2-[4-[2-(2-amino-4-oxo-4,6,7,8-tetrahydro-3h-pyrimido[5,4 - b] [1,4]thiazin-6 - yl)ethyl] benzoyl(or thienylboronic)amino] entandikwa acid or its lower alkilany ether, the method of inhibiting the growth and proliferation of cells and a method of inhibiting the enzyme


(57) Abstract:

This invention relates to new compounds of General formula I

< / BR>
where Ar denotes phenyl, thienyl or thienyl, substituted lower alkyl, or lower alkylamino ether, inhibiting the enzyme glacialinterglacial, as well as to a method of inhibiting the growth and proliferation of cells of microorganisms or higher organisms by introducing a host an effective amount of compound I or its ester, and to a method of inhibiting in vitro glacialinterglacial effect on the enzyme effective amount of the compounds of formula I or its ester. The compounds of formula I get the multistage process involving the interaction of the compounds of formula III with a compound IV in the presence of a base to obtain compound V, which when exposed to acid in the solvent leads to the production of compound VI, which restore, and the obtained compound VII hydrolyzing to obtain compound VIII and compound VIII or a compound VII, where R3- hydrogen, is subjected to the influence hydrochloride diapir glutamic acid and, if necessary, hydrolyzing the compound obtained. 3 S. and 3 C.p. f-crystals, 3 tables.

A large class of antiproliferative means includes antimetabolites connection. A concrete subclass of antimetabolites, known as antifolates and antipholi, is an antagonist of vitamin folic acid. Usually antifolates are very similar in structure with folic acid and include typical P-benzoylamino part of folic acid. Glutamate part folic acid accepts a double negative charge at physiological pH values. Therefore, this compound and its analogs have an active energy, rail transport system across the cell membrane, and have a metabolic effect.

Glycinebetaine-formyltransferase (GARFT) is palatability enzyme in the metabolic pathway of purine biosynthesis de novo. This path is critical for the division and proliferation of cells. It is known that the stop of this metabolic pathway has antipole who studied for their ability to inhibit GARFT. It was reported that the prototype, specifically, the strong binding of the inhibitor GARFT, 5,10-dictatorially acid, and antitumor activity. See F. M. Muggia, "Folat antimetabolites inhibitory to de novo purine synthesis" in New Drugs, Concepts and Results in Cancer Chemotherapy, p. 65 - 87, Kluwer Academic Peblishers, Boston (1991).

The present invention relates to a new class of compounds containing a residue of glutamic acid or its ester. These compounds are effective in the inhibition of the enzyme glycinebetaine-formyltransferase (GARFT), and growth and proliferation of cells of higher organisms and microorganisms such as bacteria, yeast and fungi. The invention relates also to methods of using these compounds as inhibitors of the enzyme.

As described above, the connection according to the invention possess anti-proliferative activity, a property that can be expressed in terms of antitumor activity. The connection according to the invention can be used by itself or can be a precursor, which is converted in vivo to the active cadinene. Preferred compounds of the invention are active in the inhibition of the enzyme GARFT. In particular, preferred compounds active in inhibiting the growth of the cell line L1210, to the us to be active in inhibiting the growth of bacteria such as gram-negative bacteria Escherichia coli, which can be grown in culture.

Another aspect of the present invention relates to a therapeutic method of inhibiting the growth and proliferation of cells of higher organisms or microorganisms, which provides an introduction to the host an effective amount of the compounds of the present invention. Compounds of the invention, in particular suitable for the treatment of mammals such as human, for the treatment of birds. In particular, the preferred therapeutic method involves the introduction of a host an effective amount of the compounds of the present invention for inhibition of GARFT.

Many of antiproliferative drugs described herein, or their pharmaceutically acceptable salts can be used in the method of treatment according to the invention. Compounds can be introduced in the form of a pharmaceutically acceptable composition containing a diluent or carrier such as described above.

Doses of the compounds preferably include pharmaceutical dosage units containing an effective amount of the active compound. The term "effective amount" denotes an amount sufficient for inhibition politiska metered units.

Approximate daily dosage unit for spinal host contains up to one gram of active compound per kilogram of body weight of the host, preferably half a gram, more preferably 110 mg and most preferably about 50 mg or less per kilogram of body weight of the host. The selected dose can be administered warm-blooded animal or a mammal, such as man, in need of treatment by inhibiting folate metabolic pathways, by any known method dose, including local manner, for example, ointment or cream; orally, rectally, e.g. in the form of a suppository; parenterally by injection or long-term intravaginal, vnutrimyshernm, intrabronchial, custom or intraocular administration.

Compounds of the present invention can be characterized as exhibiting one or more antiproliferative effect, antibacterial, antiparasitic action, antiviral action, antipsoriatic action Antiprotozoal action anticoccidial action, anti-inflammatory action, an immunosuppressive effect or antifungal action. These compounds are especially suitable for the manifestation protivoopujolevoe deatiny compounds able to inhibit GARFT and presents[2-[4-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido[5,4-b] [1,4] thiazin-6-yl)ethyl] benzoyl or thienylboronic)amino]pentadienoic acid of formula Ia

< / BR>
where Ar denotes phenyl, thienyl or thienyl, substituted lower alkyl;

or lower alkylamine ethers.

Preferably Ar is selected from

< / BR>
In particular, preferably Ar is represented by group

< / BR>
and the connection d is-(2-[4-[2-(2-amino-4-oxo-4,6,7,8 - tetrahydro-3H-pyrimido[5,4-b][1,4]thiazin-6-yl)ethyl)benzoylamine pentadienoic acid.

It is preferable that the compound is diethyl ether d-(2-[4-[2-(2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido [5,4-b] [1,4]-thiazin-6-yl)ethyl]benzoylamine]pentadienoic acid.

It should be noted that the use of instructions of the present invention to a specific problem or environment will be within the abilities of a person skilled in the art in light contained in the invention indications. Examples of products of the present invention and representative way of their acquisition and allocation shown in the following examples.

Example 1. Stage 1A. (1) 1,1-Dimethoxy-3-in-2-ol

< / BR>
In the mixed solution 1,037 g (10,55 mmol) trimethylsilylacetamide. After 10 min at -78oC is added dropwise a solution of 1.21 g (10.46 mmol) of dimethylacetal glyoxal in 5 ml of tetrahydrofuran (THF). After incubation for 1 hour at -78oC the reaction is quenched with 1 ml of H2O and leave to warm to room temperature, diluted with ethyl acetate and washed with saturated solution of NaCl. The aqueous layer was again extracted with ethyl acetate and the combined organic layer is dried over MgSO4and concentrate under reduced pressure. To 785 mg of the obtained yellow oil dissolved in tetrahydrofuran (THF), add 5.8 ml of a 1.0 M solution of tetrabutylammonium fluoride in tetrahydrofuran (THF). After heating for 1 hour at 50oC volatile part is evaporated and the residue purified flash-chromatography on silica gel with elution with a mixture of methylene chloride and ethyl acetate (9 : 1). So get 412 mg (total yield 60%) olkinuora alcohol, compound (1), in the form of a colorless oil. IR-spectrum (connection nerasbavlennom): 3441 (wide), 3277, 2944, 2839, 1636, 1450, 1196, 1084 cm-1.

