AU735729B2 - Bisethers of 1-oxa, aza and thianaphthalen-2-ones as phospholamban inhibitors - Google Patents

Description

WO 99/15523 PCT/FI98/00755

I

BISETHERS OF 1-OXA, AZA AND THIANAPHTHALEN-2-ONES AS PHOSPHOLAMBAN INHIBITORS The present invention relates to new therapeutically active compounds and salts and esters thereof as well as new intermediates. The invention also relates to pharmaceutical compositions containing these compounds as active ingredients. The compounds of the invention have phospholamban inhibiting properties and are useful in the treatment of heart failure and stunned myocardium.

Compounds of the present invention have the structure represented by formulae or (II): 0' 0' R9 in which R1 is hydrogen, alkyl, alkenyl, aryl, arylalkyl, hydroxyalkyl, halogenalkyl, alkoxy, COR10, CONR1 R11, OR10, NR1 COR11 or NR1 R1 1, where R10 is hydrogen, alkyl, alkenyl, aryl, arylalkyl, hydroxyalkyl, halogenalkyl, alkoxy or hydroxy and R11 is hydrogen, alkyl, aryl, arylalkyl, alkoxy, aryloxy, hydroxy or acyl, or in case where X is

NR

1 1, can R1 also be carboxylalkyl, R6 is hydrogen, alkyl, alkenyl, aryl, arylalkyl, WO 99/15523 PCT/FI98/00755 2 R2 and R7 mean hydrogen, alkyl, aryl, arylalkyl, alkenyl, COR1 0 CONR1iR11, halogen, trifluoromethyl, nitro or cyano, where R10 and R11 are defined as above, R3 is hydrogen, alkyl, aryl or arylalkyl, A means alkyl or substituted alkyl, m is 0-2 and n is 1-3, Y means O, NR11 or S, where R11 is the same as above, X means O, NR11 or S, where R11 is the same as above, R4, R5, R8 and R9 mean independently one of the following groups: So- fT HN-

N

H 0 H 0^ N H HN,

N

O

I HC O NN

N-N

H

or in case where X is NR11, can R4, R5, R8 and R9 also independently mean HOOC-, R1200C-, H2NCO- or HOHNCO- wherein R12 means alkyl, arylalkyl or aryl, and wherein each aryl residue defined above by itself or as part of another group may be substituted, and pharmaceutically acceptable salts and esters thereof.

In one class of preferred compounds and pharmaceutically acceptable salts and esters thereof are compounds of formula wherein R2 is hydrogen. In a subclass of this class of compounds and pharmaceutically acceptable salts and esters thereof R1 is hydrogen, C1-6 alkyl, C2-6 alkenyl, C6-10 aryl, C7-12 arylalkyl, C1-6 hydroxyalkyl, C1-6 halogenalkyl or C1-6 alkoxy. In a group of this subclass of compounds and pharmaceutically acceptable salts and esters thereof, Y is O or S, preferably 0; and X is O. In another group of this subclass of compounds and pharmaceutically acceptable salts and esters thereof, Y is O or S, preferably 0; and X is NR11, where R11 is hydrogen, C1-6 alkyl, C6-10 aryl, C7-12 arylalkyl, C1-6 alkoxy, C6-10 aryloxy, hydroxy, C1-6 alkanoyl or Ci -6 carboxyalkyl. In a subgroup of these group of compounds and pharmaceutically acceptable salts and esters thereof, R3 is hydrogen, C1-6 alkyl, C6-10 aryl, or C7-12 arylalkyl, preferably C1-6 alkyl, most preferably methyl. In a family of these WO 99/15523 PCT/FI98/00755 3 subgroups of compounds and pharmaceutically acceptable salts and esters thereof, A is preferably straight-chain or branched C1-4 alkylene.

In another preferred class of compounds and pharmaceutically acceptable salts and esters thereof, compounds have formula (II) wherein R7 is hydrogen. In a subclass of this class of compounds and pharmaceutically acceptable salts and esters thereof R6 is hydrogen, C1-6 alkyl, C6-10 aryl, C7-12 arylalkyl; and n is 1,2 or 3, preferably 1 or 2. In a group of this subclass of compounds and pharmaceutically acceptable salts and esters thereof, Y is O or S, preferably 0; and X is O. In another group of this subclass of compounds and pharmaceutically acceptable salts and esters thereof, Y is O or S, preferably 0; and X is NR11, where R11 is hydrogen, C1-6 alkyl, C6-10 aryl, C7-12 arylalkyl, C1-6 alkoxy, C6-10 aryloxy, hydroxy, C1-6 alkanoyl or C1-6 carboxyalkyl. In a subgroup of these group of compounds and pharmaceutically acceptable salts and esters thereof, A is preferably straight-chain or branched C1-4 alkylene.

Each aryl residue in each of these preferred classes of compounds by itself or as part of another group, may be substituted by 1 to 3, preferably 1 or 2, most preferably one of fluorine, chlorine, bromine, iodine, trifluoromethyl, amino, C1-4 alkyl, C1-4 alkoxy, phenyl, naphthyl, halophenyl, halonaphthyl, benzyl, phenethyl, halobenzyl, halophenethyl, naphthylmethyl, naphthylethyl, C4-7 cycloalkyl, C1-4 alkyl (C4-7)cycloalkyl, hydroxy, mono- (C -4)alkylamino, di-(C1-4)alkylamino, C1-6 alkanoylamino, phenylcarbonylamino, naphthylcarbonylamino nitro, cyano, thiol, or C1-6 alkylthio.

Brief description of the drawings FIG. 1A shows the effect of the compound of Example 1c (50 and 100 pM) on the Ca 2 uptake rate into the cardiac muscle SR vesicles.

FIG. 1B shows the effect of the compound of Example 1c (50 and 100 pM) on the Ca 2 uptake rate into the fast skeletal muscle SR vesicles.

FIG. 2A shows the development of stunned myocardium and the subsequent decrease in the left ventricular systolic pressure.

FIG. 2B shows the complete inhibition of the development of stunned myocardium by the compound of Example 8g.

The compounds of the invention can be prepared from the 1,3dihydroxy substituted heteroaromatics by alkylation of the dihydroxy WO 99/15523 PCT/FI98/00755 4 compounds by suitable alkylating agents, for example by chloroacetonitrile or bromoacetic ester according to the following Scheme 1, wherein R1,R2, R3, X and Y are the same as defined above, R' is a protecting group for the hydroxyl, e.g. methyl, benzyl or tetrahydropyranyl.

SCHEME 1 OH R3 CK>

N

B O 0 Y 0

K

2 CO, DMF 60-120*C,

K

2

CO,,DMF

60-12OoC, 1-5 h

(IV)

o R3 0 x Y JN R2 NaN, NHC, DMF O R3 O R1 O X Y O R2 SNaOH or HCI

(VI)

80-120°C, 1-3 h

(VII)

The cyano compound (IV) described above is used to prepare the 1,2,4oxadiazole and 1,2,4-thiadiazole derivatives using the methods described in J.

Med. Chem. 1996, 39, 5228-5235.

