-(3-amino-4-propoxy)-2-phenylethyl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 31)

In a procedure similar to the synthesis of compound No. 30, compound No. 31 is obtained from (2-hydroxy-2-phenylethyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid and 1-bromopropyl-3-amine hydrobromide. This material is purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 40-60% acetonitrile/H2O (0.1% TFA). MS (ESI) m/z 361 (M++1).

Example 8

Synthesis of additional dehydrobenzperidol compounds of formula I-D

This example describes the synthesis methodology dehydrobenzperidol modulators of Trp-p8 formula I-D.

Scheme 7

[2-(2-Bromo-phenyl)ethyl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid

A 100-ml round-bottom flask equipped with bar mesalc is, download CH2C12(30 ml), 2-bromophenethylamine (1.0 g, 5.00 mmol) and triethylamine (684 μl, of 5.05 mmol). The reaction solution is treated with motorglider (1,02 g of 5.05 mmol) in one portion and stirred at ambient temperature for 30 minutes. The reaction mixture was diluted with CH2Cl2(50 ml) and washed with water (3×100 ml). The organic layer is dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain a thick oil (1.8 g). The product is used in the next stage without purification.

The TFA salt of {2-[2-(2-aminoethylamino)phenyl]ethyl}amide 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 33)

25-ml reaction vessel for a microwave oven equipped with a bar stirrers, download pure diaminoethane (10 ml), [2-(2-bromophenyl)ethyl]amidon 2-isopropyl-5-methylcyclohexanecarboxylic acid (1.5 g, 4.1 mmol) and Cu powder (390 mg, 6,147 mmol, 1.5 EQ.). A reaction chamber is subjected to heating in a microwave oven at 180°C for 40 minutes. The reaction mixture is transferred into a round bottom flask and concentrated. The residue is dissolved in DMSO (1 ml) and purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 10-40% acetonitrile/H2O (0.1% TFA). Pure fractions unite, concentrate, and lyophilizers obtaining colorless solid substances is TBA (1 g, 52%). (MS (ESI) m/z 346 (M++1).

[2-(2-Cyanophenyl)ethyl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid

20-ml reaction vessel for a microwave oven equipped with a bar stirrers, load [2-(2-bromophenyl)ethyl]amidon 2-isopropyl-5-methylcyclohexanecarboxylic acid (1.54 g, 4.2 mmol), CuCN (0,60 g, 6.4 mmol) and NMP (10 ml). A reaction chamber is subjected to heating in a microwave oven at 180°C for 40 minutes. The reaction mixture is transferred into a round bottom flask and concentrated. The residue is purified using flash chromatography on silica gel (10% mixture of ethyl acetate/hexane for elution) to give a colorless solid (1.25 g, 81%).

[2-2-Aminomethylphenol)ethyl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 34)

A 100-ml round-bottom flask equipped with bar stirrers, load [2-(2-cyanophenyl)ethyl]amidon 2-isopropyl-5-methylcyclohexanecarboxylic acid (1.25 g, 4.0 mmol) and methanol (50 ml). NiCl2(1,14 g, 8,8 mmol) and NaBH4(0.64 g, a 16.8 mmol). NaBH4add in small portions over a period of 30 min and stirred for 1 hour. Added NaBH4(0.20 g) in the reaction mixture and stirred for additional 20 minutes. The reaction mixture was passed through a layer of celite and concentrate under reduced pressure. OS is atok black dissolved in a minimum amount of acetonitrile and passed through the cartridge with silica gel C18, and purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 10-40% acetonitrile/H2O (0.1% TFA). Pure fractions unite, concentrate, and lyophilizers obtaining a colourless solid (1.1 g). MS (ESI) m/z 317 (M++1).

Example 9

Synthesis of additional compounds of formula I-C

This example describes a methodology for the synthesis of modulators of Trp-p8 formula I-C.

Scheme 7

(2-Oxo-2-phenylethyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid

20-ml round-bottom flask equipped with bar stirrers, load (2-hydroxy-2-phenylethyl)amidon 2-isopropyl-5-methylcyclohexanecarboxylic acid (100 mg, 0.33 mmol) and acetic acid (1 ml). To the reaction mixture slowly add a solution of CrO3(36 mg, 0,363 mmol, 1.1 EQ.) in acetic acid (500 μl) and water (100 μl). The reaction mixture was stirred at ambient temperature for 15 min and diluted with ethyl acetate (30 ml) and a saturated solution of NaHCO3(aq.) (30 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×10 ml). The combined organic phases are washed with saturated. NaHCO3(3×10 ml), H2O (10 ml), saturated salt solution (10 ml), dried over Na2SO4filter and will contentresult under reduced pressure. The residue is purified using flash chromatography on silica gel (mixture of 30-50% ethyl acetate/hexane for elution) to obtain the specified title compound as a white solid (92 mg, 93%).

(2-Amino-2-phenylethyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid

25-ml reaction vessel for a microwave oven equipped with a bar stirrers, load (2-oxo-2-phenylethyl)amidon 2-isopropyl-5-methylcyclohexanecarboxylic acid (80 mg) and ammonium (1.5 ml, 7M in methanol). A drop of acetic acid and NaCNBH3(20 mg) are added to the reaction mixture and subjected to heating in a microwave oven at 80°C for 80 minutes, the Residue extracted into ethyl acetate (30 ml) and saturated NaHCO3(aq.) (30 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×10 ml). The combined organic phases are washed with saturated. NaHCO3(3×10 ml), H2O (10 ml), saturated salt solution (10 ml), dried over Na2SO4filter and concentrate under reduced pressure to obtain a solid (75 mg).

Tert-butyl methyl ether (1-{2-[(2-isopropyl-5-methylcyclohexanone)amino]-1-phenylethanol}ethyl) - carbamino acid

15-ml reaction vessel equipped with a bar mixers, loading the fair THF (15 ml), Boc-(R)-alanine (52 mg, 0,273 mmol), HOBt (37,87 mg, 0,273 mmol), EDCI (53 mg, 0,273 mmol) and TEA (37 μl, 0,273 mmol). The reaction mixture was stirred for 15 min, while adding (2-amino-2-phenylethyl)amide 2-isopropyl-5-methylcyclohexanecarboxylic acid (75 mg, 0,248 mmol) and stirred for additional 3 hours. To the reaction mixture are added ethyl acetate (10 ml) and H2O (10 ml). A heterogeneous mixture is transferred into a separating funnel where the aqueous phase is separated and re-extracted with ethyl acetate (2×10 ml). The combined organic phases are washed with 1 N. HCl (2×10 ml), H2O (1×10 ml), saturated NaHCO3(3×10 ml), saturated salt solution (10 ml), dried over MgSO4filter and concentrate under reduced pressure. The residue is purified using flash chromatography on silica gel (20% mixture of ethyl acetate/hexane for elution) to obtain the specified title compound as a colourless solid (30 mg).