1H NMR spectrum (CDCl3) 2.42 (broad, 1H), 2.49 (s, 1H), 3.51 (s, 3H), 3.53 (s, 3H), 4.36 (broad, 2H).

Elemental analysis:

Calculated for C6H10O30.35 H2O: C, 52.81; H, 7.90.

Found: C At 52.87; H, 7.86.

IR-spectrum (connection nerasbavlennom): 3451 (wide), 2953, 2838, 1717, 1607, 1437, 1310, 1283, 1120, 1082 cm-1.

1H NMR spectrum (CDCl3) 2.46 (broad, 1H), 3.55 (s, 3H), 3.56 (s, 3H), 3.92 (s, 3H), 4.45 (d, 1H, J = 5.4 Hz), 4.60 (d, 1H, J = 5.3 Hz), 7.52 (d, 2H, J = 8.3 Hz), 7.98 (d, 2H, J = 8.3 Hz).

Elemental analysis:

Calculated for C14H16O5: C, 63.62; H 6.10.

Found: C, 63.14; H, 6.14.

Stage 1c

(3) Methyl ester of 4-(3-hydroxy-4,4-dimethoxymethyl)benzoic acid

< / BR>
The solution containing 14.37 g (54.38 mol) of the compound (2) and 1.40 g of 5% Pd on coal in 175 ml of ethanol, hydronaut when the hydrogen pressure 275790.28 PA in the Parr apparatus. After 2.5 hours the reaction mixture is filtered and the catalyst washed with ethanol and methanol. After concentration under reduced pressure the residue is dissolved in methylene chloride and filtered through a small layer of silica gel with elution with methylene chloride, then with a mixture of methylene chloride and ethyl acetate (1 : 1) for UDA.

IR-spectrum (connection nerasbavlennom): 3495 (wide), 2953, 1721, 1611, 1437, 1283, 1109, 1080 cm-1.

1H-NMR spectrum (CDCl3) 1.72 - 1.93 (m, 2), 2.75 - 2.93 (m, 2H), 3.39 (s, 3H), 3.44 (s, 3H), 3.58 (s, 1H), 3.90 (s, 3H), 4.13 (d, 1H, J = 6.1 Hz), 7.29 (d, 2H, J = 8.1 Hz), 7.95 (d, 2H, J = 8.2 Hz).

Elemental analysis:

Calculated for C14H20O50.20 H2O: C, 61.84; H, 7.56.

Found: C, 61.83; H, 7.57.

Stage 1d

(4) Methyl ester of 4-(3-methanesulfonate-4,4-dimethoxymethyl)-benzoic acid

< / BR>
To a stirred solution of 206 mg (0.77 mmol) of a saturated alcohol, compounds (3) and 0.16 ml (1.15 mmol) of triethylamine in 5 ml of methylene chloride at 0oC added 0.07 ml (0.085 mmol) of methanesulfonamide. After incubation for 20 min at 0oC add 0.02 ml of methanesulfonanilide. After another 30 min the reaction mixture was poured into a saturated solution of NaHCO3and extracted twice with methylene chloride. The combined organic layers dried over MgSO4and the solvent is removed under reduced pressure. This compound was sufficiently pure for use in the next stage. An analytical sample is obtained by purification of the flash-chromatography on silica gel with elution with a mixture of methylene chloride and ethyl acetate (20 : 1). Thus recip is, 611, 1437, 1352, 1283, 1177, 1109, 1078 cm-1.

1H-NMR spectrum (CDCl3) 2.04 (m, 2H), 2.77 - 2.91 (m, 2H), 3.09 (C. 3H), 3.41 (s, 3H), 3.45 (C. 3H), 3.90 (s, 3H), 4.38 (d, 1H, J = 5.5 Hz), 4.64 (m, 1H), 7.29 (d, 2H, J = 8.2 Hz), 7.96 (d, 2H, J = 8.2 Hz).

Elemental analysis:

Calculated for C15H22O7S: C, 52.01; H, 6.40; S 9.26.

Found: C, 52.08; H, 6.44; S 9.25.

Stage 1e

(5) Methyl ether (4-(3-methanesulfonate-4 - oxobutyl)-benzoic acid

< / BR>
In a mixed solution of 600 mg (1.73 mmol) of dimethylacetal (nelfinavir) (compound 4) in 5 ml of methyl chloride at 0oC add 1 ml of H2O and 1 ml triperoxonane acid. The reaction mixture is heated to room temperature and then boiled for 24 hours under reflux. The cooled reaction mixture is diluted with ethyl acetate and twice successively washed with saturated NaCl solution, a saturated solution of NaHCO3then again with saturated NaCl solution, dried (MgSO4) and volatile products are removed under reduced pressure. So get mesilate, the compound (5), which is used without purification.

The NMR spectrum (CDCl3) 2.20 (m, 2H), 2.85 (m, 2H), 3.17 (s, 3H), 3.91 (s, 3H), 4.95 (DD, 1H, J = 4.2 Hz, 8.4 Hz), 7.29 (d, 2H, J = 8.1 Hz), 7.99 (d, 2H, J = 8.2 Hz), 9.59 (s, 1H).

Stage 1f

(6) Methyl who(2.28 mol) of nelfinavir, compounds (5) and 0.40 ml (2.29 mol) of N,N-diisopropylethylamine in dimethylformamide (DMF) is added 0.48 ml (3.44 mol) of 4-methoxy -- colortool. After incubation for 3 hours at room temperature, the reaction mixture was poured into 0.5 N HCl and extracted twice with ethyl acetate. The combined organic layers are washed twice with saturated NaCl solution, dried (MgSO4) and concentrate under reduced pressure. The resulting aldehyde, compound (6), sufficiently pure for use without further purification in the next stage.

IR-spectrum (KBr) 2930, 1715, 1703, 1611, 1512, 1282, 1244, 1107 cm-1.

1H-NMR spectrum (CDCl3) 1.80 - 2.16 (m, 2H), 2.75 (m, 2H), 2.99 (m, 1H), 3.53 (AB, 2H, J = 13.4 Hz), 3.81 (s, 3H), 3.90 (s, 3H), 6.83 (d, 2H, J = 8.5 Hz), 7.12 (d, 2H, J = 8.2 Hz), 7.19 (d, 2H, J = 8.6 Hz), 7.90 (d, 2H, J = 8.2 Hz), 9.27 (d, 1H, J = 4.2 Hz).