The syntheses are shown in Scheme 2, wherein R1, R2, R3, X and Y are the same as defined above.

WO 99/15523 PCT/FI98/00755 SCHEME 2 O R3 R1 R (IV) R2 NHOH X HC, DMSD

(VIII)

ImCS, BF 3 xEt 2

H

KN 0r O R3 S-N R1 EtCOCI, DBU

(IX)

Cy-N The other heterocyclics as groups R4, R5, R8 and R9 are prepared as described in Bioorg. Med. Chem. Lett., 1994, 4, 45-50.

The dihydroxyaromatics (III) are made by use of the literature methods. The coumarins (XIV), (XVI) and (XX) are made by the use of the Knoevenagel condensation or von Pechmann reaction as presented in Scheme 3 and 4, where R1,R2 and R3, are the same as defined above, Z is alkyl, aryl, arylalkyl or alkenyl and R' is a protecting group for the hydroxyls e.g. methyl, benzyl or tetrahydropyranyl.

WO 99/15523 PTF9/05 PCT/F198/00755 6 SCHEME 3.

N

ZnCI 2 HCI, Et 2 0 Zn(Hg)-HCI

(XIII)

R3000H(Rl1)CO2Et

HOIBOH

(XII)

HBr

(XIV)

WO 99/1 5523 PCT/F]98/00755 7 SCHEME 4.

AOH R3 HJt OH R1

(XV)

0 HCI, EtOH OH R3 4 C, R~ 1

(XVI)

H 0 _0 (XVII) 4 R (XVIII) 14OH C001 R' R2 Pipendine or DBU

OR

Rl

RO"(::(XIX)

R 2 SHBr or pyridinex HCI

OH

R1 HO 0 0(XX) R 2 The quinolinones are prepared by the Knorr reaction as described in Scheme 5, wherein R1, R1 1 and R3 are the same as defined above, X is a halogen.

WO 99/15523 WO 99/ 5523PCT/F198/00755 8 SCHEME R3 -Ri 0 140-1600C

H

2 S0 4

(XXI)

(XXII)

(XXIII)

N

0 R3 1

H

NaH, R 11

X

N

0 R3 O06 N 0 4Pyndinex HCI OH R3 He N 0 Rll

(XIV)

(Xv) WO 99/15523 PCT/F198/00755 9 The cyclic compounds (II) can be prepared correspondingly from compound (XXXI) which can be prepared according to the Scheme 6, wherein R2 and R6 are the same as defined above, R' is a protecting group for the hydroxyls e.g. methyl, benzyl or tetrahydropyranyl.

SCHEME 6.

'OH

HCI, EtOH

(XXVI)

(XXVII)

R6

(XXIX)

B

(XXVIII)

HCI, EtOH

(XXX)

R' R2 HBr or pyridinex Ha CH 2 R6

(XXXI)

Cyclic quinolinone compounds (II) can be prepared correspondingly from (XXVI) using Scheme Salts and esters of the compounds, when applicable, may be prepared by known methods. Physiologically acceptable salts are useful as active medicaments, however, preferred are the salts with alkali or alkaline earth metals. Physiologically acceptable esters are also useful as active medicaments. Examples are the esters with aliphatic or aromatic alcohols.

The term "alkyl" as employed herein by itself or as part of another group includes both straight, branched and cyclized chain radicals of up to 18 carbon atoms, preferably 1 to 8 carbon atoms, most preferably 1 to 4 carbon atoms. The term "lower alkyl" as employed herein by itself or as part 1 of another group includes straight, branched and cyclized chain radicals of 1 to 7, preferably 1 to 4, most preferably 1 or 2 carbon atoms. Specific 15 examples for the alkyl and lower alkyl residues, respectively, are methyl, ethyl, propyl, isopropyl, butyl, tert. butyl, pentyl, cyclopentyl, hexyl, f *cyclohexyl, octyl, decyl and dodecyl including the various branched chain isomers thereof.

Throughout the description and claims of the specification the word "comprise" and variations of the word, such as "comprising" and "comprises", is not intended to exclude other additives, components, integers or steps.

The term "acyl" as employed herein by itself or as part of another group refers to an alkylcarbonyl or alkenylcarbonyl group, the alkyl and alkenyl groups being defined above.

The term "aryl" as used herein by itself or as part of another group refers to a monocyclic or bicyclic group containing from 6 to 10 carbon atoms in the ring portion. Specific examples for aryl groups are phenyl, naphthyl and the like.

"Aroyl" means in a corresponding way an arylcarbonyl group.

S

The term "alkoxy" as employed herein by itself or as part of another group 15 includes an alkyl group as defined above linked to an oxygen atom. "Aryloxy" means in a corresponding way an aryl group linked to an oxygen atom.

S S The term "substituted" as used herein in connection with various residues refers to halogen substituents, such as fluorine, chlorine, bromine, iodine or 20 trifluoromethyl group, amino, alkyl, alkoxy, aryl, alkyl-aryl, halogen-aryl, cycloalkyl, alkylcycloalkyl, hydroxy, alkylamino, alkanoylamino, arylcarbonylamino, nitro, cyano, thiol, or alkylthio substituents.

S

W:\NciI\spccnki\93 508-98 doc WO 99/15523 PCT/FI98/00755 11 The "substituted" groups may contain 1 to 3, preferably 1 or 2, most preferably 1 of the above mentioned substituents.

Compounds of the invention may be administered to a patient in therapeutically effective amounts which range usually from about 0.1 to 500 mg per day depending on the age, weight, condition of the patient, administration route and the phospholamban inhibitor used. The compounds of the invention can be formulated into dosage forms using the principles known in the art. It can be given to a patient as such or in combination with suitable pharmaceutical excipients in the form of tablets, dragees, capsules, suppositories, emulsions, suspensions or solutions. Choosing suitable ingredients for the composition is a routine for those of ordinary skill in the art. It is evident that suitable carriers, solvents, gel forming ingredients, dispersion forming ingredients, antioxidants, colours, sweeteners, wetting compounds and other ingredients normally used in this field of technology may be also used. The compositions containing the active compound can be given enterally or parenterally, the oral route being the preferred way. The contents of the active compound in the composition is from about 0.5 to 100 preferably from about 0.5 to about 20 per weight of the total composition.

EXPERIMENTS

Experiment 1. Effect on calcium uptake into the SR vesicles prepared from cardiac and fast skeletal muscle The inhibitory effect of a given compound on phospholamban can be demonstrated by measuring the effect of the compound on calcium uptake into the SR vesicles prepared from cardiac tissue and into SR vesicles prepared from fast skeletal muscle (psoas Both kind of SR vesicles contain Ca 2 +-ATPase but the vesicles from the fast skeletal muscle do not contain phospholamban (Hoh JFY, "Muscle fiber types and function", Current Opinion in Rheumatology, 4:801-808,1992). An increase in the calcium uptake into the SR vesicles prepared from cardiac tissue but not into the SR vesicles prepared from fast skeletal muscle indicates that the compound relieves the inhibitory effect of phospholamban on SR Ca 2 ATPase and thus acts as a phospholamban inhibitor. Since phospholamban represses both the rates of relaxation and contraction in the mammalian heart through its inhibitory effects on the cardiac SR Ca2+-ATPase, a WO 99/15523 PCT/FI98/00755 12 compound relieving these effects is potentially useful in the treatment of heart failure.