The TFA salt of [2-(2-aminodiphenylamine)-2-phenylethyl]amide of 2-isopropyl-5-methylcyclohexanecarboxylic acid (compound No. 28)

5-ml round-bottom flask equipped with bar stirrers, load 10% TFA/CH2Cl2and tert-butyl ester (1-{2-[(2-isopropyl-5-methylcyclohexanone)amino]-1-phenylethanol}ethyl) - carbamino acid (30 mg) and stirred for 1 hour. TFA is removed when p is low pressure and the residue is dissolved in 30% mixture of acetonitrile/H 2O (0.1% TFA) and purified using preparative HPLC (Ultro 120 (10 μm) C18Q) using a gradient of 10-40% acetonitrile/H2O (0.1% TFA). Pure fractions unite, concentrate, and lyophilizer to obtain a colorless solid matter (to 17.7 mg). MS (ESI) m/z 374 (M++1).

Example 10

Expression of Trp-p8 cells CHO

Cells CHO man, transfetsirovannyh Trp-p8 (referred to here as CHO/Trp-p8), are generated for use in experiments by the present invention. The expression of the polypeptide Trp-p8 in this transfectants and in the absence of any adokenai expression in nitrostilbene CHO confirmed by analysis of Western blot and immunofluorescence assay using antibodies specific for Trp-p8 (GS2.20), and flow analysis of calcium with Icilin (1-[2-hydroxyphenyl]-4-[3-nitrophenyl]-1,2,3,6-tetrahydropyrimidin-2-it) and menthol (2-isopropyl-5-methylcyclohexanol). Nitrostilbene CHO cells are used to establish the specificity of the effects of compounds observed with CHO/Trp-p8.

Example 11

Mediaremote Trp-p8 decrease in the survival rate of cells after exposure of cells CHO/Trp-p8 for candidate compounds in agonists Trp-p8 at 37°C

This example describes a survival analysis using ATP suitable for screening effective agonists Trp-p8. Analysis vyzyvaet the use of ATP described here uses CHO cells expressing exogenous cDNA Trp-p8. This example further provides that agonists Trp-p8 of the present invention are effective in reducing the survival of cells expressing Trp-p8.

The intracellular concentration of ATP decreases very quickly when metabolically active cells undergo necrosis and/or apoptosis. The concentration of ATP and, as a consequence, the relative cell survival can be measured by established methods using commercially available reagents. In the methodology of screening for agonists described here, the connection that specific reduces cell survival CHO/Trp-p8, referred to as agonist.

As the primary screening for the efficiency and specificity for agonist, as nitrostilbene CHO, and cells CHO/Trp-p8 are exposed to 1 or 10 μm of these compounds in 1% dimethyl sulfoxide (DMSO) or 1% DMSO (control) in 96-well-plate with black walls, black bottom, processed for cell culture. DMSO is a solvent for all the investigated compounds. After 24-26 hours at 37°C the cells lyse and the concentration of ATP is determined using chemiluminescent analysis using commercially available reagents - Cell Titer-Glo (Promega; Madison, WI). Relative is the survival rate (%), expressed as ATP levels in cells treated with compounds, which is expressed as a percentage of ATP levels in cells treated only DMSO, is a measure of the activity of the connection candidate for agonists than % survival rate, the more potent agonist is Trp-p8. The values of EC50are determined for the most active candidates agonists Trp-p8 at 37°C by measuring the survival rate at 8-10 different concentrations of agonists. (EC50is defined here as the concentration of agonist, with a 50% reduction in the relative survival of cells).

Examples of agonists Trp-p8 formula I, formula I-A, formula I-B, formula I-C, formula I-D, and formula I-E that are effective in the analysis of survival with the help of ATP, presented here in tables 1-5, the data EC50are indicated as follows: A = <0,020 μm; B = 0,021-0,050 μm; C = 0,051-0,10 mm.

In tables 1-5 summarizes the patterns are of the form:

Where chemical names are given for X and/or Y. where names are for "X/Y", names are names, with the inclusion of the group of nitrogen.

Cell survival CHO/Trp-p8 after treatment illustrative agonists Trp-p8 is presented in figure 1.

Table 1
Examples of compounds of formula I-A
Conn. No.StructureEC50X
1A2-(2-Aminodiphenylamine)-4-methoxyphenyl
2AN-(2-amino-ethyl)-2-amino-5-methylsulfinylphenyl
3A1-(2-Aminoethoxy)naphthalene-2-yl
4And2-(2-Aminoethylamino)-4-methylsulfinylphenyl
5AndN-(2-amino-ethyl)-5-methoxybenzamide
6In2-(2-Aminoethylamino)-4-methoxime the Il
7In2-(2-Amino-3-hydroxypropylamino)-4-methoxyphenyl
8In3-(2-Aminoethylamino)naphthalene-2-yl
9InN-(2-amino-ethyl)-2-aminobenzamide
10In2-(2-Amino-3-hydroxypropylamino)-4-methoxyphenyl
112-(2-Aminoethylamino)phenyl
122-(2-Amino-3-hydroxybutyrate)-4-methoxybenzylamine
132-(2-Aminoethylamino)-4-methoxyphenyl

Table 2
Examples of compounds of formula I-B
Conn. No.StructureEC50X
14A2-(2-amino-ethyl)-5-furan-2-yl-2H-pyrazole-3-yl
15And2-(2-Aminopropyl)-5-phenyl-2H-pyrazole-3-yl
16And2-(2-amino-ethyl)-5-phenyl-2H-pyrazole-3-yl
17And2-(2-amino-ethyl)-5-(1-methyl-1H-pyrrol-2-yl)-2H-pyrazole-3-yl
18And2-(2-Aminopropyl)-5-phenyl-2H-pyrazole-3-yl
19In 2-(2-amino-ethyl)-5-(4-AMINOPHENYL)-2H-pyrazole-3-yl
20In2-(2-amino-ethyl)-5-(4-hydroxyphenyl)-2H-pyrazole-3-yl
21In2-(2-Methylaminomethyl)-5-phenyl-2H-pyrazole-3-yl
22In2-(2-Aminopropyl)-5-phenyl-2H-pyrazole-3-yl
23In2-(2-amino-ethyl)-5-(3-cyanophenyl)-2H-pyrazole-3-yl
24In2-(2-amino-ethyl)-5-(3-methoxyphenyl)-2H-pyrazole-3-yl
25Methyl ester of 4-{1-(2-amino-ethyl)-1H-pyrazole-3-yl}benzoic acid
26 2-(2-amino-ethyl)-5-(3-AMINOPHENYL)-2H-pyrazole-3-yl
272-(2-amino-ethyl)-5-(3-hydroxyphenyl)-2H-pyrazole-3-yl

