HIV inhibiting 5-amido substituted pyrimidines

Abstract

This invention concerns pyrimidine derivatives of formula ##STR00001##
having HIV (Human Immunodeficiency Virus) replication inhibiting properties, the preparation thereof and pharmaceutical compositions comprising these compounds.

Claims

1. A compound having the following structure: ##STR00082## wherein: R.sup.5 is selected from C.sub.3-7cycloalkyl; C.sub.1-6alkyloxy; aryl; Het; C.sub.1-6alkyl substituted with a radical selected from hydroxy, C.sub.1-6alkyloxy, cyano, amino, mono- and di-C.sub.1-6alkylamino, C.sub.1-6alkylcarbonylamino, aryl, Het, dioxolanyl optionally substituted with one or two C.sub.1-6alkyl radicals, tetrahydrofuranyl, pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, piperazinyl optionally substituted with C.sub.1-6alkyl or C.sub.1-6alkylcarbonyl, C.sub.1-6alkyloxycarbonyl, arylC.sub.1-6alkyloxycarbonyl, or C.sub.3-7cycloalkyl; and is C.sub.1-6alkyl substituted with two C.sub.1-6alkyloxy radicals; R.sup.6 is selected from hydrogen and C.sub.1-6alkyl; or R.sup.5 and R.sup.6 taken together with the nitrogen atom to which they are attached form pyrrolidinyl; piperidinyl; morpholinyl; piperazinyl; or piperazinyl optionally substituted with C.sub.1-6alkyl or C.sub.1-6alkylcarbonyl.

2. A compound according to claim 1, wherein: R.sup.5 is C.sub.3-7cycloalkyl; C.sub.1-6alkyloxy; aryl; Het; C.sub.1-6alkyl substituted with a radical selected from hydroxy, C.sub.1-6alkyloxy, cyano, di-C.sub.1-6alkylamino, C.sub.1-6alkylcarbonyl amino, aryl, Het, dioxolanyl substituted with two C.sub.1-6alkyl radicals, tetrahydrofuranyl, pyrrolidinyl, C.sub.1-6alkyloxycarbonyl, and C.sub.3-7cycloalkyl; R.sup.6 is hydrogen or C.sub.1-6alkyl; or R.sup.5 and R.sup.6 taken together with the nitrogen atom to which they are substituted form morpholinyl; piperazinyl substituted with C.sub.1-6alkyl.

3. A compound according to claim 1, wherein R.sup.5 is C.sub.3-7cycloalkyl; C.sub.1-6alkyloxy; C.sub.1-6alkyl substituted with a radical selected from hydroxy, C.sub.1-6alkyloxy, cyano, C.sub.1-6alkylcarbonylamino, aryl, Het, C.sub.1-6alkyloxycarbonyl; and R.sup.6 is hydrogen.

4. A compound according to claim 1, wherein each Het independently is pyridyl, thienyl, thiazolyl, furanyl, each of which may be optionally substituted with a radical selected from C.sub.1-6alkyl.

5. A compound according to claim 1, wherein each aryl independently may be phenyl optionally substituted with C.sub.1-6alkylamino, mono- or di-C.sub.1-6alkyl-amino, C.sub.1-6alkyloxy, aminosulfonyl, Het, the latter more in particular being thiadiazolyl.

6. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and as active ingredient a therapeutically effective amount of a compound as claimed in claim 1.

7. A compound according to claim 1, having the following structure: ##STR00083##

8. A compound according to claim 1, having the following structure: ##STR00084##

9. A compound according to claim 1, having the following structure: ##STR00085##

10. A compound according to claim 1, having the following structure: ##STR00086##

11. A compound according to claim 1, having the following structure: ##STR00087##

Description

EXAMPLES

Example 1

Preparation of Intermediate 2

(1) ##STR00011##

(2) N-bromosuccinimide (0.0393 mol) was added portion wise at room temperature to Intermediate 1 (0.0327 mol), the preparation of which is described in WO-03/016306, in CH.sub.3CN (100 ml). The mixture was stirred at room temperature for 4 hours. The precipitate was filtered off, washed with CH.sub.3CN and dried yielding 10.08 g of the desired end product. The filtrate was evaporated and purified by column chromatography (eluent: CH.sub.2Cl.sub.2 100; 35-70 m). The pure fractions were collected, the solvent was evaporated and the residue was crystallized from CH.sub.3CN. Yield: 2.4 g of Intermediate 2. The two fractions were collected. Total yield: 12.48 g of intermediate 2 (86%, melting point: >250 C.).