Stage 1g

(7) Methyl ester 4-[3-[1,3]dioxolane-2-yl-3-(4 - methoxybenzenesulfonyl)propyl]benzoic acid

< / BR>
The vessel containing 301 mg (0.84 mmol) of the aldehyde, compound (6), 94 μl (1.68 mmol) of ethylene glycol and 42 mg (0.17 mmol) of p-toluensulfonate pyridinium and 30 ml of benzene is heated at the boil under reflux, removing water formed by using traps Dean-stark. After 3 frequent reaction mixture was poured into a saturated solution of NaCl and EXT the nom pressure. The residue is purified flash-chromatography on silica, elwira mixture of hexanol and ethyl acetate (5:1). Such a method are specified in the header of the compound (7) with a total yield of 84%, based on dimethylacetal, the compound (4).

IR-spectrum (connection nerasbavlennom): 2949, 2886, 1721, 1611, 1510, 1435, 1279, 1248, 1177, 1111, 1034 cm-1.

1H-NMR spectrum (CDCl3) 1.67 (m, 1H), 1.99 (m, 1H), 2.57 (m, 2H), 2.84 (m, 1H), 3.72 - 4.03 (m, 6H), 3.81 (s, 3H), 3.90 (s, 3H), 4.97 (d, 1H, J = 4.6 Hz), 6.83 (d, 2H, J = 8.6 Hz), 7.09 (d, 2H, J = 8.1 Hz), 7.23 (d, 2H, J = 8.5 Hz), 7.88 (d, 2H, J = 8.1 Hz).

Elemental analysis:

Calculated for C22H26O5S: C, 65.65, H 6.51, S 7.97

Found: C, 65.72, H 6.50, S 8.07

Stage 1h

(8) Methyl ester of 4-(3-[1,3]dioxolane-2-yl-3-mercaptopropyl)benzoic acid

< / BR>
In a mixed solution of 5.70 g (14.16 mmol) of the compound (7) and 5.42 g (17.00 mmol) acetate mercury (II) in methylene chloride cooled to 0oC, is added dropwise 5 ml triperoxonane acid. After incubation for 3 hours at 0oC add a saturated solution of hydrogen sulfide in methanol and stirring is continued at 0oC for 20 min, the Reaction mixture was poured into saturated NaCl solution and extracted two times with methylene chloride. The combined organic layer is dried (MgSO44) and concentrate under reduced pressure. The residue is purified flash-chromatography on silica gel with elution with a mixture of hexanol and ethyl acetate (4:1). Thus obtain 1.99 g (50%) of the thiol, compound (8), in the form of a light yellow oil.

IR-spectrum (connection nerasbavlennom): 2951, 2886, 1719, 1611, 1435, 1281, 1179, 1144, 1111 cm-1.

1H-NMR spectrum (CDCl3) 1.64 (d, 1H, J = 7.8 Hz), 1.74 (m, 1H), 2.15 (m, 1H), 2.80 (m, 2H), 3.00 (m, 1H), 3.90 (s, 3H), 3.97 (m, 4H), 4.91 (d, 1H, J = 4.0 Hz), 7.28 (d, 2H, J = 8.1 Hz), 7.95 (d, 2H, J = 8.1 Hz).

Elemental analysis:

Calculated for C14H18O4S: C, 59.55, H 6.14

Found: C, 59.42, H 6.41

Phase 1i

(9) Methyl ester 4-[3-(2,4-diamino-6-oxo-1,6-dihydropyrimidin - 5-ylsulphonyl)-3-[1,3]dioxolane-2-yl]benzoic acid

< / BR>
In a mixed solution of 1.91 g (6.76 mmol) of the thiol, compound (8) and 1.39 g (6.78 mmol) of 5-bromo-2,4-diamino-6-oxopyrimidine in the atmosphere of argon in degassed N, N-dimethylformamide added 1.18 ml (6.77 mmol) of N,N-diisopropylethylamine. The reaction with the formed precipitate was separated by filtration, washed with water and dried in air. The filter cake was dispersed in methylene chloride. Slowly add hexane and precipitate again separated by filtration, washed with hexane and dried. Thus obtain 1.91 g (69%) of the target dioxolane, compound (9) in the form not of a white solid substance with so pl. 206 - 208oC (decomposition).

IR-spectrum (KBr): 3439, 3341, 3154, 1701, 1636, 1591, 1470, 1447, 1287 cm-1.

1H-NMR-spectrum (DMSO) 1.64 (m, 1H), 1.84 (m, 1H), 2.58 (m, 1H), 2.79 (m, 1H), 3.19 (m, 1H), 3.82 (s, 3H), 3.85 (m, 4H), 4.83 (d, 1H, J = 4.3 Hz), 6.33 (user s, 4H), 7.33 (d, 2H, J = 8.1 Hz), 7.83 (d, 2H, J = 8.1 Hz), 10.03 (s, 1H).

Elemental analysis:

Calculated for C18H22N4O5S: C, 53.19, H 5.46, N 13.78, S 7.89

Found: C, 52.98, H 5.53, N 13.60, 7.76

Stage 1j

(10) Methyl ester 4-[2-(2-amino-7-hydroxy-4-oxo-4,6,7,8 - tetrahydro-3H-pyrimido[5,4-b][1,4]thiazin-6-yl)ethyl]benzoic acid

< / BR>
Stir a suspension of 1.20 g (2.85 mmol) of dioxolane, compound (9), and 4 ml of 2 N HCl in 20 ml of tetrahydrofuran (THF) and heated to boiling under reflux for 2.5 hours. The homogeneous solution was poured slowly into a saturated solution of NaHCO3and the formed precipitate was separated. The filter is extracted with ethyl acetate. The residue (58 mg), formed between the layers separated and the thing in common is the STATCOM (43 mg) will also be combined with the first precipitate. So get 984 (98%) of amerosport, compound (10), as an orange solid: so pl. 213 - 216oC.

IR-spectrum (KBr): 3351, 3441, 1705, 1638, 1609, 1557, 1470, 1289, 1113, 1020 cm-1.

1N-AMN-range one pair of diastereomers (DMSO) 1.39 and 1.96 (m, m, 1H), 1.70 (m, 1H), 2.56 - 2.89 (m, 3H), 3.82 (s, 3H), 4.71 and 4.84 (m, m, 1H), 5.37 and 5.40 (d, d, 1H, J = 6.6 Hz), 6.06 (s, 2H), 7.20 (d, 1H, J = 4.5 Hz), 7.31 and 7.36 (d, d, 2H, J = 8.1 Hz), 7.86 and 7.88 (d, d, 2H, J = 8.0 Hz), 10.16 and 10.19 (s, 1H).

Elemental analysis:

Calculated for C16H18N4O4S 1.7 H2O; C 48.89, H 5.49, N 14.26, S 8.16

Found: C 48.78, H 5.18, N 14.00, S 8.03

Stage 1k

(11) Methyl ester 4-[2-(2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido[5,4-b] [1,4]thiazin-6-yl]benzoic acid

< / BR>
At 0oC) suspension of 1.126 g (3.1 mmol) of amerosport, compounds (10), in tetrahydrofuran (THF) is added 2.3 ml (18.64 mmol) epirate boron TRIFLUORIDE. After this add the add on parts 0,586 g (9.32 mmol) of cyanoborohydride sodium within 5 min After 30 min after addition was added 5 ml of a saturated solution of ammonia in methanol, the reaction mixture was diluted with ethyl acetate and washed with saturated solution of NaCl. The organic layer is dried (MgSO4) and the solvent is removed under reduced pressure. OST is 1). Thus receive 542 mg (50%) digidratirovannogo ether, compound (11), as an orange solid substance with so pl. 245 - 246oC (decomposition).