Method Guinea pigs (10-12) were decapited. Their hearts or the psoas muscles were excised, washed in ice-cold 0.9 NaCI and cut into pieces in a buffer containing 20 mM Tris-maleate, 0.3 M sucrose, pH 7.0. Thereafter tissue pieces were homogenized with Polytron and further with Potter strokes). The homogenate was centrifugated at 1000 g for 15 min at 4 °C.

The supernatant was collected and the pellet was resuspended into 5 ml of the buffer (20 mM Tris-maleate, 0.3 M sucrose, pH 7.0) and recentrifugated at 1000 g for 10 min at 4 oC. The obtained supernatant was combined with the earlier collected supematant and centrifugated once again at 10 000 g for 20 min at 4 The final supernatant was filtered into a bottle equipped with a magnetic stirrer. KCI was added to the filtered supernatant to achieve the final concentration of 0.6 M (at 4 The obtained solution was centrifugated at 100 000 g for 60 min at 4 The pellet was suspended in ml of the buffer containing 20 mM Tris-maleate, 0.3 M sucrose, pH 7.0 and centrifugated at 100 000 g for 60 min at 4 The obtained pellet was suspended in 5 ml of buffer containing 20 mM Tris-maleate, 0.3 M sucrose, 0.1 M KCI, pH 7.0 and stored at -80 °C until use. The protein concentration was also measured in order to standardise the separately prepared vesicle preparations.

In the calcium uptake assay, the fluorescent indicator, fluo-3 was used to detect the decrease of the extravesicular Ca2+-concentration, when the SR Ca2+ATPase was transferring Ca 2 from the extravesicular space into the SR-vesicles.

The SR-vesicles obtained above (50 pg protein/ml) were preincubated with or without the test compound at 37 °C for 5 min in the assay buffer containing 40 mM imidazole, 95 mM KCI, 5 mM NaN3, 5 mM MgCl 2 0.5 mM EGTA, 5 mM potassium oxalate, 2 pM ruthenium red, 5 pM fluo-3, pH 7.0. The free calcium was adjusted to 0.1 pM or to 0.04 pM by CaCl 2 The reaction was initiated by adding ATP (5 mM). The final reaction volume was 1.5 ml. The fluorescence of reaction mixture was measured for 3 min by using the excitation and emission wavelengths of 510 nm and 530 nm, respectively.

WO 99/15523 PCT/FI98/00755 13 Results Figures 1A and 1B show the effect of the compound of Example ic and 100 pM) on the Ca 2 uptake rate into the cardiac and fast skeletal muscle SR vesicles. It can be seen that the compound of the invention accelerated the calcium uptake into the cardiac SR vesicles but did not change the calcium uptake into the SR vesicle prepared from the fast skeletal muscle.

Table 1 shows the effects of various other phospholamban inhibitors of formula or (II) on the Ca 2 uptake rate into the cardiac and fast skeletal muscle SR vesicles. The experiments were carried out at 0.1 pM and 0.04 pM free calcium concentrations, respectively.

TABLE 1. Stimulation of the Ca 2 uptake into the vesicle preparations obtained from the ventricular myocardium and fast skeletal muscle of the guinea-pig heart.

Compound of The stimulation Example No. of Ca 2 uptake (100 pM) A B 3c** 51 0 2c 26 -1 7c 5 -17 8g* 18 0 11b 28 nd 12 32 nd 13d*** 23 nd 14c* 18 nd 18e 13 nd 21 11 nd 20 nd pM, **20 pM, pM, pM nd=not determined WO 99/15523 PCT/FI98/00755 14 Experiment 2. The effects on the left ventricular pressure derivatives Method Guinea-pigs of either sex weighing 300-400 g were used in the study.

After the guinea-pig was sacrificed by a blow on the skull and decapitated the heart was rapidly excised. The heart was then rinsed in cold oxygenated perfusion buffer. A cannula was inserted into the aorta and secured with a ligature. Retrograde perfusion began as soon as the heart was placed in a thermostatically controlled moist chamber of the Langendorff apparatus.

Modified Tyrode solution (37 equilibrated in a thermostatically controlled bulb oxygenator with carbogen (95 02 and 5% C02) was used as a perfusion buffer. The composition of the Tyrode solution was (in mM): NaCI 135; MgCI 2 x 6H 2 0 1; KCI 5; CaCl 2 x 2H 2 0 2; NaHC0 3 15; Na 2 HPO4 x 2H 2 0 1; glucose 10; pH 7.3-7.4. The experiments were carried out under constant pressure condition (50 mmHg). After a short prestabilization min) a latex balloon (size 4) was carefully placed into the left ventricle through the left pulmonary vein and the left atrium. The latex balloon was attached to a stainless-steel cannula coupled with a pressure transducer.

The latex balloon, the cannula and the chamber of the pressure transducer were filled with ethylene glycol water mixture avoiding any air-bubble.

The isovolumetric left ventricular pressure was recorded through the pressure transducer. At the beginning of the experiment, the volume of the balloon was adjusted to obtain a diastolic pressure of approximately mmHg. Before starting the experiment, the heart was allowed to stabilise further for 30 50 min with vehicle 1% DMSO) in the perfusion buffer.

After 15 min baseline recording various concentrations of the test compound were added to the perfusion buffer at 15 min intervals. The concentration range of 0.3 30 pM was tested. The vehicle concentration DMSO) was kept constant throughout the experiment.

Results The EC 50 values and maximum effects change from baseline) of various compounds of the invention on left ventricular systolic pressure are given in Table 2.

WO 99/15523 PCT/FI98/00755 TABLE 2. The EC 5 o values and maximum effects change from baseline) on left ventricular systolic pressure.

Compound of EC50 maximum Example No. (pM) effect ic 9 +52 at 30 pM 3c 4 +63 at 10 pM 5c >10 +14 at 30 pM 6c 0.5 +25 at 10 pM 7c 2.5 +29 at 10 pM 8g 2 +64 at 10 pM 9d 5 +50 at 30 pM 12 5 +22 at 10 pM 13d 10 +48 at 30 pM 14c 1.5 +25 at 10 pM 3 +37 at 10 pM 16c 10 +57 at 30 pM 18e 10 +35 at 30 pM 19e 6 +39 at 30 pM Experiment 3. Effect on the development of stunned myocardium in isolated guinea-pig Langendorff heart Method Guinea-pigs of either sex weighing 300-400 g were used in the study.

After the guinea-pig was sacrificed by a blow on the skull and decapitated the heart was rapidly excised. The heart was then rinsed in oxygenated perfusion buffer. A cannula was inserted into the aorta and secured with a ligature. Retrograde perfusion began as soon as the heart was placed in a thermostatically controlled moist chamber of the Langendorff apparatus.