Example 2

Preparation of Intermediate 3

(3) ##STR00012##

(4) A mixture of intermediate 2 (0.0247 mol), dichlorobis(triphenylphosphine)-palladium(II) (0.00494 mol) and triethylamine (0.107 mol) in ethanol (100 ml) were stirred at 100 C. for 72 hours under 15 bars pressure of carbon monoxide. The mixture was poured in water and the precipitate was filtered off, yielding 6 g of intermediate 3. The filtrate was extracted with CH.sub.2Cl.sub.2. The organic layer was dried over magnesium sulfate, filtered and the solvent was evaporated. The residue was purified by column chromatography over silica gel (eluent: CH.sub.2Cl.sub.2/MeOH 99.5/0.5; 15-40 m)). The pure fractions were collected and the solvent evaporated. Yield: 1.9 g. The two fractions were combined, yielding 7.9 g of intermediate 3 (73%, melting point: >250 C.).

Example 3

Preparation of Intermediate 4

(5) ##STR00013##

(6) A mixture of intermediate 3 (0.00456 mol), lithium hydroxide, monohydrate (0.0137 mol) in THF (20 ml) and water (7 ml) were stirred at 50 C. overnight. The THF was evaporated. The residue was diluted in water and HCl 3N was added until pH 2-3. The precipitate was filtered off, washed with water and dried. Yield: 1.78 g of intermediate 4 (95%, melting point: >250 C.).

Example 4

Amide Synthesis

(7) Method A:

(8) ##STR00014##

(9) 1-hydroxybenzotriazole (0.000183 mmol, 1.5 eq) was added to a mixture of intermediate 4 (0.00122 mmol, 1.5 eq) in THF (3 ml). Dichloromethane (3 ml) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (0.00183 mmol, 1.5 eq) were added successively to the mixture. To this solution, morpholine (0.00183 mmol, 1.5 eq) was added. The mixture was stirred at room temperature for 24 h then poured in water and K.sub.2CO.sub.3 10% and extracted with a 90/10 mixture of CH.sub.2Cl.sub.2 and methanol. The organic layer was washed with a solution of brine, dried over magnesium sulfate, filtered and the solvent evaporated. The residue was purified by column chromatography over silica gel (eluent: CH.sub.2Cl.sub.2/MeOH 99/1; SiO.sub.2 70-200). Yield: 0.055 g of compound 1 (94%, melting point: >250 C.).

Example 5

Method B

(10) ##STR00015##

(11) Thionyl chloride (7 ml) was added to intermediate 4 (0.000734 mmol). The mixture was heated to reflux 1.5 hour, then evaporated to dryness. The residue was purified by trituration in diethyl ether. Yield: 0.3 g of intermediate 5 (95%).

(12) A mixture of intermediate 5 (0.000233 mol), 2-aminoanisole (0.00035 mol, 1.5 eq) and triethylamine (0.00035 mol, 1.5 eq) in THF (5 ml) and CH.sub.2Cl.sub.2 (5 ml) was stirred at room temperature for 24 hours, then poured in water and K.sub.2CO.sub.3 10% and extracted with AcOEt. The organic layer was washed with a solution of brine, dried over magnesium sulfate, filtered and the solvent evaporated. The residue was purified by column chromatography (eluent: CH.sub.2Cl.sub.2 100% to CH.sub.2Cl.sub.2/MeOH 98/2; Kromasil 3.5 m 150*30). Yield: 0.052 g of compound 56 (53%, melting point: >250 C.).).

(13) The following tables list compounds which were or can be prepared according to the procedures described in the above examples.