Table 3
Examples of compounds of formula I-C
Conn. No.StructureEC50X
28A2-(2-Aminodiphenylamine)-2-phenylethyl
29And2-(2-Aminoethoxy)-2-phenylethyl
30And2-(2-Aminoethoxy)-2-phenylethyl
31In2-(3-Aminopropoxy)-2-phenylethyl
32 2-(2-Aminoethylamino)-2-phenylethyl

Table 4
Examples of compounds of formula I-D
Conn. No.StructureEC50X
33A2-[2-(2-Aminoethylamino)phenyl]ethyl
34And2-(2-Aminomethylphenol)ethyl
35In2-[(2-Aminoacetyl)phenylamino]ethyl

3-(2-amino-ethyl)-5-methylsulfanyl-1,3-dehydrobenzperidol-2-he
Table 5
Examples of compounds of the formula T-E
Conn. No.StructureEC50X/Y
36A3-(2-amino-ethyl)-5-methoxy-1,3-dehydrobenzperidol-2-he
37And3-(2-amino-ethyl)-5-(3-hydroxypropoxy)-1,3-dehydrobenzperidol-2-he
38And3-(2-amino-ethyl)-5-ethoxy-1,3-dehydrobenzperidol-2-he
39And3-(2-amino-ethyl)-5-methanesulfonyl-1,3-dehydrobenzperidol-2-he
40And3-(2-amino-ethyl)-5-(2-hydroxyethoxy)-1,3-dehydrobenzperidol-2-he
41AndAmide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
42And
43And3-(2-amino-ethyl)-5-methanesulfonyl-1,3-dehydrobenzperidol-2-he
44And(2-Diethylaminoethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
45And3-(2-Aminopropyl)-2,3-dehydrobenzperidol-2-he
46And[3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-yloxy]acetonitrile
46AndEthylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
48AndPyridine-3-alamid 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-shall benzoimidazol-5-carboxylic acid
49And(2-Methoxyethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
50And1-(2-amino-ethyl)-1,3-dehydrobenzperidol-2-he
51And1-(2-amino-ethyl)-1,3-dihydronaphtho[2,3-d]imidazol-2-he
52And(2-Hydroxyethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
53And3-(2-amino-ethyl)-5-propoxy-1,3-dehydrobenzperidol-2-he
54In3-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-c]pyridine-2-he
55 In(2-Diethylaminoethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-4-carboxylic acid
56InPyridine-4-alamid 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
57In3-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-b]pyridine-2-he
58In1-(3-Aminopropyl)-1,3-dehydrobenzperidol-2-he
59InPhenylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
60In[2-(2-Hydroxyethoxy)ethyl]amide of 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
61 In1-(2-amino-ethyl)-5-trifluoromethyl-1,3-dehydrobenzperidol-2-he
62In1-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-c]pyridine-2-he
63InBenzylated 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
64In3-(2-amino-ethyl)-5-(morpholine-4-carbonyl)-1,3-dehydrobenzperidol-2-he
65In3-(2-amino-ethyl)-5-(2-oxo-2-phenylethane)-1,3-dehydrobenzperidol-2-he
66In3-(2-Methylaminomethyl)-1,3-dehydrobenzperidol-2-he
673-(2-amino-ethyl)-5-butoxy-1,3-dehydrobenzperidol-2-he
68Methylphenylene 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
69Ethyl ester of 4-[3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carbonyl]piperazine-1-carboxylic acid
70Diethylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
71Ventilated 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
72(2-Hydroxy-1-hydroxymethyl-2-phenylethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
73Carbamoylmethyl 3-(2-what aminoethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
74(2-Hydroxy-1-hydroxymethylation)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid
753-(2-amino-ethyl)-5-benzyloxy-1,3-dehydrobenzperidol-2-he
761-(4-aminobutyl)-1,3-dehydrobenzperidol-2-he

Example 12

Screening and characterization of compounds-agonists Trp-p8 by measuring the influx of calcium into the cells of CHO/Trp-p8 at 37°C

This example describes the analysis of the inflow of calcium-based CHO/Trp-p8 used to further assess the activity of candidates agonists Trp-p8 of the present invention.

Calcium influx was measured using a Flexstation Microplate Fluorescent Plate Reader (Molecular Devices; Sunnyvale, CA). Typical flow analysis of calcium is carried out as follows. Cells in the medium based on DMEM/ham F-12, typically at a density of 30,000 cells/well/100 ál, placed in a 96-well plate for tissue culture with black wall, clear bottom (Greiner io-one) and then incubated for 16-20 hours at 37°C. Cells in each well is then incubated for one hour at 37°C with a mixture of Fura2-AM Fluorescent Dye/Pluronic F-27 (Molecular Probes; Eugene, Oregon) and dissolved in a medium containing Probenecid. Typical final concentrations are: 5-8 μm Fura2-AM, in 0.01% Pluronic F-27 and 2.5 mm Probenecid (an inhibitor of anion exchange, which reduces the transport hydrolyzed dye out of the cell, thereby minimizing the loss of dye during the experiment). After one hour the cells are washed in buffer solution (20 mm HEPES balanced salt solution Hanks with 1,26 mm CaCl2), a pH of 7.4, containing Probenecid at final concentration of 2.5 mm, and pre-incubated for at least 30 minutes at a temperature analysis 37°C.

Typically, the above-described buffer on the basis of HEPES/HBSS or not containing extra calcium or calcium to increase the concentration up to 2 mm, and with various concentrations of compounds (5-fold final concentration) is added to each well using a multichannel robotic device for epatirovanie. Connection pre-incubated at 37°C for at least 30 minutes before performing the analysis (at 37°C). The signals recorded in dual excitation at wavelengths of 340 and 380 nm and at the wavelength of emission of 510 nm with a bandpass filter at 495 nm. The signal is expressed as the ratio which their emission, when the excitation occurs at 340 nm, emission when the excitation occurs at 380 nm [Relative Fluorescence Units (RFU)]. The ionomycin routinely used as a positive control.

In the case of the analysis of agonist compounds at various concentrations is added to the loaded dye cells (as described above). Increase max is a measure saledate compounds as agonist. The estimated results are presented in figure 2.

Example 13

The increase in apoptosis after exposure of cells CHO/Trp-p8 for connections-agonists Trp-p8 at 37°C

This example describes the effectiveness of compounds for agonist Trp-p8 the induction of apoptosis in cells expressing Trp-p8.

Running cytometrics analysis using Annexin V/Propery iodide (PI) used for more information on the mechanism of cell death induced by compounds-agonists Trp-p8. Staining with Annexin V detects translocation of phosphatidylserine in the outer layer of the plasma membrane, characteristic events of apoptosis, whereas staining with PI shows the death of cells with compromised membranes.

Cells treated with compounds in 1% DMSO or 1% DMSO (control) for 24 to 26 hours at 37°C. Cells quickly trypsinized under controlled conditions and are colored by the th set of reagents Annexin V/PI, following the methodology provided by the supplier (for example, Southern Biotech; Birmingham, Alabama). The estimated results are presented in figure 3.