Modified Tyrode solution (37 equilibrated in a thermostatically controlled bulb oxygenator with carbogen (95 02 and 5% C02) was used as a perfusion buffer. The composition of the Tyrode solution was (in mM): NaCI 135; MgCI 2 x 6H 2 0 1; KCI 5; CaCl 2 x 2H 2 0 2; NaHC03 15; Na 2

HPO

4 x 2H 2 0 1; glucose 10; pH 7.3-7.4. The experiments were carried out under constant pressure condition (50 mmHg). After a short prestabilization min) a latex balloon attached through the stainless-steel cannula to a pressure transducer was carefully placed into the left ventricle through the left pulmonary vein and the left atrium. The latex balloon, the cannula and WO 99/15523 PCT/F198/00755 16 the chamber of the pressure transducer were filled with ethylene glycol water mixture avoiding any air-bubble. The isovolumetric left ventricular pressure was recorded through the pressure transducer. At the beginning of the experiment, the volume of the balloon was adjusted to obtain the enddiastolic pressure of approximately 5 mmHg. Before starting the experiment, the spontaneously beating heart was allowed to stabilise further for 30 min with vehicle DMSO) in the perfusion buffer.

After 15 min baseline recordings compound of Example 8g (10 pM) was added to the perfusion buffer. The heart was 15 min later exposed to the 8 minute period of global ischemia followed by reperfusion. This procedure was then repeated twice at 35 min intervals. Another series of experiments was performed with vehicle instead of compound of Example 8g. The vehicle concentration DMSO) was kept constant throughout the experiments. The baseline value was the average of the two minute recordings obtained just before compound of Example 8g or vehicle was added to the perfusion buffer. The preischemia values were the average of the two minute recordings obtained just before each ischemia period and the reperfusion values were the average of the two minute recordings obtained at 8 min during each reperfusion period.

The results are shown in Figures 2 A and B. Figure 2A shows the development of stunned myocardium and the subsequent decrease in the left ventricular systolic pressure in the control group. Figure 2B shows that the phospholamban inhibitor of Example 8g completely inhibited the development of stunned myocardium. Givens are mean SEM of 2-3 experiments.

EXAMPLES

Example 1. Preparation of 3-Benzyl-5,7-bis[(1 H methoxy]-4-methyl-2H -1 -benzopyran-2-one a) 3-Benzyl-5,7-dihydroxy-4-methyl-2H -1-benzopyran-2-one WO 99/15523 PCT/FI98/00755 17

OH

HO 0 O A solution of phloroglucinol dihydrate (20 g) and ethyl 2-benzylacetoacetate (27.5 ml) in ethanol (320 ml) was treated with dry HCI at 0°C for five hours and the solution was kept at that temperature overnight. The yellow solution was concentrated and triturated with water, the solids filtered, washed with water and dried. The resulting hydrate was thrice evaporated to dryness from toluene, triturated with pethroleum ether (bp. 40-60°C) and filtered. Yield 33,4 g (96 Melting point 258-260 °C.

1 H-NMR (DMSO-d 6 400MHz): 2.525 3H, CH3), 3.887 2 H, CH2Ph), 6.171 1H, J 2,4 Hz), 6.274 1H, J 2,4 Hz), 7.167-7.279 5H, Ph), 10.2 1H, OH), 10.47 1H, OH).

b) 3-Benzyl-5,7-bis(cyanomethoxy)-4-methyl-2H-1-benzopyran-2-one 0 0 0 Chloracetonitrile (6.86 potassium carbonate (23.9 g) and 12.2 g of the product from example la were stirred in 120 ml of DMF at 100°C under nitrogen for two hours. The reaction mixture was cooled and poured into ice water. The solids were filtered and washed with water. Yield 13.8 g (88 Melting point 147-154°C.

1 H-NMR (DMSO-d 6 400MHz): 2.525 3H, CH3), 3.969 2H, CH2Ph), 5.307 2H, OCH2CN), 5.314 2H, OCH2CN), 6.814 1 H, J Hz), 6.940 1H, J 2.5 Hz), 7.18-7.292 5H, Ph).

WO 99/15523 PCT/FI98/00755 18 c) 3-Benzyl-5,7-bis[(1 H -tetrazol-5-yl)methoxy]-4-methyl-2H -1benzopyran-2-one The product from example 1b (1 sodium azide (0.42 g) and ammonium chloride (0.34 g) were stirred in DMF (5 ml) under nitrogen at 100 °C for 5 hours. The reaction mixture was allowed to cool down and then poured into ice water. The pH of the solution was adjusted to 10-11 and then the solution either extracted once with ethyl acetate or filtered through CELITE. The solution was acidified to pH 2 with hydrochloric acid, kept at 5°C and filtered. Yield 0.96 g (81 Melting point 229-233°C.

1 H-NMR (DMSO-d 6 400MHz): 2.468 3H, CH3), 3.937 2H, CH2Ph), 5.596 2H, OCH2Tet), 5.602 2H, OCH2Tet), 6.832 1H, J 2.4 Hz), 6.851 1H, J 2.4 Hz), 7.171-7.283 5H, Ph).

Example 2. Preparation of 7,8,9,10-Tetrahydro-1,3-bis[(1 H yl)methoxy]-7-phenyl-6H-dibenzo[b,d]pyran-6-one a) 7,8,9,10-Tetrahydro-1,3-dihydroxy-7-phenyl-6H-dibenzo[b,d]pyran- 6-one A solution of phloroglucinol (0.7 g) and 2-ethoxycarbonyl-3phenylcyclohexanone (1,5 g) in ethanol was treated with dry HCI as described in example 1 a. The product was first recrystallized from ethanolwater and then triturated with ether. Yield 0.61 g.

WO 99/15523 PCT/F198/00755 19 1 HNMR (DMSO-d 6 400MHz): 1.38-1 .52 (in, 1 1.57-1.66 (in, 1 H), 1.69-1.78 (in, 1 1. 86-1.96 (in, 1 2.9-3.02 (in, 1 3.3-3.4 1H), 4.050 1 6.157 1 H, J 2.4 Hz), 6.297 1 H, J 2.4 Hz), 7.076- 7.265 (in, 5 10. 153 1 10.456 1 H).

b) 7,8,9,1 0-Tetrahydro-1 ,3-bis(cyanomethoxy)-7-phenyl-6Hdibenzo[b,d]pyran-6-one The product from example 2a (0.5 g) was treated with chioroacetonitrile (0.25 g) and potassium carbonate (1.12g) in DMF (5 ml) as described in example l b. Yield 0.6 g.

1 H-NMR (DMSO-d 6 400MHz): 1.38-1.58 (in, 1 1.6-1.7 (in, 1 H), 1.7-1.76 (in, 1 1.89-1.99 (in, 1 2.9-3.03 (in, 1 3.2-3.28 (m 1 H), 4.111 1 5.314 2H), 5.349 2H), 6.840 1 H, J 2.5 Hz), 6.925 1 H, J 2.5 Hz), 7.108-7.274 (in, c) 7,8,9,1 0-Tetrahydro-1 ,3-bis[(1 H -tetrazol-5-yl)methoxy]- 7-phenyl- 6H-dibenzo[b,d]pyran-6-one The product from example 2b (0.6 g) was treated with sodium azide (0.2 g) and ammonium chloride (0.17 g) in DMF (5 ml) as in example 1c. The product was recrystallized from a mixture of DMVF, ethanol and water (approximately Yield 0.41 g. Melting point: 153-1 5400.