(14) TABLE-US-00001 TABLE 1 Phys. Data Com- and pound Meth- stereo- No. od R chemistry 1 A (E) Yield 94% >250 C. 2 A (E) Yield 69% >250 C. 3 A (E) Yield 51% >250 C. 4 A 0 (E) Yield 60% 230 C. 5 A (E) Yield 38% 238 C. 6 A (E/Z/93/7) Yield 48% 221 C. 7 A (E) Yield 50% 175 C. 8 A (E) Yield 43% 218 C. 9 A (E) 62% 214 C. 10 A (E) Yield 78% 246 C. 11 A (E) Yield 60% 227 C. 12 A (E) Yield 67% 238 C. 13 A (E) Yield 62% >250 C. 14 A 0 (E) Yield 32% 246 C. 15 A (E) Yield 63% >250 C. 16 A (E/Z:96/4) Yield 19% >250 C. 17 A (E) Yield 25% >250 C. 18 A (E/Z:96/4) Yield 27% 226 C. 19 A (E) Yield 45% >250 C. 20 A (E) Yield 44% >250 C. 21 A (E/Z:98/2) Yield 39% >250 C. 22 B (E) Yield 68% 226 C. 23 B (E) Yield 28% >250 C. 24 B 0 (E) Yield 26% 226 C. 25 A (E) Yield 37% >250 C. 26 A (E) Yield 58% >250 C. 27 A (E) Yield 31% >250 C. 28 A (E) Yield 59% >250 C. 29 A (E) Yield 39% >250 C. 30 A (E) Yield 18% >250 C. 31 A (E) Yield 53% >250 C. 32 A (E) Yield 50% >250 C. 33 A (E) Yield 45% >250 C. 34 A 0 (E) Yield 9% >250 C. 35 A (E) Yield 15% >250 C. 36 A (E) Yield 14% >250 C. 37 A (E) Yield 65% >250 C. 38 A (E) Yield 43% >250 C. 39 A (E) Yield 36% >250 C. 40 A (E) Yield 43% 199 C. 41 A (E) Yield 56% >250 C. 42 A (E) Yield 21% >250 C. 43 A (E) Yield 56% 226 C. 44 A 0 (E) Yield 66% 245 C. 45 A (E) Yield 66% 222 C. 46 A (E) Yield 40% >250 C. 47 A (E) Yield 85% 221 C. 48 A (E) Yield 77% 250 C. 49 A (E) Yield 57% >250 C. 50 A (E) Yield 11% >250 C. 51 A (E) Yield 49% 225 C. 52 A (E) Yield 32% 260 C. 53 A (E/Z 93/7) mp >250 C. 54 A 0 (E/Z 94/6) mp >250 C. 55 A (E/Z 94/6) mp >222 C. 56 B E mp >250 C. 57 A E

Example 6

Preparation of Intermediate 11

(15) ##STR00074## ##STR00075##

(16) Sodium hydride (60% in oil, 0.036 mol, 1.1 eq.) was added to a stirred solution of 2,6-dimethyl-4-hydroxy-benzaldehyde (0.033 mol, 1.1 eq.) in dioxane (50 ml). Stirring was continued for 10 min before adding 1-methyl-2-pyrrolidinone (50 ml). After another 10 min, intermediate 2 (0.033 mol) was added and the whole mixture was heated at reflux for 18 hours. After cooling down, water and ice were added. The pure product was obtained by filtration. Yield 11.2 g (98%) of Intermediate 7.

(17) Potassium tertbutoxyde (0.026 mol, 1.5 eq.) was added to a solution of diethyl-phosphonoacetonitrile (0.026 mol, 1.5 eq.) in THF (60 ml) at 5 C. under nitrogen. Stirring was maintained 60 min before intermediate 7 (0.017 mol) was added and the whole mixture was stirred 12 hours at room temperature. After cooling down, water was added and the extraction conducted with dichloromethane. The organic layer was dried over magnesium sulfate, filtered and the solvent evaporated. The pure product was obtained by crystallization in ether of the crude. Yield 3.6 g (56%) of Intermediate 8.

(18) Intermediates 9, 10 and 11 were prepared following the same procedures as those previously described in examples 1-4.

Example 7

(19) Following the procedures of example 5 or 6 the following compounds were prepared:

(20) TABLE-US-00002 TABLE 2 Phys. Data and Compound No. R stereo-chemistry 58 (E) >250 C. Yield 62% 59 (E) 118 C. Yield 53% 60 (E) 134 C. Yield 57% 61 0 (E) >250 C. Yield 53% 62 (E) 132 C. Yield 55%

Formulation Examples

Capsules

(21) A compound of formula (I) is dissolved in organic solvent such as ethanol, methanol or methylene chloride, preferably, a mixture of ethanol and methylene chloride. Polymers such as polyvinylpyrrolidone copolymer with vinyl acetate (PVP-VA) or hydroxyl-propylmethylcellulose (HPMC), typically 5 mPa.Math.s, are dissolved in organic solvents such as ethanol, methanol methylene chloride. Suitably the polymer is dissolved in ethanol. The polymer and compound solutions are mixed and subsequently spray dried. The ratio of compound/polymer is selected from 1/1 to 1/6. Intermediate ranges can be 1/1.5 and 1/3. A suitable ratio can be 1/6. The spray-dried powder, a solid dispersion, is subsequently filled in capsules for administration. The drug load in one capsule ranges between 50 and 100 mg depending on the capsule size used.