IR-spectrum (KBr): 3358, 2936, 1721, 1644, 1595, 1537, 1447, 1346, 1281 cm-1.

1H NMR-spectrum (DMSO) 1.72 (m, 1H), 1.90 (m, 1H), 2.80 (m, 3H), 3.22 (m, 1H), 3.52 (m, 1H), 3.82 (s, 3H), 6.00 (s, 2H), 6.65 (s, 1H), 7.37 (d, 2H, J = 8.1 Hz), 7.87 (d, 2H, J = 8.1 Hz), 10.05 (s, 1H).

Elemental analysis:

Calculated for C16H18N4O3S: C, 55.47, H 5.24, N 16.17, S 9.26

Found: C, 55.31, H 5.29, N 16.09, S 9.17

Stage 11

(12) 4-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido-[5,4-b] [1,4] thiazin-6-yl)ethyl)benzoic acid

< / BR>
A solution of 530 mg (1.53 mol) of the ester, compound (11), and 10 ml and 1 H of NaOH solution is stirred at room temperature for 30 minutes a Homogeneous solution is carefully acidified (pH 4) with concentrated HCl. After cooling in an ice bath poorly-orange precipitate was separated by filtration and air-dried, the Residue is then suspended in ethanol and the ethanol is then removed under reduced pressure. So get 468 mg (91%) acid, compound (12), which decomposes at temperatures above 310oC.

IR-spectrum (KBr): 3285, 3086, 2928, 1698, 1642, 1611, 1576, 1449, 1348 cm-1.

1H NMR-spectrum (DMSO) 1.72 (m, 1H), 1.89 (m, 1H), 2.78 (nty analysis:

Calculated for C15H16N4O3S 1.20 H2O: C, 50.89, H 5.24, N 15.83, S 9.06

Found: C, 50.70, H 4.92, N 15.58, S 8.87

Stage 1m

(13) Diethyl ether 2-[4-[2-(2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido[5,4-b] [1,4]thiazin-6-yl)ethyl]benzoylamine]-pentandiol acid

< / BR>
In a mixed solution of 397 mg (1.19 mmol) of the acid compound (12), 169 mg (1.25 mmol) of hydrate of 1-hydroxybenzotriazole (HOBT), 0.22 ml (1.25 mmol) of N, N-diisopropylethylamine and 300 mg (1.25 mmol) of the hydrochloride of diethyl ether L-glutamic acid in 15 ml of N,N-dimethylformamide added 240 mg (1.25 mmol) of the hydrochloride of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC). After incubation for 18 hours at room temperature, the reaction mixture was poured into ice saturated NaCl solution and the formed precipitate was separated, washed with water and dried in air. Purify the residue flash-chromatography on silica gel with elution with a mixture of methylene chloride and methanol (9: 1). So get 357 mg (58%) of the desired product (13) as a slightly orange solid with so pl. 132 - 136oC.

IR-spectrum (KBr): 3333, 1732, 1645, 1572, 1535, 1449, 1343, 1203, 1020 cm-1.

1H-NMR-spectrum (DMSO) 1.15 (t, 3H, J = 7.3 Hz), 1.17 (t, 3H, J = 7.3 Hz), 1.72 (m, 1H), 1.88 - 2.10 (m, 3H), 2.42 (t, 2H, J = 7.4 Hz), 2.79 (m, 3H), 3.2 is), 8.64 (d, 1H, J = 7.41 Hz), 10.24 (s, 1H).

Elemental analysis:

Calculated for: C24H31N5O6S:

C 55.69, H 6.04, N 13.53, S 6.19

Found: C, 55.41, H 6.11, N 13.48, S 6.12

Stage 1n

(14) 2-[4-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido-[5,4-b] [1,4]thiazin-6-yl)ethyl]benzoylamine]entandikwa acid

< / BR>
A mixture of 320 mg (0,618 mmol) of glutamate, the compounds (13), and 6 ml of 1 H NaOH solution stirred at room temperature for 3 hours, neutralized with concentrated HCl, then slightly acidified with 2 H HCl. After cooling slightly yellow precipitate is collected and air-dried. The filter cake is transferred into a mixture of ethanol and acetonitrile and the remaining water is removed by azeotropic distillation. Thus obtained 220 mg (77%) decollate, compound (14), S. T. pl. 188 - 190oC.

IR-spectrum (KBr): 3348 (wide), 2930, 1717, 1642, 1539, 1505, 1348 cm-1.

1H NMR-spectrum (DMSO) 1.71 (m, 1H), 1.92 (m, 2H), 2.08 (m, 1H), 2.34 (t, 2H, J = 7.4 Hz), 2.79 (m, 3H), 3.20 (m, 1H), 3.55 (m, 1H), 4.38 (m, 1H), 6.07 (s, 2H), 6.68 (s, 1H), 7.31 (d, 2H, J = 8.1 Hz), 7.80 (d, 2H, J = 8.2 Hz), 8.53 (d, 1H, J = 7.7 Hz), 10.11 (s, 1H), 12.40 (user. s, 2H).

Elemental analysis:

Calculated for C20H23N5O6S 1.5 H2O

C 49.17, H 5.36, N 14.34, S 6.56

Found C at 48.77, H 4.97, N 14.0,7, S 6.54

Example 2

Perinovic ring systems [Henris, R. N. , Lazarus, R. A., Benkovic, S. J. Preparation of 2-Amino-4(3H)-oxopyrimido[5,4-b] [1,4] thiazines (5-Thiapterins) and Their Evaluation as Cofactors for Phenylalanine Hydroxylase. J. Med. Chem., 1983, 26, 559 - 563].

The method of obtaining thiophene derivatives may be represented by the following reaction scheme.

The schema. Synthesis pirimidilmyetanov

< / BR>
< / BR>
Conditions for reactions

(a) ethyl ester of bromothiophene, (Ph3P)2PdCl2, CuJ, Et3N, CH3CN;

(b) H2with 5% Pd/C, EtOH;

(c) TsOH, EtOH;

(d) TsCl, Et3N, CH2Cl2;

(e) NaN3, DMF or CH3CN ;

(f) (BOC)2O, H2with 5% Pd/C, THF;

(g) TBSCl, Et3N, CH2Cl2;

(h) MsCl, Et3N, CH2Cl2;

(i) TBAF, THF;

(j) NaH, THF;

(k) MsCl, Et3N, CH2Cl2;

(l) KSAc, acetone;

(m) (CH3OCO)2CHCl, K2CO3, MeOH;

(n) TFU, CH2Cl2;

(o) (CH3)3O BF4CH2Cl2;

(p) guanidine hydrochloride, NaOEt, EtOH;

(q) NaOH;

(r) diethyl ester of L-glutamate, EDS, NOWT, DIEA, DMF);

(s) NaOH.