Example 14

The in vitro screening of compounds antagonists Trp-p8 using survival analysis cells on the basis of the protection of cells expressing Trp-p8 for toxic compounds-agonists

This example describes the system analyses for the identification and characterization of candidate compounds in antagonists Trp-p8.

Antagonists Trp-p8 are identified through the use of survival analysis of cells using the cells of CHO/Trp-p8 at 37°C (see example 11) with the following modifications. In the context of the present invention compounds that protect cells CHO/Trp-p8 from the toxic effect of the control agonist, thereby supporting cell survival CHO/Trp-p8, exposed to agonists Trp-p8, defined as antagonists. As the primary screening for antagonists cells CHO/Trp-p8 are exposed to 10 μm of the compounds in 1% dimethyl sulfoxide (DMSO) or 1% DMSO plus toxic concentration control agonist. The relative survival rate at 10 microns, determined as described in example 11, is a measure of the potential compound as an antagonist of Trp-p8 - the higher the survival rate, the more potent is antagon the Art. The estimated results are presented in figure 4.

Example 15

Screening in vitro using flow analysis of calcium compounds-antagonists Trp-p8 on the basis of their ability to suppress the inflow of calcium induced by agonists Trp-p8, cells CHO/Trp-p8

This example describes the system in vitro used for additional screening and characterization of candidates antagonists Trp-p8.

Antagonists Trp-p8 also examined and characterized using flow analysis of calcium at 37°C as described in example 12, with the following two differences: (1) the connection is pre-mixed with control agonist or only the control agonist is added to the cells, and suppression of the response to the agonist is a measure saledate compounds as the antagonist, and (2) the compound at various concentrations are added to the cells followed by the addition of the control agonist after 2-3 minutes, and suppression of the response induced by agonist, is a measure saledate compound as an antagonist. The estimated results are presented in figure 5.

Example 16

The system of animal models for the analysis of in vivo efficacy candidates for agonists and antagonists of Trp-p8 for cancer treatment

This example causes the system of animal models suitable on what I determine the effectiveness of in vivo candidate modulators of Trp-p8, including both agonists and antagonists.

Grafted tumors prostate cancer person expressing Trp-p8 (LuCaP, from Dr. Robert Vessella''s lab at the University of Washington estimated using in situ hybridization, quantitative the polymerase chain reaction and immunohistochemistry using polyclonal rabbit antibodies specific to proteins, T-904), as well as cell lines, obtained using genetic engineering to ekspressirovali Trp-p8, which includes cell line CHO (Chinese hamster ovary) and EL-4 Thymoma mice), used to establish the model for tumors of mice. Expressiona Trp-p8 in the transfectants confirmed by analysis of Western blot and immunofluorescence assay using antibodies (GS 2,20), specific for Trp-p8, as well as through response to known agonists in functional analysis influx of calcium. In addition to this line transfected cells are sensitive to destruction under the action of agonists Trp-p8, as shown by the analyses of survival with the help of ATP and apoptosis (as described in examples 11 and 13).

Model tumors in mice was determined by subcutaneous injection of cells CHO/Trp-p8 mice SCID. Expression of Trp-p8 in the tumor, isecheno of these mice confirmed using RT-PCR, immunohistochemistry and analysis of Western blot. Further development of the model tumor realizes what I'm using grafted tumors prostate cancer man described above, animicheskih naked mice or SCID mice and using transfectants EL4/Trp-p8 in normal mice. Grafted prostate tumors from other sources and other cell lines that can be obtained using genetic engineering to ekspressirovali Trp-p8, are also potential candidates for building additional systems models.

Based on the results of the evaluation in vitro and in vivo will be the selected set agonists Trp-p8 to determine the efficacy in mice. The in vitro evaluation should include selenocystine in the analysis of cell death, solubility in water, research associate in the plasma and metabolic stability (the ability to metabolize compounds in the liver, as determined by the use of hepatocytes and/or microsomes of mice). The in vivo evaluation should include studies of the pharmacokinetics and toxicity. Selected compounds will be administered to mice with tumors expressing Trp-p8, using different methods [orally, intravenously, intraperitoneally, subcutaneously, intramuscularly]. The reduction of tumors and survival of these mice will be evaluated at different doses of these compounds. Compounds that are most effective against tumors, will be selected for additional studies.

Example 17

Experimental characterization nekotorykh connections

This example describes the experimental characterization and the results for several exemplary low molecular weight modulators of Trp-p8 formula I, which indicate the connection I, II, III and IV. Their chemical formula and molecular weight are given in table 6.

Table 6
The chemical formula and molecular weight
ConnectionIIIIIIIV
Chemical formulaC21H31N3O3C20H29N3O2C22H32N4OC21H34N2O2
Molecular weight373343368346

Activity in vitro

As shown in table 8, the compounds show a high degree selenolate and specificity in relation to the death of cells that Express Trp-p8. Usually >1000x higher concentration of the texts required for cell death, where there is no Trp-p8, compared with cells that Express Trp-p8, Compounds II, III and IV show similar activity in this assay, whereas Compound I is about 3 times more potent.

Table 8
The results of the analysis using ATP survival for several preferred compounds
ConnectionIIIIIIIV
EC50CHO/Trp-p8
(µm)
0,0030,010,010,01
EC50the original CHO
(µm)
>10>10>10>10

Activity in vivo

As illustrated in figa and 9B, the compounds I, II and III provide a prolonged exposure after a single oral administration as in rodents (figa)and Beagle dogs (pigv). Compared with mice, rats require approximately double oral dose (relative to body weight) to achieve the CPA is removable platen, and dogs require less than a third dose. In accordance with the supported levels in plasma (t1/2 ~9 hours) one oral dose provides prolonged persistent response for the model grafted tumors CHO/Trp-p8.

As illustrated in figa and 10B, oral dosing of these compounds provides a strong reaction for the model grafted tumors CHO/Trp-p8. Significant inhibition of tumor growth was observed after a single dose, not more than 10 mg/kg, and little toxicity was observed at 100 mg/kg; therapeutic window >10x.

As illustrated in Fig.7, the compounds of formula I generate a significantly shorter exposure with a single intraperitoneal injection compared with oral administration. As illustrated in figa and 8B, intraperitoneal (IP) injection of a mouse these compounds leads to shorter reaction model grafted tumors CHO/Trp-p8 and looks less persistent after cessation IP dosing.

To demonstrate that the effectiveness mediasuite through Trp-p8, 11 illustrates the evaluation of compound I in a coherent model of CHO (Trp-p8-). In accordance with the proposed mechanism of action of compound I does not show significant efficacy at 100 mg/kg for this model; this dose 10 times higher than the effective dose of the same model CHO/Trp-p8.