WO 99/15523 PCT/FI98/00755 1 H-NMR (DMSO-d 6 400MHz): 1.38-1.5 1 1.5-1.6 1 H), 1.69-1.76 1 1.87-1.96 1 2.9-3.05 1 3.2-3.3 1 H), 4.094 1H), 5.602 2H), 5.643 2H), 6.832 1H, J 2.3 Hz), 6.851 1H, J 2.3.Hz), 7.089-7.212 Example 3. Preparation of 3-Benzyl-5,7-bis[( 2,5-dihydro-5-oxo-4H 1,2,4-oxadiazol-3-yl)- methoxy]-4-methyl-2H -1-benzopyran-2-one a) 3-Benzyl-5,7-bis[(hydroxyamidino)methoxy]-4-methyl-2H-1benzopyran-2-one

HO

0 0

H

2

N

N'OH

Triethylamine (1.94 ml) was added to a suspension of hydroxylamine hydrochloride (0.97 g) in DMSO (2 ml) and the resulting mixture stirred at room temperature for thirty minutes. The crystals were filtered and washed with THF. The filtrate was concentrated and the product from example 1b g) added. This solution was kept at 75 °C overnight. The reaction mixture was treated with ice water, the pH adjusted to 11 and the solids filtered, washed with water, and dried. Yield 0.5 g. Melting point: 155-1600C.

1 H-NMR (DMSO-d 6 400MHz): 2.56 3H, CH3), 3.938 2H), 4.466 2H), 4.486 2H), 5.565 H, NH2), 5.709 2H, NH2), 6.658 1H, J 2.4 Hz), 6.692 1H, J 2.4 Hz), 7.168-7.284 5H, Ph), 9.346 1H, OH), 9.362 1H, OH).

b) 3-Benzyl-5,7-bis[(ethoxycarbonyloxyamidino)methoxy]-4-methyl- 2H -1 -benzopyran-2-one WO 99/15523 PCT/FI98/00755 21

H

2 N 0 o' N Ethyl chloroformiate (0.45 ml) was added to a solution of the product from example 3a (1 g) and pyridin (0.38 ml) in DMF (5 ml) at 0°C. The reaction mixture was kept at that temperature for an additional 30 minutes and then ice water added. The solids were filtered and washed with water.

Yield 1.63 g. Melting point 87-92°C.

1 H-NMR (DMSO-d 6 400MHz): 1.215-1.256 6H), 2.553 3H), 3.947 2H), 4.140-4.198 4H), 4.566 2H), 4.599 2H), 6.688 (d, 1 H, J 2.4 Hz), 6.718 1H, J 2.4 Hz), 6.792 2H, NH2), 6.818 2H, NH2), 7.171-7.285 c) 3-Benzyl-5,7-bis[( 2,5-dihydro-5-oxo-4H -1,2,4-oxadiazol-3-yl)methoxy]-4-methyl-2H -1 -benzopyran-2-one

H

N 0

N

0 00 0-N The product from the previous example (1.5 g) and DBU (0.8 ml) in DMF (5 ml) was stirred at room temperature overnight. The reaction mixture was treated with ice water and acidified. The solids were filtered and washed with water. The resulting solid mass was taken in 0.1 N sodium hydroxide WO 99/15523 PCT/FI98/00755 22 solution, treated with activated carbon and finally acidified. Yield 0.64 g.

Melting point: 130-1360C.

1 H-NMR (DMSO-d 6 400MHz): 2.524 3H), 3.954 2H), 5.187 (s, 2H), 5.215 2H), 6.748 1H, J 2.4 Hz), 6.834 1H, J 2.4 Hz), 7.158-7.289 5H), 12.8 2H).

Example 4. Preparation of 7,8,9,10-Tetrahydro-bis[(1 H yl)methoxy] -1,3-dihydroxy-6H-dibenzo[b,d]pyran-6-one a) 7,8,9,10-Tetrahydro-1,3-dihydroxy-6H-dibenzo[b,d]pyran-6-one

OH

HO 0 O Phloroglucinol (1 g) and ethyl 2-oxocyclohexane carboxylate (1.32 g) were stirred in 75 sulfuric acid (10 ml) overnight, the mixture poured into ice water and filtered. Yield: 1.55 g.

1 H-NMR (DMSO-d 6 400MHz): 1.65 4H), 2.345 2H), 3.037 (b, 2H), 6.138 1H, J 2.4 Hz), 6.245 1H, J= 2.4 Hz), 10.069 1H, OH), 10.322 1H, OH).

b) 7, 8, 9, 10-Tetrahydro-bis(cyanomethoxy)-1,3-dihydroxy-6Hdibenzo[b,d]pyran-6-one 0 0 The product from the previous example (0.5 chloroacetonitrile (0.34 g) and potassium carbonate (1.5 g) in DMF (5 ml) were reacted as in example lb. Yield: 0.44 g.

1 H-NMR (DMSO-d 6 400MHz): 1.68 4H), 2.41 2H), 3.00 (b, 2H), 5.297 2H), 5.309 2H), 6.797 1 H, J 2.4 Hz), 6.899 1 H, J 2.4 Hz).

WO 99/15523 PCT/F198/00755 23 c) 7,8,9,1 0-Tetrahydro-bis[( 1 H -tetrazol-5-yl)methoxy] -1 ,3-dihydroxy- 6H-dibenzo[b,d]pyran-6-one "I 0 q~ I

N-N

H

1 The product from the previous example (0.4 g) was treated with sodium azide 18 g) and ammonium chloride (0.14 g) in DMVF (2.5 ml) as in example 1c. The product was recrystallized from ethanol-DMF Yield 0. 17 g. Melting point 283-2860C.

1 H-NMVR (DMSO-d 6 400MHz): 1.626 4H), 2.393 2H), 2.971 (b, 2H), 5.583 2H), 5.599 2H), 6.811 2H).

Example 5. Preparation of 5,7-Bis[(1 H-tetrazol-5-yl)methoxyl-4phenyl-2 H-i -benzopyran-2-one a) 5 ,7-Dihydroxy-4-phenyl-2 H-i -benzopyran-2-one

I

OH

HOA000 A solution of phloroglucinol (2.00 g) and ethyl benzoylacetate (3.05 g) in ethanol (30 ml) was treated with dry HOI as described in example 1la. The product was recrystallized from ethanol-water Yield 3.0 g (75 1 H-NMR (DMSO-d6, 300 MHz): 5.739 1 H, CH=C), 6.155 1 H, J =2.3 Hz), 6.263 1 H' J 2.3 Hz), 7.305-7.381 (in, 5H, Ph), 10.084 1 H, OH), 10.368 1 H, OH).