The instructions above were obtained optically pure thiophene derivatives, based on the known optically pure acetylene [Nemec, M., Janda, M. , Srogl, J. , Stibor, I. The Synthesis of 4-Substituted 2-Thiophenecarboxylic Acids. Collect. Czech. Chem. Co93, 72, 6-13].

The process began with the stage catalyzed by palladium join optically pure form acetylene 5 as ethyl ether 2-bromothiophene-5-carboxylic acid and 3-methyl derivative, followed by reduction and hydrolysis of the acid with getting diol intermediates 4a or 4b. For compounds C-6(R) the nitrogen atom in position 8 was introduced by monocotyledonae primary alcohol, followed by substitution with sodium azide. For compounds C-6(S) hydroxyl group was converted by protection of the primary alcohol in the form of a silyl ether, metilirovaniya secondary alcohol, removal of the protective group and education directed epoxide with sodium hydride. The epoxide was opened selectively at a first position where it is less difficult with the help of sodium azide. Aspidosperma was restored in the presence of BOC-anhydride with obtaining alcohols 3a-d. The sulfur atom in the 5 position was introduced by substitution of the appropriate nelfinavir with KSAc. Acetyl protective group was removed under alkaline conditions in the presence of dimethylcarbonate, and when the protective group of the SIDE was removed, the free amine was spontaneously collisionally in the lactam 2a-d. Lactam alkylation using trimethylolpropane guanidine in ethanol by boiling under reflux with obtaining 2-amino-4-(3H)-oxopyrimidine[5,4-b][1,4]-thiazines. Hydrolysis of ethyl esters, peptide binding and hydrolysis was completed the synthesis of glutamic acids 1a-d.

Spectra1H NMR was determined using conventional electrically spectrometer QE-300 operating in a field of 300 MHz. The chemical shift was in Micros () and set the standards for comparison so that in CDCl3CHCl3corresponds 7,26 memorial plaques, and in DMSO-d6DMSO corresponds 2,49 m D. Resolution standards and peaks denoted as follows: s - singlet, d - doublet, DD - doublet of doublets, t - triplet, Shir.with. - broad singlet, Shir.Dr. broad doublet, Shir. - wide signal, m - multiplet. The mass spectrum was determined in the center of the mass spectrometry research Institute Scripa. Infrared absorption spectra were obtained on a FTIR MIDAC Corporation. Elemental analyses were performed by using Atlantic Microlab Inc., Norcroas, GA or MHW Laboratories, Phoenix, AZ and presented the findings at the established elements with an accuracy of 0.4% of theoretical values. N,N-dimethylformamide (DMF) was dried over activated (250oC) molecular sieves. Tetrahydrofuran was subjected to distillation of nitrobenzophenone in nitrogen atmosphere. Et2O refers to diethyl ether, DIEA refers to diisopropylethylamine to 1-(3-(dimethylamino)propyl)-3-ethylcarbodiimide hydrochloride. Chromatography was performed using silica gel 60 (Merck Art 9385). Thin-layer chromatography (TLC) was performed on pre-prepared sheets of silicon dioxide 60 F264(Merck Art 5719). The melting point is defined on the unit MelTcMp and they are not in point.

5-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido- [5,4-b][1,4]thiazin-8(R)-yl)ethyl] thiophene-2-carboxylic acid: not quite white solid: so pl. 283 - 285oC decomp.; []589+ 71,0o(C = 0,60, 1N NaOH); IR (KBr) 3256 (Shir), 2942, 1707, 1641, 1612, 1464, 1364 cm-1;1H NMR (DMSO-d6) 1.72 (m, 1H), 1.89 (m, 1H), 2.81 - 3.04 (m, 2H), 3.16 - 6.2 (m, 3H, overlap of H2O), 6.08 (s, 2H), 6.68 (s, 1H), 6.92 (d, 1H, J = 4.0 Hz), 7.52 (d, 1H, J = 3,7 Hz), 10,12 (s, 1H), 12.80 (width, 1H). Anal. (C13H14N4O3S20,60 H2O) C, H, N, S.

2(S) [[[5-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido- [5,4-b] [1,4]thiazin-6-(R)-yl]thiophene-2-yl]carbonyl]amino]- pentadecanol acid (1a): not quite white solid; T. pl. 191 - 194oC, foams; []589+ 61,9o(C = 0.65, 1N NaOH); IR (KBr) 3389, 3235, 2924, 1701, 1624, 1545, 1340 cm-1;1H NMR (DMSO-d6) 1.70 - 2.04 (m, 4H), 2.29 (d, 2H, J = 7.5 Hz), 2.90 (m, 2H), 3.13 - 3.53 (m, 3H, partially obscured H2O), 4.29 (m, 1H), 6.30 (s, 2H), 6.77 (s, 1H), 6.89 (d, 1H, J = 3,7 Hz), 7.56 (d, 1H, J = 3,7 Hz) and 8.50 (d, 1H, J= 3,7 Hz), 8.50 (d, 1 is-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido [5,4-b] [1,4] thiazin-6(S)-yl)ethyl]thiophene-2-carboxylic acid: it is not a solid substance; so pl. 258 - 261oC decomp.; [ ]589- 81,3o(c=0,63, 1H NaOH); IR (KBr) 3254 (Shir. ), 2918, 1692, 1685, 1453, 1352 cm-1;1H NMR (DMSO-d6) 1.72 (m, 1H), 1.89 (m, 1H), 2.80 - 3.03 (m, 2H), 3.18 - 3.52 (m, 3H, overlap of H2O), 6.09 (s, 2H), 6.80 (s, 1H), 6.92 (d, 1H, J=3,7 Hz), 7.52 (d, 1H, J=3,7 Hz), 10.20 (s, 1H), 12.80 (Shir. , 1H); BMC calculated for C13H14N4O5S2(M+Na+) 361,0405 found 361,0390.

2(S) [[[5-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H - pyrimido[5,4-b][1,4] thiazin-6(S)-yl)ethyl] thiophene-2-yl]-carbonyl]amino] pentadecanol acid (1b): not quite white solid; T. pl. 220oC decomp.; [ ]589-57,1o(C= 0,61, 1H NaOH); IR (KBr) 3363, 3094, 2926, 1711, 1641, 1605, 1559, 1454, 1400, 1333, 1279 cm-1;1H NMR (DMSO-d6) 1.70 - 2.05 (m, 4H), 2.29 (t, 2H, J= 7.4 Hz), 2.87 (m, 2H), 3.15 - 3.48 (m, 3H, overlap of H2O), 4.29 (m, 1H), 6.05 (s, 2H), 6.66 (s, 1H), 6.89 (d, 1H, J=3,7 Hz), 7.65 (d, 1H, J= 3,7 Hz), 8.50 (d, 1H, J=7,7 Hz), 10.05 (s, 1H), 12.50 (width, 2H). Anal. (C18H27N5O8S21,4 H2O) C, H, N, S.