(22) Film-Coated Tablets

(23) Preparation of Tablet Core

(24) A mixture of 100 g of a compound of formula (I), 570 g lactose and 200 g starch is mixed well and thereafter humidified with a solution of 5 g sodium dodecyl sulfate and 10 g polyvinylpyrrolidone in about 200 ml of water. The wet powder mixture is sieved, dried and sieved again. Then there is added 100 g microcrystalline cellulose and 15 g hydrogenated vegetable oil. The whole is mixed well and compressed into tablets, giving 10.000 tablets, each comprising 10 mg of the active ingredient.

(25) Coating

(26) To a solution of 10 g methylcellulose in 75 ml of denaturated ethanol there is added a solution of 5 g of ethylcellulose in 150 ml of dichloromethane. Then there is added 75 ml of dichloromethane and 2.5 ml 1,2,3-propanetriol. 10 g of polyethylene glycol is molten and dissolved in 75 ml of dichloromethane. The latter solution is added to the former and then there is added 2.5 g of magnesium octadecanoate, 5 g of polyvinyl-pyrrolidone and 30 ml of concentrated color suspension and the whole is homogenized. The tablet cores are coated with the thus obtained mixture in a coating apparatus.

(27) Antiviral Spectrum:

(28) Because of the increasing emergence of drug resistant HIV strains, the present compounds were tested for their potency against clinically isolated HIV strains harbouring several mutations. These mutations are associated with resistance to reverse transcriptase inhibitors and result in viruses that show various degrees of phenotypic cross-resistance to the currently commercially available drugs such as for instance AZT and delavirdine.

(29) The antiviral activity of the compound of the present invention has been evaluated in the presence of wild type HIV and HIV mutants bearing mutations at the reverse transcriptase gene. The activity of the compounds is evaluated using a cellular assay which was performed according to the following procedure.

(30) The human T-cell line MT4 is engineered with Green Fluorescent Protein (GFP) and an HIV-specific promoter, HIV-1 long terminal repeat (LTR). This cell line is designated MT4 LTR-EGFP, and can be used for the in vitro evaluation of anti-HIV activity of investigational compounds. In HIV-1 infected cells, the Tat protein is produced which upregulates the LTR promoter and finally leads to stimulation of the GFP reporter production, allowing to measure ongoing HIV-infection fluorometrically. Analogously, MT4 cells are engineered with GFP and the constitutional cytomegalovirus (CMV) promoter. This cell line is designated MT4 CMV-EGFP, and can be used for the in vitro evaluation of cytotoxicity of investigational compounds. In this cell line, GFP levels are comparably to those of infected MT4 LTR-EGFP cells. Cytotoxic investigational compounds reduce GFP levels of mock-infected MT4 CMV-EGFP cells.

(31) Effective concentration values such as 50% effective concentration (EC50) can be determined and are usually expressed in M. An EC50 value is defined as the concentration of test compound that reduces the fluorescence of HIV-infected cells by 50%. The 50% cytotoxic concentration (CC50 in M) is defined as the concentration of test compound that reduces fluorescence of the mock-infected cells by 50%. The ratio of CC50 to EC50 is defined as the selectivity index (SI) and is an indication of the selectivity of the anti-HIV activity of the inhibitor. The ultimate monitoring of HIV-1 infection and cytotoxicity is done using a scanning microscope. Image analysis allows very sensitive detection of viral infection. Measurements are done before cell necrosis, which usually takes place about five days after infection, in particular measurements are performed three days after infection.

(32) The columns IIIB, L100I, etc. in the table list the pEC.sub.50 values against various strains IIIB, L100I, etc.

(33) Strain IIIB is wild type HIV strain

(34) MDR refers to a strain that contains mutations L100I, K103N, Y181C, E138G, V179I, L2214F, V278V/I and A327A/V in HIV reverse transcriptase.