As illustrated in Fig, model LuCP apparently reacts comparable or slightly better what is the model of CHO/Trp-p8. CHO/Trp-p8 is a fast-growing tumor; treatment with compound I slows the growth, but does not cause regression. In contrast, LuCaP is a slow-growing tumor; treatment causes a statistically significant regression and growth inhibition. In this case, the model grafted tumor LuCAP shows the levels of Trp-p8, comparable with the model of CHO/Trp-p8, as measured by immunohistochemistry of tumor tissue, isecheno of the mouse.

The highest dose, oral injected mice, 100 mg/kg, does not lead to significant toxicity for any of the compounds. Because a single oral dose of 10 mg/kg of compound I provides a significant efficiency for the model grafted tumor CHO/Trp-p8, a therapeutic window >10 is achieved by the compounds of formula I. It expands with Toxicological experiments on rats, where connections could be implemented at higher levels of dosing. In Toxicological studies carried out on rats, oral doses of 250 mg/kg does not induce any observable toxic effects. A single dose of 500 mg/kg and 1000 mg/kg leads to toxicity in the range of mild to moderate, but not reaching the MTD. These data, representing the minimum therapeutic window, obtained by using the compounds of formula I, are given in the table is e 7.

Table 7
Oral dose for rats (mg/kg)Observed
toxicity
Comparable oral dose for mice (mg/kg)Multiple the smallest effective oral dose for mice
250no12512,5
500light25025
1000moderate50050

Although the present invention is described in detail in the form of illustrations and examples for purposes of clarity of understanding, changes and modifications may be made without deviating from the scope of the present invention, which is limited only by the framework of the attached claims.

1. The compound of formula I:

or its pharmaceutically acceptable salt, where R1represents H, or R1and R2together with a group of nitrogen may form

where a, b, C and D independently are selected from the group consisting of CR1aand N; where at least one of a, b, C and D represents CR1a; where R1aselected from the group consisting of H, -ORi, -SRii, -S(O)Riii, -C(O)NRvRviand CF3where Riselected from the group consisting of methyl, ethyl, propyl, hydroxyethyl, hydroxypropyl, 2-oxo-2-phenylethyl, butyl, acetonitrile, and benzyl; Rii, Riiiand Rivrepresents methyl; Rvand Rviindependently selected from the group consisting of H, methyl, ethyl, hydroxyethyl, hydroxypropyl, diethylaminoethyl, phenyl, pyridinyl, methoxyethyl, hydroxyethoxyethyl, benzyl, phenylethyl, 2-hydroxy-1-hydroxymethyl-2-phenylethyl and carbamoylmethyl, or Rvand Rvitogether form a morpholine or complex ethyl ester piperazine;
R2selected from the group consisting of phenyl, naphthyl, pyrazolyl and C1-C8alcelaphine;
R3represents a C1-C8alkylen;
R4selected from the group consisting of H, C1-C8the alkyl and-C=NH(NH2).

2. The compound of formula I-A:

or its pharmaceutically acceptable salt, where a, b, C and D represent CR2; where R2independently represents a residue selected from the group consisting of H, R6O - and Rsub> 6S-, where R6represents a C1-C8alkyl; or two adjacent R2can be combined with the formation of a single phenyl group; and
R1represents H;
R3selected from the group consisting of-NR7C(O)-, -C(O)NR7-, -O - and-NR7-, where R7represents H;
R4represents a C1-C8alkyl and hydroxys1-C8alkyl; and
R5represents N.

3. The compound according to claim 2 or its pharmaceutically acceptable salt, where
R6represents 1, 2, 3, 4, 5 or 6 carbon atoms.

4. The compound according to claim 2 or its pharmaceutically acceptable salt, where balance

selected from the group consisting of 2-(2-aminodiphenylamine)-4-methoxyphenyl, N-(2-amino-ethyl)-2-amino-5-methylsulfinylphenyl, 1-(2-aminoethoxy)naphthalene-2-yl, 2-(2-aminoethylamino)-4-methylsulfinylphenyl, N-(2-amino-ethyl)-5-methoxybenzamide, 2-(2-aminoethylamino)-4-methoxyphenyl, 3-(2-aminoethylamino)naphthalene-2-yl, N-(2-amino-ethyl)-2-aminobenzamide, 2-(2-aminoethylamino)phenyl and 2-(2-aminoethylamino)-4-methoxyphenyl.

5. The compound of formula I-B:

or its pharmaceutically acceptable salt, where R1represents H;
R2represents a substituted or unsubstituted phenyl, benzoyl, furanyl or pyrrolyl;
Rsub> 3represents a C1-C8alkyl;
R4selected from the group consisting of N and C1-C8the alkyl.

6. The compound according to claim 5 or its pharmaceutically acceptable salt, where R3represents 1, 2, 3, 4, 5 or 6 carbon atoms, or R2represents a group selected from the group consisting of phenyl, furan, methylpyrrole, methylbenzoate, AMINOPHENYL, hydroxyphenyl, cyanophenyl and methoxyphenyl.

7. The compound of formula I-C:

or its pharmaceutically acceptable salt, where R1represents H;
R2represents phenyl;
R3selected from the group consisting of-NHC(O)R5-, -OR5and other5-, where R5represents a C1-C8alkyl; and
R4represents N.

8. The connection according to claim 7 or its pharmaceutically acceptable salt, where R2represents phenyl, or R5selected from the group consisting of methylene, ethylene, propylene and butylene.

9. The compound of formula I-D:

or its pharmaceutically acceptable salt, where R1represents H;
R2represents phenyl or phenylamino;
R3selected from the group consisting of methylene, ethylene, methylamino, ethylamino and acetyl; and
R4represents N.

10. The compound according to claim 5, or the th pharmaceutically acceptable salt, where R4selected from the group consisting of H and methyl.

11. The compound of formula I-E:

or its pharmaceutically acceptable salt, where a, b, C and D independently are selected from the group consisting of CR1and N; where at least one of a, b, C and D represents CR1; where R1selected from the group consisting of H, -ORi, -SRii, -S(O)Riii, -S(O)2Riv-C(O)NRvRviand CF3where Riselected from the group consisting of methyl, ethyl, propyl, hydroxyethyl, hydroxypropyl, 2-oxo-2-phenylethyl, butyl, acetonitrile, and benzyl; Rii, Riiiand Rivrepresents methyl; Rvand Rviindependently selected from the group consisting of H, methyl, ethyl, hydroxyethyl, hydroxypropyl, diethylaminoethyl, phenyl, pyridinyl, methoxyethyl, hydroxyethoxyethyl, benzyl, phenylethyl, 2-hydroxy-1-hydroxymethyl-2-phenylethyl and carbamoylmethyl, or Rvand Rvitogether form a morpholine or complex ethyl ester piperazine; and when two adjacent remnant of a, b, C and D represent CR1two R1can be combined with the formation of a single phenyl group;
R2represents a C1-C8alkylen; and
R3represents H, C1-C8alkyl or-C=NH(NH2).