5-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido [5,4-b] [1,4] thiazin-6(R)-yl)ethyl] -4-methylthiophene-2-carboxylic acid: not quite white solid: so pl. 253oC decomp.; [ ]589+80,7o(C=0,29, 1H NaOH); IR (KBr) 3339 (Shir), 2922, 1641, 1589, 1451, 1346, 1269 cm-1;1H NMR (DMSO-d6) 1.64 (m, 1H), 1.82 (m, 1H), 2.09 (s, 3H), Leno for C14H16N4O2S2(M+Na+) 370,0002 found 375,0570.

2(S) [[[5-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido [5,4-b][1,4] thiazin-6(R)-yl)ethyl]-4-methylthiophene-2-yl]-carbonyl]-amino] pentadecanol acid (1c): not quite white solid; T. pl. 210oC decomp.; [ ]589+64,4o(C= 0.45, and 1H NaOH); IR (KBr) 3341 (Shir.), 2928, 1701, 1638, 1536, 1449, 1340 cm-1;1H NMR (DMSO-d6) 1.78 - 2.05 (m, 4H), 2.10 (s, 3H), 2.28 (t, 2H, J=7.0 Hz), 2.82 (m, 2H), 3.45 (m, 3H, overlap of H2O), 4.25 (m, 1H), 5.98 (s, 2H), 6.6 (s, 1H), 7.54 (s, 1H), 8.38 (d, 1H, J=7,7 Hz), 10.05 (s, 1H), 12.5 (Shir.s, 2H). Anal. (C19H23N6O6S20,7 H2O) C, H, N, S.

5-[2-(2-Amino-4-(3H)-oxo-5,6,7,8-tetrahydropyrimido [5,6-b] [1,4] thiazin-6(S)-yl)ethyl] -4-methylthiophene-2-carboxylic acid: [ ]589-77,9o(C= 0,58, 1H NaOH);1H NMR (DMSO-d6) 1.62 - 1.73 (m, 1H), 1.79 - 1.92 (m, 1H), 2.12 (s, 3H), 2.81 - 2.89 (m, 3H), 3.16 - 3.26 (m, 1H), 3.50 - 3.58 (m, 1H), 6.21 (Shir. s, 2H), 6.76 (Shir.s, 1H), 7.45 (s, 1H), 10.24 (width, 1H), 12.76 (width, 1H); Anal. (C14H18N4O3S21,4 H2O) C, H, N, S.

2(S) [[[5-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido [5,4-b][1,4] thiazin-6(S)-yl)ethyl]-4-methylthiophene-2-yl]- carbonyl]-amino]pentadecanol acid (1d): [ ]589- 36,8o(C=0,57, 1H NaOH);1H NMR (DMSO-d6) 1.61 - 1.72 (m, 1H), 1.76 - 1.92 (m, 2H), 1.99-1H), 8.44 (d, 1H, J= 7,7 Hz), 10.12 (Shir.s, 1H), 12.45 (Shir. 2H). Anal. (C19H23N3O8S20,75 H2O) C, H, N, S.

Example 3

Biological and biochemical assessment

Test in vitro

Cell growth in the presence of compounds of the present invention evaluated using cell lines of murine leukemia L1210 (ATCC CCL219). Cell line incubated in medium RPMI 1640 containing 5% V / V heat inactivated serum embryo calf without antibiotics.

The value of the IC50determine in 160 µl of microculture containing 1500 (L1210) cells and installed in the plate with 96 wells in a nutrient medium, supplemented with 50 ME/ml penicillin and 50 µg/ml streptomycin. Growth was measured for 3 days under continuous exposure to various concentrations of the test compounds (14), was added 4 hours after seeding the cells, by the method of Mosmann MITT-tetrazole-resume (Immunol. Meth. 65, 55 - 63 (1983)), modified by Alley et al. (Cencer Res. 48, 589 - 601 (1988)). Water-insoluble compounds are dissolved in dimethyl sulfoxide and diluted to a final concentration of 0.5% solvent in cell cultures.

Determination of the inhibition constant Ki Gar-transformylase

The inhibition constant GAR adding enzyme) by spectrophotometric analysis Young et al. (Biochemistry 23, 3979-3986 (1984)) and monitoring the reaction at 295 nm. Apply GAR transformylase domain of this enzyme in humans. Variable substrate is 10-formyl-5,8-deesaster with concentrations of 0.83 μm, 1.25 μg, 2.5 μm and 5 μm, while the concentration of the other substrate, GAR (glycinebetaine) support constant at 20 μm. The formula for determining contains 20 mm Hepes pH 7.5, 20 μm GAR and variable amounts of 10-formyl-5,8-deesaster and inhibitor. Inhibitor (14) is used in the form of five concentrations ranging from 0 to about 3 Ki. Data analysis is applied on the graph in the form of dependence of the reaction rate on the reciprocal concentration of 10-formyl-5,6-deesaster. The inhibition constant determined again from the graph of the dependence of the angles of inclination of these lines obtained for each concentration of inhibitor, the concentration of the inhibitor.

Biochemical analyses. Activity GART was measured using a modified method Young. The reaction mixture contained the catalytic domain GART man, 20 μm , - GAR, 10 or 20 μm FDDF, variable concentrations of inhibitor GART 50 mm HEPES-KOH, pH 7.5, and 50 mm KCl. The reaction was initiated by addition of enzyme, and then the control reaction was carried out by the absorption of y 294 nm at 20aliciously speed in stationary phase concentration of inhibitor and substrate. On the basis of the dependence of the apparent Ki(Kiapp. ) concentration FDDF found that the type of the investigated competitive inhibition relative to FDDF, and showed that he describes in the following way: Kiapp.=Ki+(Ki/Km)[FDDF]. The Michaelis constant for FDDF,Kmdetermined independently on the basis of the dependence of the catalytic rate on the concentration of FDDF, it was shown that it is 0.6 μm. Data definitions as Kmand Kiwere approximated nonlinear methods for control or Michaelis equation of Michaelis for competitive inhibition that is suitable. The data resulting from the inhibition of durable binding, analyzed and Kiwas determined by approximatevalue these data to equation durable binding Morrison nonlinear methods.