b) 5,7-Bis(cyanomethoxy)-4-phenyl-2H- 1 -benzopyran-2-one WO 99/15523 PCT/FI98/00755 24 N 0 The product from previous example (1.00 g) was treated with chloroacetonitrile (0.62 g) and potassium carbonate (2.72 g) in DMF (5 ml) as described in example lb. The reaction mixture was poured into ice water and the mixture extracted with ethyl acetate. Ethyl acetate was washed with 1 M NaOH, dried with sodium sulfate and evaporated. The product was recrystallized from isopropanol. Yield 0.41 g (31 1 H-NMR (DMSO-d6, 300 MHz): 4.845 2H, OCH2CN), 5.344 (s, 2H, OCH2CN), 6.086 1H, CH=C), 6.770 1H, J 2.4 Hz), 7.040 1H, J 2.4 Hz), 7.320-7.443 5H, Ph).

c) 5,7-Bis[(1 H-tetrazol-5-yl)methoxy]-4-phenyl-2H-1 -benzopyran-2one HN N ;ZO HN 0 0

N=N

The product from previous example (0.40g) was treated with sodium azide (0.16 g) and ammonium chloride (0.14 g) in DMF (2 ml) at 100 °C for 2 hours. The product was isolated as described in example ic. Yield: 0.40 g (79 Melting point 222-224 °C.

1 H-NMR (DMSO-d6, 400 MHz): 5.148 2H, OCH2Tet), 5.649 (s, 2H, OCH2Tet), 5.968 1 H, CH=C), 6.811 1 H, J 2.3 Hz), 6.962 (d, 1H, J 2.3 Hz), 6.994-7.185 5H, Ph).

Example 6. Preparation of 7,8,9,10-Tetrahydro-1,3-bis[(1 yl)methoxy]-8-phenyl-6H- dibenzo[b,d]pyran-6-one WO 99/15523 PCT/FI98/00755 a) 7,8,9,10-Tetrahydro-1,3-dihydroxy-8-phenyl-6H-dibenzo[b,d]pyran- 6-one H O A solution of phloroglucinol (1.56 g) and ethyl hexane-carboxylate (2.52 g) in ethanol (25 ml) was treated with dry HCI as described in example la. The precipitate was filtered and washed with water and EtOH. Yield 1.0 g (32 1 H-NMR (DMSO-d6, 400 MHz): 1.72-1.82 1H), 2.01 1H), 2.317-2.387 1H), 2,707-2,763 1H), 2,830 1H), 3,041 1H), 3.35 and 3.40 1H), 6.174 1H, J 2.3 Hz), 6.277 1H, J 2.3 Hz), 7.200- 7.350 5H, Ph), 10.131 1H, OH), 10.401 1H, OH).

b) 7,8,9,10-Tetrahydro-1,3-bis(cyanomethoxy)-8-phenyl-6Hdibenzo[b,d]pyran-6-one

N

o 0o The product from previous example (1.0 g) was treated with chloroacetonitrile (0.57 g) and potassium carbonate (1.0 g) in DMF (5 ml) as described in example 1b. DMF was evaporated and residue dissolved in EtOAc. Ethyl acetate was washed with 1 M NaOH, dried with sodium sulfate and evaporated. The product was recrystallized from acetone-isopropanol Yield 0.50 g (40 1 H-NMR (DMSO-d6, 300 MHz): 1.75-1.88 1H), 2.05 1H), 2.38- 2.48 1H), 2.77-2.85 1H), 2.90 1H), 3.07 1H), 3.22 and 3.28 (b, 1 5.316 2H, OCH2CN), 5.331 2H, OCH2CN), 6.829 1 H, J Hz), 6.939 1H, J 2.5 Hz), 7.210-7.380 5H, Ph).

WO 99/15523 PCT/F198/00755 26 c) 7,8,9,1 0-Tetrahydro- 1, 3-bis[( 1 H-tetrazol-5-yl)methoxy]-8-phenyl- 6H- dibenzo[b,d]pyran-6-one Th rdutfo peiuseape 03 a retdwthsdu aid g ad mmNumclrd(00g)iDM(2m)a100fo 3. orThe product wasm peiosolapilte same0 mane was aeinh xapleic.

Yield 0.30 g (82 Melting point 235-245 00.

1 H-NMR (DMSO-d6, 400 MHz): 1.70-1.80 (in, 1 1.96 1 2.38- 2.446 (in, 1 2.836 (in, 2H), 3.052 1 3.252 and 3.301 1 5.604 2H, OCH2CN), 5.632 2H, OCH2CN), 6.827 1 H, J 2.5 Hz), 6.858 1 H, J 2.5 Hz), 7.209-7.35 1 (mn, 5H, Ph).

Example 7. Preparation of 5,7-Bis[(1 H-tetrazol-5-yl)methoxy]-4methyl-3-(2-phenylethyl)-2H-1 benzopyran-2-one a) 5,7-Dihydroxy-4-methyl-3-(2-phenylethyl)-2H-1 -benzopyran-2-one &0 A solution of phloroglucinol (0.87 g) and ethyl 2-(2-phenylethyl)acetoacetate (1.62 g) in ethanol (30 ml) was treated with dry HCl as described in example 1la. Yield: 1.77 g (87 Melting point 248-252 00.

1 H.NMR (DMSO-d6, 300 MHz): 2.413 3H, 0H-3), 2.652-2.782 (in, 4H, 0H20H2), 6.151 1 H, J 2.4 Hz), 6.256 1 H, J 2.4 Hz), 7.183- 7.304 (in, 5H, Ph), 10.137 1 H, OH), 10.369 1 H, OH).

b) 5,7-Bis(cyanomethoxy)-4-methyl-3-(2-phenylethyl)-2H-1 benzopyran-2-one WO 99/15523 PCT/F198/00755 27 <0 N 0 0 The product from previous example (0.90 g) was treated with chloroacetonitrile (0.48 g) and potassium carbonate (2.1 g) in DMF (5 ml) at 100 00 for 0.5 hours. The product was isolated as described in example l b.

Yield 1.00 g (88 Melting point 179-1 83 0

C.

1 H-NMR (DMSO-d6, 300 MHz): 2,384 3H, CH3), 2.699-2,754 (in, 2H, CH2CH2), 2.805-2.841 (in, 2H, 0H20H2), 5,302 4H, OCH2CN), 6,790 1 H, J 2.5 Hz), 6.909 1 H, J 2.5 Hz), 7.190-7.307 (in, 5H, Ph).

c) 5,7-Bis[( 1 H-tetrazol-5-yi)methoxy]-4-methyl-3-(2-phenylethyl)-2H- 1 benzopyran-2-one

N-I

The~~1 prdc Nrmpeiu xml 04g a rae ihsdu azide (.15 g) Nmoimclrd 01 nDF( t100 o 2. orThe product wasm peisolaeus dsbdi example (04g a rae i. Yiu038 g (78 Melting point 248-250 00.

1 H-NMR (DMSO-d6, 400 MHz): 2.368 3H, CH3), 2.668-2.707 (in, 2H, CH2CH2), 2.783-2.822 (in, 2H, 0H20H2), 5.593 2H, OCH2Tet), 5.604 2H, OCH2Tet), 6.819 1 H, J 2.3 Hz), 6.834 1 H, J 2.3 Hz), 7.161-7.291 (in, 5H, Ph).