12. The connection 11 or f is matemticas acceptable salt, where R1represents H or methoxy.

13. The connection 11 or its pharmaceutically acceptable salt, where R1represents a
(a) ORiand where Riselected from the group consisting of methyl, ethyl, propyl, hydroxyethyl, hydroxypropyl, 2-oxo-2-phenylethyl, butyl, acetonitrile, and benzyl; or optional
(b) -S(O)Riiiand where Riiirepresents methyl;
(c) -S(O)2Rivand where Rivrepresents methyl;
(d) -C(O)NRvRviwhere Rvand Rviindependently selected from the group consisting of H, methyl, ethyl, hydroxyethyl, hydroxypropyl, diethylaminoethyl, phenyl, pyridinyl, methoxyethyl, hydroxyethoxyethyl, benzyl, phenylethyl, 2-hydroxy-1-hydroxymethyl-2-phenylethyl and carbamoylmethyl;
(e) -C(O)NRvRviwhere Rvand Rvitogether form a morpholine or ethyl ester piperazine;
(f) CF3; or
R2selected from the group consisting of methylene, ethylene, propylene and butylene; or
R3represents N.

14. A method of treating diseases associated with receptor Thr-R8 in a mammal, comprising the introduction of a combination of compounds according to claim 1 of the formula I according to claim 2 of formula I-A according to claim 5 of the formula I-B according to claim 7 of formula I-C, according to claim 9 of the formula I-D or according to claim 11 of formula 1, or a combination thereof, or a pharmaceutical the ski acceptable salt and one or more therapeutic agents against cancer; or where the specified combination is entered at the same time.

15. Pharmaceutical composition having modulating Thr-R8 activity containing a compound according to claim 1 of the formula I according to claim 2 of formula I-A according to claim 5 of the formula I-B according to claim 7 of formula I-C, according to claim 9 of the formula I-D or according to claim 11 of formula 1, or a combination thereof, or their pharmaceutically acceptable salt and a pharmaceutically acceptable excipient; and optionally a pharmaceutically acceptable carrier or agent.

16. The connection method selected from the group consisting of cancer, induction of apoptosis and/or necrosis in cells expressing Thr-R8, decrease the survival rate of cells and/or inhibiting cell growth, treatment of a disease in a mammal, medical diagnosis or therapy, treatment of a disease or condition in a mammal, which shows the involvement of receptors Thr-R8, and is a desirable modulation functions of the receptors, and modulation Thr-R8 functions of receptors and treatment of diseases associated with expression of Thr-R8, and a compound selected from the group consisting of compounds according to claim 1 of formula I, according to claim 2 of formula I-A according to claim 5 of the formula I, according to claim 7 of formula I-C, according to claim 9 of the formula I-D or according to claim 11 of formula I-E, or combinations thereof, or their pharmaceutically acceptable salts.

17. A method of identifying agonists Thr-R8, which is includes a stage of contacting cells, expressing Thr-R8, and cells not expressing Thr-R8, with a compound selected from the group consisting of compounds according to claim 1,formula I, according to claim 2,of formula I-A according to claim 5,of formula I-B according to claim 7,formula I-C, according to claim 9,formula I-D or according to claim 11 of formula 1, or its pharmaceutically acceptable salts within the time and in a quantity sufficient to reduce the survival of these cells expressing Thr-R8, but not these cells not expressing Thr-R8.

18. The compound according to claim 5 or its pharmaceutically acceptable salt, where balance

selected from the group consisting of 2-(2-amino-ethyl)-5-furan-2-yl-2H-pyrazole-3-yl, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(1-methyl-1H-pyrrol-2-yl)-2H-pyrazole-3-silt, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(4-AMINOPHENYL)-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(4-hydroxyphenyl)-2H-pyrazole-3-yl, 2-(2-methylaminomethyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-aminopropyl)-5-phenyl-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(3-cyanophenyl)-2H-pyrazole-3-yl, 2-(2-amino-ethyl)-5-(3-methoxyphenyl)-2H-pyrazole-3-yl, methyl ester 4-{1-(2-amino-ethyl)-1H-pyrazole-3-yl}-benzoic acid, 2-(2-amino-ethyl)-5-(3-AMINOPHENYL)-2H-pyrazole-3-yl and 2-(2-amino-ethyl)-5-(3-hydroxyphenyl)-2H-pyrazole-3-Il.

19. The connection according to claim 7 or its pharmaceutically acceptable salt, where br/>
selected from the group consisting of 2-(2-aminodiphenylamine)-2-phenylethyl, 2-(2-aminoethoxy)-2-phenylethyl, 2-(2-aminoethoxy)-2-phenylethyl, 2-(3-aminopropoxy)-2-phenylethyl and 2-(2-aminoethylamino)-2-phenylethyl.

20. The connection according to claim 9, or its pharmaceutically acceptable salt, where balance

represents 2-[2-(2-aminoethylamino)phenyl]ethyl, 2-(2-aminomethylphenol)ethyl and 2-[(2-aminoacetyl)phenylamino]ethyl.

21. The connection 11 or its pharmaceutically acceptable salt, where balance

selected from the group consisting of 3-(2-amino-ethyl)-5-methoxy-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-(3-hydroxypropoxy)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-ethoxy-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-methanesulfonyl-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-(2 hydroxyethoxy)-1,3-dehydrobenzperidol-2-it, amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-5-methylsulfanyl-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-methanesulfonyl-1,3-dehydrobenzperidol-2-it, (2-diethylaminoethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-aminopropyl)-2,3-dehydrobenzperidol-2-it, [3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-yloxy]acetonitrile, ethylamide 3-(2-amino-ethyl)--oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, pyridine-3-ylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-methoxyethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 1-(2-amino-ethyl)-1,3-dehydrobenzperidol-2-it, 1-(2-amino-ethyl)-1,3-dihydronaphtho[2,3-d]imidazol-2-it, (2-hydroxyethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-5-propoxy-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-C]pyridine-2-(2-diethylaminoethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-4-carboxylic acid, pyridine-4-ylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-b]pyridine-2-it, 1-(3-aminopropyl)-1,3-dehydrobenzperidol-2-it, phenylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, [2-(2-hydroxyethoxy)ethyl]amide of 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 1-(2-amino-ethyl)-5-trifluoromethyl-1,3-dehydrobenzperidol-2-it, 1-(2-amino-ethyl)-1,3-dihydroimidazo[4,5-C]pyridine-2-it, benzylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, 3-(2-aminoethyl)-5-(morpholine-4-carbonyl)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-(2-oxo-2-phenylethane)-1,3-dehydrobenzperidol-2-it 3-(2-methylaminomethyl)-1,3-dehydrobenzperidol-2-it 3-(2-amino-ethyl)-5-butoxy-1,3-dihydro Intimidator-2-it, methylphenylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, ethyl ester 4-[3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carbonyl]piperazine-1-carboxylic acid diethylamide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, phenetylamine 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-hydroxy-1-hydroxymethyl-2-phenylethyl)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, carbamoylmethyl 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid (2-hydroxy-1-hydroxymethylation)amide 3-(2-amino-ethyl)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid, N-{2-[2-oxo-2,3-dihydrobenzoic-1-yl]ethyl}guanidine, 3-(2-amino-ethyl)-5-benzyloxy-1,3-dehydrobenzperidol-2-it 1-(4-aminobutyl)-1,3-dehydrobenzperidol-2-it.