The ability of inhibitors GART to act as a substrate for FPGS were measured using an enzyme derived from rat liver. FPGS were obtained in partially purified form from rat liver essentially by the method of Moran and Colman. Rats Sprague-Dawley were killed by inhalation of CO2. The liver was removed and through the portal vein was perfesional 20 ml ice 20 mm HEPES. a pH of 7.4; 250 mm sucrose; 50 mm 2-mercaptoethane for perfusion, weighed, then mixed in twice its volume weight (in wet state) volume of fresh buffer perfusion, containing 0.5 mm PMSF, 50 μg/ml soybean trypsin inhibitor and 0.5 mm EDTU until a homogeneous mass. The homogenate was filtered through cheesecloth and the filtrate was centrifuged at 130000 x g for 60 minutes resulting supernatant was filtered through a fine glass wool to remove suspended fats and added ATF to 2.5 mm. Protein was besieged by 30% saturated ammonium sulfate and centrifuged at 30,000 x g for 15 minutes the resulting precipitate resuspendable in a small volume of a mixture of 20 mm Tris-Cl pH of 7.4; 50 mm 2-mercaptoethanol, 5 mm MgCl2; 5 mm ATP, and 25 μg/ml soybean trypsin inhibitor, and then centrifuged as above. Of the supernatant was removed suspended fats and proteins besieged 50% saturated ammonium sulfate, and then centrifuged as above, to obtain a precipitate. Besieged thus the protein was stable when stored at -70oC for more than 4 months. Because FPGS quite unstable in solution, frozen sediment FPGS resuspendable for analysis, as described below, and immediately applied. The precipitate was melted and resuspendable a minimal amount of buffer (20 was developed to remove residual solids. The supernatant was absoluely by passing through a column of Sephadex G25, balanced in 20 mm Tris. a pH of 7.4; 50 mm 2-mercaptoethanol, 5 mm MgCl2, 5 mm ATP, and 25 μg/ml soybean trypsin inhibitor and centrifuged to remove residual solids. The supernatant was absoluely by passing through a column of Sephadex G25, balanced in 20 mm Tris, pH 74; 50 mm 2-mercaptopurine and 5 mm MgCl2. Fractions containing protein were pooled and the concentration of ATP was brought to 5 mm. The protein concentration was determined by Bradford method, using BSA (bovine serum albumin as protein standard. Typically, the protein concentration was 3 mg/ml Typical mixture for analysis contained 0.7 mg partially purified protein FPGS with 200 mm Tris-Cl pH of 8.5; variable concentration of the inhibitor GART or folic acid; 5 mm ATP; 10 mm MgCl2; 30 mm KCl; 50 mm 2-mercaptoethanol; 1 mm glutamic acid and 2 µci3H-glutamic acid in a volume of 0.5 ml, the Reaction was initiated by addition of enzyme and were incubated for 60 min at 37oC. the Reaction is then extinguished 0.5 ml of charcoal and put on ice for the complete absorption of the product (getting the solution coal and significant clearance3H-glutamic acid were performed as described RAS 10 mm glutamate with a pH of 6.8; 10 mm 2-mercaptoethanol, in each case with resuspending and centrifugation. Products at the end was suirable from coal 1 ml of a solution of ammonia in ethanol (3 M NH4OH; 60% ethanol) resuspendable and centrifugation. The radioactivity in 1 ml of the supernatant eluent containing tritium-labeled product was measured by scintillation counting after adding 1 ml of water. The kinetic parameters Kmand Vmaxwere determined by nonlinear approximatevalue substrate according to product education for the equation of Michaelis.

The dissociation constant (Kd) mFBP was determined for inhibitors GART method of competitive binding with the use of membrane-associated FBP cells derived from cultured KB cells. Adherent cells scraped from the flasks, washed once with ice saline solution with phosphate buffer (RFB) and centrifuged at 5000 x g for 5 min at 4oC. the Precipitate of KB cells were stored at -70oC, activity of mFBP was stable for at least 6 months. Cellular precipitate containing approximately 2108cells resuspendable in 10 ml of buffer for suspension (KH2PO4-KOH, pH of 7.4; 10 mm EDTU; 10 mm 2-mercaptoethanol), quickly about the P>C. the Precipitate was released from endogenous associated folate-resuspending in 20 ml of acidic buffer (50 mm KH2PO4-KOH, pH 3.5; 10 mm EDTU; 10 mm 2-mercaptoethanol) and centrifuged as above. The precipitate is then resuspendable in 20 ml of buffer for suspension at pH 7.4 and centrifuged as above. Sediment resuspendable in 5 ml of buffer for suspension with a pH of 7.4, containing no EDTU. The protein content was quantitatively determined by the method Bradforda using BSA (bovine serum albumin as protein standard. Normal outputs for this method were 4-5 mg total membrane protein 2108cells. This suspension was used as a source associated with the membrane FBP person. In this analysis of competitive binding to the ligand-competitor was given the opportunity to compete with3H-folic acid for binding to mFBP. The reaction mixture contained 50 to 100 µg of cell membrane protein, containing 3-6 pmol (3-6 nm) FBP, 17,25 pmol3H-folic acid (17,25 nm of 0.5 µci) and different concentrations of ligand-competitor in 1 ml of a mixture of 50 mm KH2PO4-KOH, pH of 7.4; 10 mm 2-mercaptoethanol. Binding assays were performed at 25oC. because of the rapid binding and very IU the of 30 min in the absence of3H-folic acid. Then add3H-folic acid and the mixture was left to reach equilibrium within 2.5 hours. Complete, the reaction mixture was passed through nitrocellulose filters in vacuum to trap the cell membranes with an associated3H-folic acid. Caught membrane was then washed 4 times with 1 ml buffer for the reaction. The amount of bound3H-folic acid was measured by scintillation account nitrocellulose membrane. The data obtained was nonlinear approximatively, as described above. mFBP Kdfor3H-folic acid used to calculate competitor Kdreceived direct titration mFBP3H-folate and subsequent nonlinear approximatevalue data for the equation of a strong binding Kdfound that it is equal 6014 microns.

Measurement IC50in tissue culture. Cell lines L1210 and CCRF-CEM were obtained from the American type culture collection. Cell line L1210/C1920 was developed for resistance to fostriecin and had no function restored Plantago media. The L1210 cells were grown in medium RPM1-1640, supplemented with 5% cialisbuynow fetal bovine serum. Cell lines CEM and L1210/C1920 were grown in RPMI medium, gopalan>in humid thermostat. Inhibition of cell growth was measured by the modified method of Mosmann. Cells in the middle of the log phase of each cell line was diluted up to 18500 cells/ml in fresh culture medium RPMI-1640, supplemented cialisbuynow fetal bovine serum, and then aliquot of sample was injected into the columns (rows) from 2 to 12 titration microplate with 96 wells. Column 1 filled with the same volume, 135 μl of fresh medium without cells for use as control experiment. The microplate was then placed in a thermostat at 37oC in air containing 5% CO2. After 4 hours, the microplate was removed from thermostat and then a solution of a medicinal product, 15 μl/well, when binary dilutions, were added to column 12 to 4. Wells containing test compound was obtained in four replications on each tablet. In the wells in columns 1 and 2 of the tablets was added 15 μl of medium without the test compound. Column 3 received 15 μl of diluent for the drug. The cells were then returned to the incubator for 72 hours (cell line L1210 and C1920) or 120 hours (cell line CEM). For reversible experiments the medium was supplemented with 100 μm gipoksantina. After exposure to the medicinal product into each well of all Mick is food for tissue culture, after that the cells were returned to thermostat. After 4 hours all the tablets were removed from the incubator and centrifuged at 1200 rpm./min for 7 minutes. The medium was aspirated and each well of all tablets was added 150 μl of DMSO. The contents of the tablets were then mixed with a slow speed stirrer type vortex for 1 hour in the dark at room temperature. Content metabolized MTT was measured by the spectrophotometric method at 540 nm on a kinetic microdensitometer Molecular Devices VmaxTM. The concentration of drug required to reduce cell growth by 50%, as measured by the MTT metabolism was determined by interpolation between O. D. (minus control experiment) directly above and below the O. D. 50% of the control (negative control experience).