Example 8. Preparation of 5,7-Bis[(1 H-tetrazol-5-yI)methoxy]-1 ,3dibenzyl-4-inethyl-2( 1 H)-quinolinone a) 2-Benzyl-3-oxobutanoic acid WO 99/15523 PCT/FI98/00755 28 O N O I

H

(5 g) was added in portions to a preheated (160 ethyl 2-benzyl acetoacetate (15 ml) under nitrogen and kept at that temperature for 60 minutes. The cooled solution was diluted with heptaneethyl ether and filtered. Yield 5.2 g (49 1 -H-NMR (DMSO-d6, 300 MHz): 2.183 3H), 3.069 2H, J 7.2 Hz), 3.923 1 H, J 7.2 Hz), 6.616 (dd. 1 H, J 2.3 Hz), 6.765 2H, J 2.3 Hz), 7.13- 7.3 5H), 10.123 1H).

b) 3-Benzyl-5,7-dimethoxy-4-methyl-2(1 H)-quinolinone 0 N 0 I H The product from the previous example (1.2 g) was added to a preheated (85 methanesulfonic acid (3.5 ml) and kept at that temperature for 15 minutes. The solution was allowed to cool and then treated with ice water. The product was filtered, washed with sodium bicarbonate and water. Yield 1.08 g (95 1 -H-NMR (300 MHz):2.486 3H), 3.785 3H), 3.808 3H), 3.985 2H), 6.315 1H, J 2.4 Hz), 6.472 1H, J 2.4 Hz), 7.1-7.3 5 H), 11.52 1 H).

c) 3-Benzyl-5,7-dihydroxy-4-methyl-2(1 H)-quinolinone

OH

H N 0

H

The product from the previous example (1 g) was refluxed under nitrogen in pyridine hydrochloride (5 g) for twenty minutes. The reaction WO 99/15523 PCT/FI98/00755 29 mixture was treated with water and the product filtered. Yield 0.9 g (100 Melting point: 307 312 oC.

1 -H-NMR (300 MHz):2.503 3H), 3.942 2H), 6.102 1H, J 2.3 Hz), 6.187 1H, J 2.3 Hz), 7.1-7.25 5H), 9.725 1H), 9.984 (s, 1H), 11.285 1H).

d) 1,3-Dibenzyl-5,7-dimethoxy-4-methyl-2(1 H)-quinolinone

O

N 0 The product from the example 8b (1 potassium t-butoxide (0.62 g) and benzyl bromide (0.68 ml) were stirred in DMSO (10 ml) at 60 °C for 4 hours. The reaction mixture was treated with water, extracted with toluene and evaporated. The product was triturated with ethyl ether and filtered.

Yield 0.5 g (39 1 -H-NMR (400 MHz):2.537 3H), 3.708 3H), 3.826 3H), 4.124 2H), 5.56 2H), 6.413-6.434 2H), 7.154- 7.332 e) 1,3-Dibenzyl-5,7-dihydroxy-4-methyl-2(1 H)-quinolinone.

OH

H N 0 The product from the previous example (2 g) was treated with pyridine hydrochloride (10 g) as described in example 8c. The product was extracted with ethyl acetate and evaporated. Yield 1,4 g (75 1 -H-NMR (400 MHz):2.570 3H), 4.076 2H), 5.450 2H), 6.135 1H, J 2.2 Hz), 6.199 1H, J 2.2 Hz), 7.128 7.333 10 9.83 1 10.166 1 H).

f) 5,7-Bis(cyanomethoxy)-1,3-dibenzyl-4-methyl-2(1 H)-quinolinone.

WO 99/1 5523 PCT/F198/00755 The product from the previous example (1.4 g) was treated with chloroacetonitrile (0.76 g) and K2003 (2.5 g) in DMF (20 ml) as described in example 1 b. Yield 1.5 g (89 1 -H-NMR (400 MHz):2.555 3H), 4.146 2H), 5.214 2H), 5.275 2H), 5.578 2H), 6.735 2H), 7.13-7.33 (in, g) 5,7-Bis[(l1H-tetrazol-5-yl)methoxy]- 1,3-dibenzyl-4-methyl-2(l1H)quinolinone.

The product from the previous example (1 .3 g) was treated with sodium azide (0.41 g) and ammonium chloride (0.34 g) as described in example 1ic. Yield: 0.69 g (45 1 -H-NMR (400 MHz):2.471 3H), 4.113 2H), 5.477 2H), 5.55 2H1), 5.574 2H), 6.670 1 H, J 2.1 Hz), 6.775 1 H, J 2.1 Hz), 7.13-7.32 (in, 10 H).

Example 9. Preparation of 5,7-Bis[(1 H-tetrazol-5-yl)methoxy] -3benzyl- 1,4-dimethyl-2( 1 H)-quinolinone.

a) 3-Benzyl-5,7-dimethoxy-1 ,4-dimethyl-2( 1 H)-quinolinone.

WO 99/15523 PCT/F198/00755 31 The product from example 8b (0.5 t-BuOK (0.2 g) and methyl iodide (0.4 ml) were stirred in DMSO (5 ml) at 35 oC for two days. The reaction mixture was treated with water and extracted with toluene. The product was purified by column chromatography using toluene-ethyl acetateacetic acid 8 2: 1 as the eluent. Yield 0.24 g (46 1 -H-NMR (300 MHz):2.51 3H), 3.632 2H), 3.846 3H), 3.896 3H), 4.047 2H), 6.468 1 H, J 2.3 Hz), 6.558 1 H, J 2.3 Hz), 7.1-7.26 b) 3-Benzyl-5,7-dihydroxy-1,4-dimethyl-2(1 H)-quinolinone.

OH

1 The product from the previous example (0.2 g) was treated with pyridine hydrochloride (2 g) as described in example 8c and the product extracted with ethyl acetate. Yield 0.16 g (89 1 -H-NMR (400 MHz):2.567 3H), 3.515 3H), 4.005 2H), 6.244 1 H, J 2.3 Hz), 6.268 1 H, J 2.3 Hz), 7.08-7.25 5H), 9.879 (s, 1H), 10.113 (s,1H).

c) 5,7-Bis(cyanomethoxy)-3-benzyl-1,4-dimethyl-2(1 H)-quinolinone.

N 0 The product from the previous example (0.15 chloroacetonitrile 0.08 g) and K2CO3 (0.28 g) were reacted in DMF (2 ml) as described in example lb. Yield 0.16 g (84 1 -H-NMR (400 MHz): 2.524 3H), 3.658 3H), 4.079 2H), 5.292 2H), 5.379 2H), 6.766 1 H, J 2.3 Hz), 6.855 1 H, J 2.3 Hz), 7.13-7.24 (m WO 99/15523 PCT/FI98/00755 32 d) 5,7-Bis[(1 H-tetrazol-5-yl)methoxy] -3-benzyl-1,4-dimethyl-2(1 H)quinolinone.