22. The connection 11 or its pharmaceutically acceptable salt, where the specified connection is a 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-5-methoxy-1,3-dehydrobenzperidol-2-it.

23. The compound 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-5-methoxy-1,3-dehydrobenzperidol-2-one or its pharmaceutically acceptable salt.

24. The amide compound 3-(2-amino-ethyl)-1-(2-isopropyl-5-methylcyclohexanone)-2-oxo-2,3-dihydro-1H-benzoimidazol-5-carboxylic acid or E. what about the pharmaceutically acceptable salt.

25. The compound according to any one of p, 23 or 24 or its pharmaceutically acceptable salt in isolated and purified form.



 

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22 cl, 10 tbl, 11 ex

FIELD: chemistry.

SUBSTANCE: invention relates to a method of producing novel 2-(2-acylvinyl)indole derivatives of formula:

characterised by that 2-azidobenzylfurans are boiled in high-boiling organic solvents, such as bromobenzene or chlorobenzene or para-xylene or ortho-dichlorobenzene or diphenyl oxide or butyl acetate or dimethyl sulphoxide.

EFFECT: obtaining novel 2-(2-acylvinyl)indole derivatives.

2 tbl, 9 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention relates to compound of formula (I): or to its pharmaceutically acceptable ester, amide, carbamate, solvate or salt, including salt of such ester, amide or carbamate and solvate of such ester, amide, carbamate or salt, where values R1, R2, R3, R4, R5 and R6 are given in item of the formula, with the exception: 4-[3-(4,5-dihydro-1H-imidazol-2-yl)-2-(3,5-dimethylisoxazol-4-yl)indole-1-yl]phenol; 1-(4-hydroxyphenyl)-2-(4-methylimidazol-1-yl)-1H-indole-3-carbonitryl; 1-(4-hydroxyphenyl)-2-(1H-pyrazol-3-yl)-1H-indole-3-carbonitryl; 1-(3-chloro-4-hydroxyphenyl)-2-(1-methyl-1H-pyrazol-4-yl)-1H-indole-3-carbonitryl; 1-(4-hydroxyphenyl)-2-prop-1-inyl-1H-indole-3-carboxylic acid amide.

EFFECT: compounds I possess affinity of binding with estrogen receptor of p-subtype, which makes it possible to use them in pharmaceutical composition and in treatment or prevention of state, associated with disease or disorder, associated with activity of estrogen receptors of β-subtype.

27 cl, 271 ex

FIELD: chemistry.

SUBSTANCE: present invention relates to novel alkyl [(S)-1-((S)-2-{5-[4-(4-{2-[(S)-1-((S)-2-methoxycarbonylamino-3-methyl-butyryl)-pyrrolidin-2-yl]-3H-imidazol-4-yl}-buta-1,3-diinyl)-phenyl]-1H-imidazol-2-yl}pyrrolidine-1-carbonyl)-2-methyl-propyl]-carbamate naphthalene-1,5-disulphonates of general formula 1, which are NS5A inhibitors and can be used as an active component for producing a pharmaceutical composition and an antiviral medicinal agent for treating and preventing viral diseases caused by hepatitis C virus (HCV) and hepatitis GBV-C virus. In general formula 1 R denotes C1-C3alkyl.

EFFECT: obtaining novel compounds.

6 cl, 2 tbl, 6 ex

FIELD: chemistry.

SUBSTANCE: invention relates to use of 2-nitroheterylthiocyanates, particularly 4-rhodano-5-nitropyrimidine and 2-rhodano-3-nitripyridine derivatives of general formula (I), optionally in crystalline form or in form of pharmaceutically acceptable addition salts thereof with acids or bases, having activity on fungal strains, fungal infection agents, for producing pharmaceutical compositions that are suitable for local application. The compounds are also active on strains that are resistant to existing drugs. In general formula (I) X=N or C-R3, R1 denotes a proton, a saturated or unsaturated linear alkoxy radical having 1-5 carbon atoms; a cycloalkyloxy radical having up to 6 carbon atoms; a saturated linear alkylmercapto radical having 1-3 carbon atoms; an amino radical having 1-10 carbon atoms, selected from a saturated or unsaturated linear mono- or dialkylamino radical or a cycloalkylamino radical, cyclic amino radical. Each of the cyclic groups can be substituted with 1-2 methyl groups, or a benzylamino group; R2 denotes a proton, a saturated or unsaturated linear alkyl radical having 1-5 carbon atoms, or a cyclic aliphatic radical having up to 6 carbon atoms, trifluoromethyl, styryl or methylmercapto group; R3 denotes a trifluoromethyl, formyl, acetyl, nitro, benzoyl, cyano group or an alkoxycarbonyl substitute having 1-3 carbon atoms in the alkoxy group.

EFFECT: improved properties of compounds.

5 cl, 3 tbl, 21 ex

FIELD: chemistry.

SUBSTANCE: invention relates to novel substituted 6-(1-piperazinyl)-pyridazines of formula , where R1 is chlorine, trifluoromethyl or cyano; R2 is phenyl or phenyl substituted with a halogen; R*1 is hydrogen, C1-4-alkyl or pyridinylmethyl; X is -O-, -NH-, -CH2-, -CH(OH)-, -SO2-, -CO-, -NH-CH2-, -O-CH2-, 1,2-ethendiyl or ethyndiyl; or a pharmaceutically acceptable addition salt or solvate thereof as well as pharmaceutical compositions containing said novel compound as an active ingredient.

EFFECT: improved properties of derivatives.

8 cl, 4 tbl, 6 ex

FIELD: chemistry.

SUBSTANCE: invention relates to substituted quinoxaline-type bridge piperidine compounds of formula

,

or pharmaceutically acceptable derivatives thereof, where: a equals 0; b is an integer selected from 0 or 1; each R5 is independently selected from -H; R1 is -(C9-C14)bicycloalkyl, each substituted with 1 or 2 independently selected R3 groups; each R3 independently selected from -(C1-C4)alkyl. The invention also relates to a pharmaceutical composition, capable of modulating ORL-1 receptor function, based on said compound.

EFFECT: obtaining novel compounds which can be used in medicine to treat pain, memory disorder, obesity, constipation, depression, dementia, Parkinsonism, anxiety, cough, diarrhoea, high blood pressure, epilepsy, anorexia, urinary incontinence or drug dependence.

27 cl, 2 tbl, 19 ex

FIELD: chemistry.