(35) TABLE-US-00003 L100I + K103N + IIIB L100I K103N K103N Y181C Y181C Y188L MDR Co. No pEC.sub.50 C pSi pEC.sub.50 pEC.sub.50 pEC.sub.50 pEC.sub.50 pEC.sub.50 pEC.sub.50 pEC.sub.50 22 9.10 3.90 7.70 7.00 9.20 7.70 7.80 7.30 5.70 38 9.10 3.70 7.60 7.20 9.00 7.80 7.70 7.60 5.70 55 9.00 4.00 7.80 7.10 8.40 7.00 7.70 7.20 5.70 12 8.90 4.10 8.60 7.70 8.60 7.90 7.80 8.00 6.00 6 8.80 > 4.20 6.80 5.90 8.30 6.40 7.00 6.50 5.00 7 8.80 4.00 8.10 7.30 8.70 7.20 7.70 7.30 5.70 3 8.70 4.00 8.30 7.90 8.50 7.20 7.80 7.60 5.60 11 8.70 3.40 8.10 7.30 8.40 7.10 7.40 7.60 5.80 44 8.70 > 4.10 7.90 7.50 8.60 8.00 8.00 8.10 5.90 9 8.60 > 4.00 8.30 7.60 8.50 7.70 7.90 7.80 6.00 10 8.60 > 4.00 8.40 7.40 8.50 7.20 7.80 7.70 6.30 14 8.60 > 4.00 8.60 7.80 8.60 8.00 7.90 7.90 6.20 54 8.60 > 4.00 8.40 7.80 8.70 7.90 8.00 8.00 6.00 37 8.60 3.60 8.30 7.70 8.80 8.10 7.90 7.90 6.00 45 8.60 > 4.00 8.20 8.70 7.80 7.70 6.00 2 8.50 > 3.90 7.20 6.70 7.90 6.30 6.90 6.40 5.00 53 8.50 > 3.90 8.30 7.60 8.20 7.50 7.70 7.50 6.30 15 8.50 3.70 7.90 7.20 8.50 7.70 7.80 7.70 6.70 19 8.40 3.60 8.40 7.30 8.40 7.70 7.70 7.70 6.00 23 8.40 > 3.80 8.30 7.90 8.10 7.80 7.80 6.10 51 8.40 3.00 7.10 6.90 8.20 7.20 7.20 7.00 5.60 5 8.30 > 3.70 7.60 7.30 8.00 6.80 7.00 7.00 5.40 20 8.30 3.60 7.80 7.00 8.30 7.50 7.40 7.30 5.60 41 8.30 > 3.70 8.10 7.30 8.40 7.70 7.80 7.10 5.80 27 8.20 > 3.60 7.30 8.10 7.40 7.40 7.10 5.50 46 8.10 > 3.50 8.10 7.80 8.40 7.40 7.50 6.80 26 7.90 3.00 7.60 6.50 7.90 7.40 7.30 7.10 5.30 25 7.90 3.00 7.60 7.40 7.80 7.50 7.10 7.30 5.00 32 7.90 3.10 7.40 7.80 7.10 7.10 7.00 5.00 30 7.90 > 3.30 7.00 6.60 7.80 7.00 7.00 6.50 5.30 43 7.90 > 3.30 7.30 6.90 8.00 7.10 7.60 7.10 5.50 21 7.80 2.80 8.40 7.80 8.30 7.80 7.80 7.80 6.30 24 7.80 3.00 7.60 7.10 7.30 7.60 7.10 5.30 31 7.80 3.10 7.50 7.80 7.30 7.20 7.10 5.00 28 7.80 3.10 7.30 7.80 7.30 7.10 6.90 5.10 29 7.80 3.00 7.10 6.90 7.70 7.00 7.10 6.80 5.00 33 7.80 > 3.20 7.80 7.80 7.80 7.90 7.30 7.20 6.40 49 7.80 3.00 7.70 7.20 7.80 7.30 7.10 7.10 6.30 47 7.70 3.10 7.20 6.90 7.60 6.90 7.30 7.00 6.30 48 7.70 3.10 6.90 6.30 7.70 6.70 6.90 6.50 5.00 16 7.60 2.60 7.10 6.30 8.40 6.50 6.70 6.30 5.10 36 7.60 > 3.00 6.90 6.10 7.60 6.90 6.90 6.40 5.00 58 8.70 > 4.10 6.50 8.20 7.00 6.80 5.50 59 8.60 3.80 7.20 6.60 8.20 7.10 7.50 6.60 5.40 60 8.50 3.50 7.10 6.50 7.90 7.20 7.50 6.80 5.00 61 8.50 > 3.90 7.70 7.30 8.40 7.60 7.70 7.00 5.50 62 8.80 4.00 8.30 7.40 8.60 7.50 7.70 7.40 5.50