The molecular mechanics calculations on the connection 13. These calculations were carried out using batchmin rool with version 3.5 program Vacromodel. Models were built connection 13 with pseudoequilibrium and pseudoaxial orientations and loading generated using MORAK 6. Each of the ligands were placed in the active site of the enzyme laying pyrimidine and glutamate parts on the upper part of the crystal structure of compound 5 and the substructure of managemeni shown in the tables.

1. 2-[4-[2-(2-Amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido[5,4-b][1,4]thiazin-6-ethyl] benzoyl(or thienylboronic)amino]intentionaly acid, having the formula Ia

< / BR>
where Ar denotes phenyl, thienyl or thienyl, substituted lower alkyl,

or lower acrylic ester.

2. Connection on p. 1, where Ar is selected from

< / BR>
3. Connection on p. 1, where Ar denotes

< / BR>
and which is the d-2-[4-[2-(2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido[5,4-b][1,4]thiazin-6-yl)ethyl]benzoylamine]pentandiol acid.

4. Connection on p. 1, which is diethyl ether d-2-[4-[2-(2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimido[5,4-b] [1,4] thiazin-6-yl)ethyl] benzoylamine]pentandiol acid.

5. Method of inhibiting the growth and proliferation of cells of microorganisms or higher organisms, characterized in that the owner is in need of such inhibition is administered an effective amount of a compound according to any one of paragraphs.1 to 4.

6. Method of inhibiting in vitro glacialinterglacial (GARET), characterized in that the enzyme is affected effective amount of a compound according to any one of paragraphs.1 to 4.


Same patents:

,5,6]-6-amino-3-azabicyclo [3.1.0] gex-3-yl)- 6-fluoro-1-(2,4-differenl)-1,4 - dihydro-4-oxo-1,8 - naphthiridine-3-carboxylic and methanesulfonic acid and its preparation" target="_blank">

The invention relates to a new form of the anhydrous salt methanesulfonic acid and 7-([1,5,6]-6-amino-3-azabicyclo[3.1.0] Gex-3-yl)-6-fluoro-1-(2,4-differenl)- 1,4-dihydro - 4-oxo-1,8-naphthiridine-3-carboxylic acid, to a method of use of the compounds in the treatment of bacterial infections in mammals, especially humans, and to pharmaceutical compositions useful for him

The invention relates to pyrazolopyrimidines General formula I and their pharmaceutically acceptable salts, where A is the group NR1R2or CR'1R'2R11, R1- H or C1-C6-alkyl, unsubstituted or substituted certain substituents, such as HE, F, CL and others, or C2-C6alkenyl; or C2-C6-quinil; R2-C1-C6-alkyl, unsubstituted or substituted certain substituents, such as HE, C1-C6-alkoxy and others; or C2-C6alkenyl or2-C6-quinil, or furanyl; and (C1-C4-alkylene)phenyl which may be substituted by 1 to 3 substituents: CL, F, C1-C4-alkyl, and one Deputy:1-C6-alkoxy, CF3, NO2, NH2; or (C1-C4-alkylen) hetaryl where hetaryl - thienyl, possibly substituted by CL, benzothiazyl, pyridyl, chinoline, furanyl, benzofuranyl, thiazolyl, benzothiazolyl, pyrrolyl, pyrrolidinyl, 1-benzylpiperidine, tetrahydropyranyl; or (C1-C4-alkylen)cyclopropyl; or NR1R2form hetaryl selected from the group consisting of pyrrolidyl, possibly substituted benzyl, pyrrolidinyl, possibly substituted by benzyl or HE, the IIR>-C6-alkyl; R3is hydrogen, C1-C6-alkyl, O-(C1-C6alkyl), S(C1-C4- alkyl); R4- C1-C6- alkyl, or S(O)n(C1-C6)-alkyl, where n= 0-2, R5- 2,4,6-substituted phenyl CL, C1-C6-alkyl, CF3; R11-N., HE, or COO- (C1-C2alkyl), provided that the group CR'1R'2R11not an alkyl straight chain; and when R3is N, then R4isn't C1-C6the alkyl

The invention relates to medicine, in particular to the experimental therapy, and for the prevention and treatment of atherosclerosis

The invention relates to new compounds of the formula

< / BR>
to pharmaceutically acceptable additive salts of the acid and stereoisomers of these compounds, which are used as antagonists of mediators and have a high activity against Central nervous system

The invention relates to new nucleoside analogues of 1,3-oxathiolane and their use for treating viral infections, HIV infection, hepatitis b, more specifically to the (-)-4-amino-5-fluoro-1-(2-oxymethyl-1,3-oxathiolan-5-yl)-(1H )-pyrimidine-2-ONU (I) and its pharmaceutically acceptable derivative containing pharmaceutical compositions

The invention relates to medicine

The invention relates to a new method of treatment of patients, such as people with benign prostatic hyperplasia (BPH), which includes treatment by assigning a therapeutically effective amount of an inhibitor 5- reductase in combination with blocker1- adrenergic receptor

FIELD: organic chemistry, chemical technology.

SUBSTANCE: invention relates to an intermediate compound, i. e. tert.-butyl-(E)-(6-{2-[4-(4-fluorophenyl)-6-isopropyl-2-[methyl(methylsulfonyl)amino]-pyrimidine-5-yl}-(4R,6S)-2,2-dimethyl[1,3]dioxane-4-yl]acetate that can be used in synthesis of compound of the formula (IV)

eliciting inhibitory effect on activity of HMG-CoA-reductase and, therefore, can be used for preparing pharmaceutical agents for treatment, for example, hypercholesterolemia, hyperproteinemia and atherosclerosis. Also, invention relates to a method for preparing indicated intermediate compound by reaction of the new parent compound - diphenyl-[4-(4-fluorophenyl)-6-isopropyl-2-[methyl(methylsulfonyl)amino]pyrimidine-5-ylmethyl]phosphine oxide with tert.-butyl-2-[(4R,6S)-6-formyl-2,2-dimethyl-1,3-dioxane-4-yl]acetate in the presence of a strong base in simple ether or aromatic solvents or their mixtures at temperature in the range from -200C to -900C. Also, invention relates to a method for preparing of compound of the formula (IV) wherein R1 means hydrogen atom or pharmaceutically acceptable cation and to a method for preparing intermediate compounds of the formula (VI):

wherein each P1 and P2 represents independently (C1-C4)-alkyl or group:

and wherein P3 represents (C1-C8)-alkyl. Applying new intermediate compounds and proposed methods provide enhancing quality and yield of compounds.

EFFECT: improved preparing methods.

9 cl, 1 tbl, 8 ex