SUBSTANCE: invention relates to novel pyrrole derivatives of the formula (1): or pharmaceutically acceptable salts thereof, where: R1 denotes H, halogen; R2 denotes an 8-10-member bicyclic hydrocarbon group, optionally substituted, or a bicyclic heterocyclic group consisting of one or two atoms selected from nitrogen, oxygen and sulphur and 5-9 carbon atoms, optionally substituted, where the optional substitute is halogen, lower alkyl, OH, lower alkoxy, oxo, NO2, CN; R3 denotes H.

EFFECT: compounds have inhibiting action of production of IL-6, which enables use thereof in a pharmaceutical composition and when treating a range of diseases.

12 cl, 1 tbl, 10 ex

FIELD: chemistry.

SUBSTANCE: invention relates to novel pyrimidine derivatives of general formula (I), or pharmaceutically acceptable salts thereof, where X1 denotes N, N-R3, C-R3 or O; X2 denotes N, CH or C-CH3; X3 denotes N or C, where X1, X2 and X3 all do not simultaneously denote N; X4, X5 each independently denotes C or N; X6 denotes N or C-R1 ; where at least two and not more than four of X1, X2, X3, X4, X5 and X6 denotes N; and where the bonds between X1 and X2, X2 and X3, X3 and X4, X4 and X5, X5 and X1, as well as X5 and X6 can each independently be single or double, or , can form an aromatic ring, under the condition that a chemically stable structure is formed; R denotes -(cyclohexyl)R2; R1 denotes a halide, nitro group, -CN, -CH2CN, -OH, -NH2, -COOH or -Y1R4; Y1 denotes -O-, -SO2, -NHSO2-, -C(O)O- or a bond; R4 denotes (C1-C6)alkyl, phenyl or benzyl, each of which is substituted with 0-3 times with a hydroxy group or a halogen; n equals 0, 1 or 2; R3 denotes H or (C1-C6)alkyl; R2 denotes H, -OH, =O or -Y2-Y3-Y4-R5, where Y2 denotes -C(O)-, -C(O)NRa-, -NH- or a bond; Y3 denotes (C1-C6)alkylene or a bond; Y4 denotes -NRa-, -S-, -SO2-, -NRaC(O)-, -NHSO2- or a bond; R5 denotes (C1-C6)alkyl, (C3-C10)cycloalkyl, a 5-6-member heterocyclyl containing 1-2 heteroatoms selected from N and O, where R5 is optionally substituted with -OH or -NHRa; where each Ra independently denotes hydrogen or (C1-C6)alkyl; Z denotes hydrogen. The invention also relates to a pharmaceutical composition based on a compound of formula (I).

EFFECT: obtaining novel pyrimidine derivatives which are useful for therapeutic or preventive treatment of a c-Jun N-terminal kinase-mediated disorder.

14 cl, 3 tbl, 26 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention relates to compound of formula (I): or to its pharmaceutically acceptable ester, amide, carbamate, solvate or salt, including salt of such ester, amide or carbamate and solvate of such ester, amide, carbamate or salt, where values R1, R2, R3, R4, R5 and R6 are given in item of the formula, with the exception: 4-[3-(4,5-dihydro-1H-imidazol-2-yl)-2-(3,5-dimethylisoxazol-4-yl)indole-1-yl]phenol; 1-(4-hydroxyphenyl)-2-(4-methylimidazol-1-yl)-1H-indole-3-carbonitryl; 1-(4-hydroxyphenyl)-2-(1H-pyrazol-3-yl)-1H-indole-3-carbonitryl; 1-(3-chloro-4-hydroxyphenyl)-2-(1-methyl-1H-pyrazol-4-yl)-1H-indole-3-carbonitryl; 1-(4-hydroxyphenyl)-2-prop-1-inyl-1H-indole-3-carboxylic acid amide.

EFFECT: compounds I possess affinity of binding with estrogen receptor of p-subtype, which makes it possible to use them in pharmaceutical composition and in treatment or prevention of state, associated with disease or disorder, associated with activity of estrogen receptors of β-subtype.

27 cl, 271 ex

FIELD: biotechnologies.

SUBSTANCE: invention relates to new salts of a compound 1:

,

which represents hemi- or monosalts with C4 organic dibasic acid, which have improved properties when used. The invention also refers to method of specified salts production.

EFFECT: improved acid stability.

24 cl, 8 dwg, 4 tbl, 39 ex

FIELD: biotechnologies.

SUBSTANCE: invention relates to new derivatives of 4-aminopyrimidine, having antagonistic activity in respect to the receptor H4. In the formula : R1 is selected from C1-8alkyl, C3-8cycloalkyl-C0-6alkyl, aryl-C1-6alkyl, the group of the formula R2 and R3 together with the atom N, with which they are linked, creating the saturated heterocyclic group, which may be a 4-7-member monocyclic or 9-member condensed bycyclic one, where the specified heterocyclic group may contain up to two atoms N and does not contain any other heteroatoms, and may be unnecessarily substituted with one or more substitutes, independently selected from C1-4alkyl and NRaRb, provided that if the heterocyclic group either contains 2 atoms N and is not substituted with the group NRaRb, or comprises 1 atom N and is substituted with one group NRaRb; or R2 is H, and R3 is pyrrolidinyl substituted with C1-4alkyl group; where Ra is H or C1-4alkyl; Rb is H or C1-4alkyl; R4 and R5 is H; R6 is aryl group; n is equal to 1; and aryl is phenyl group, unnecessarily substituted with halogen. The invention also relates to a pharmaceutical composition containing the specified compounds, to application of compounds for production of a medicinal agent for treatment of such diseases as an allergic, immunological or inflammatory diseases or pain, and also to a method for production of the specified compounds.

EFFECT: higher efficiency of application of compounds in treatment of diseases.

34 cl, 30 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: present invention refers to compounds of formula (I) or pharmaceutically acceptable salts thereof wherein A, R1, R2, R3 and m are specified in the patent claim. The present invention also refers to the number of specific compounds, and to a pharmaceutical composition containing the above compounds effective for inhibition of kinases, such as glycogen synthase kinase 3 (GSK-3), Rho kinase (ROCK), Janus kinase (JAK), AKT, PAK4, PLK, CK2, KDR, MK2, JNK1, aurora, pim 1 and nek 2.

EFFECT: preparing the specific compounds and pharmaceutical composition containing the above compounds effective for kinase inhibition.

18 cl, 393 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to compounds of formula I and formula IV wherein the radical values are such as specified in cl. 1 and 4 of the patent claim, as well as to their therapeutically acceptable salts. Besides, the invention refers to a composition for treating cancer on the basis of the compounds of formula I, to using the compounds of formula I for preparing the therapeutic agent for treating cancer, as well as to using it for treating cancer.

EFFECT: there are prepared and described the new compounds which inhibit anti-apoptotic Bcl-2 and Bcl-x protein activity.

17 cl, 481 ex

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