HN N O

N

N-NH

The product from the previous example (0.15 g) was treated with NaN3 (57 mg) and NH4CI (47 mg) in DMF (2 ml) as described in example ic. Yield 0.115 g. Melting point: 250-253°C.

1 -H-NMR (400 MHz): 2.451 3H), 3.649 3H), 4.042 2H), 6.792 1H, J 2.2 Hz), 6.833 1H, J Hz), 7.1-7.25 Example 10. Preparation of 3-Benzyl-5,7-bis[(2-methyl-1 H yl)methoxy]-4-methyl-2H-1-benzopyran-2-one and the three isomers.

N T

-N

0 00 N I 0.07 ml of methyl iodide was added to a solution of 0.2 g of the product from example 1c and 0.31 g of K2CO3 in 2 ml of DMF and the mixture stirred at room temperature for 4 hours. The reaction mixture was poured into ice water and filtered. Yield 0.2 g as a mixture of four regioisomers, melting point 71-76oC.

1 H-NMR (DMSO-d 6 400MHz): 2.47 CH3), 2.48 CH3), 3.93 (s, CH2Ph), 4.11 NCH3), 4.12 NCH3), 4.15 NCH3), 4.38 NCH3), 4.40 NCH3), 5.51 OCH2), 5.52 OCH2), 5.62 OCH2), 5.67 (s, OCH2), 6.84-6.91 2H), 7.16-7.28 (m,5H, Ph).

WO 99/15523 PCT/F198/00755 33 Example 11. Preparation of 3-Benzyl-5,7-bis[1-(1 H yl)ethoxy]4-methyl-2 H -1-benzopyran-2-one, mixture of stereoisomers a) 3-Benzyl-5,7-bis-[(1-cyano)ethoxy)-4-methyl-2 H -1-benzopyran- 2-one The product from example la (1 2-chlorpropionitrile (0.7 g) and potassium carbonate (2 g) were heated in DMF (15 ml) under nitrogen at 110°C for sixty minutes. The mixture was treated with water, filtered and washed with 1 N NaOH and water. Yield 1.2 g.

1 H-NMR (DMSO-d 6 300MHz): 1.74-1.78 (t t, 6 H, CH-CH3), 2.53 3 3.97 2H), 5.58-5.66 2H, CH-CH3), 6.87 1H), 6.99 1H), 7.18-7.31 b) 3-Benzyl-5,7-bis[1-(1 H -tetrazol-5-yl)ethoxy]4-methyl-2 H-1benzopyran-2-one, mixture of stereoisomers.

The product from the previous example (0.5 sodium azide (0.18 g) and ammonium chloride (0.15 g) were heated in DMF (7 ml) at 100 °C for minutes. The product was treated with water, extracted with ethyl acetate and evaporated. Yield 0.57 g. Melting point 91-1040C.

WO 99/15523 PCT/F198/00755 34 1 H-NMR (DMSO-d 6 300MHz): 1.69-1.77 (in, 6 H, CH-0H3), 2.54 (s, 3H), 3.94 2H), 6.10-6.17 2H, CH-0H3), 6.65 (dd, 1 6.74 (dd, 1 H), 7.13-7.30 (in, Example 12. Preparation of 5,7-Bis(carboxymethoxy)- 1,3-dibenzyl-4methyl-2(1 H)-quinolinone HO,,r 0

OH

The product from example 8f (0.2 g) was ref luxed in a solution of concentrated hydrochloric acid (3 ml) and acetic acid (2 ml) for one hour.

The product was filtered at 25 00. Yield 0.14 g.

1 H-NMR (300 Mhz, DMSO-d6): 2.63 0H-3), 4.14 2H, CH2Ph), 4.66 (s, 2 H, OCH2000H), 4.79 2H, OCH2000H), 5.53 2H, NCH2Ph), 6.41 1 H, J 2.2 Hz), 6.45 1 H, J 2.2 Hz), 7.13-7.34 (in, 10 H, Ph).

Example 13. Preparation of 3-Benzyl-5,7-bis[(1 yl)methoxy]-1 -(4-fluorobenzyl)-4-inethyl-2(1 H)-quinolinone a) 1 -Benzyl-5,7-dimethoxy-3-(4-fluorobenzyl)-4-methyl-2(l1H)quinolinone The product from example 8b (2 potassium -tert-butoxide (0.87 g) and 4-fluorobenzylchloride (1.12 g) were heated in DMVSO (20 ml) at 6000C for three hours as in example 8d. Yield 1.28 g.

WO 99/15523 PCT/F198/00755 1 HNMR (400 Mhz, DMSO-d6): 2.53 3H), 3.73 3H), 3.83 (s, 3H), 5.55 2H), 6.43 2H), 7.12-7.2 (in, 5 7.26-7.28 (in, 4H).

b) 3-Benzyl-5,7-dihydroxy-l1-(4-fluorobenzyl)-4-methyl-2(l1H)quinolinone

OH

HO N 0

F

The product from previous example (1.25 g) were heated in pyridine hydrochloride (12.5 g) at about 225 00 for 9 minutes. Yield 1 g.

1 HNMR (300 Mhz, DMSO-d6): 2.56 3H), 4.07 2H), 5.4 2H), 6.13 1 H, J 2.1 Hz), 6.20 1 H, J 2.1 Hz), 7.12-7.28 (mn, 9H), 9.88 (s, 1 10.22 I1H).

c) 3-Benzyl-5,7-Bis(cyanomethoxy)-1 -(4-fluorobenzyl) -4-methyl- 2(1 H)-quinolinone The product from the previous example (1 CICH2CN (0.43 g) and K2003 (1.42 g) were heated in DMF (8 ml) at 120 00 for one hour. Yield 0.94 g.

1 H-NMR (300 Mhz, DMSO-d6): 2.55 3H), 4.14 2H), 5.25 (s, 2H), 5.28 2H), 5.57 2H), 6.74 2H, ArH), 7.1 -7.3 (in, 9H).

WO 99/15523 PCT/F198/00755 36 d) 3-Benzyl-5,7-bis[( 1 H-tetrazol-5-yl)methoxy]- 1 luorobenzyl)-4methyl-2(1 H)-quinolinone

,N

HN~ 0 0 N 0 HN

F

The product from the previous example (0.5 sodium azide (0.14 g) and ammonium chloride 12 g) were heated in DMF (5 ml) at 120 00 for min. The product was triturated with acetonitrile. Yield 0.28 g. Melting point: 126-132 00.

1 HNMR (300 Mhz, DMSO-d6): 2.48 3H), 4.11 5.51 2H), 5.55 2 5.58 2 6.67 1 H, J 2.1 Hz), 6.78 1 H, J 2.1 Hz).

Example 14. Preparation of 5,7-Bis[(1 H-tetrazol-5-yl)methoxy]-3-(4chlorobenzyl)-4-methyl-2 H-i -benzopyran-2-one a) 3-(4-Chlorobenzyl)-5,7-dihydroxy-4-methyl-2H-1 -benzopyran-2-one

OH

HO 0O 0

CI

A solution of phloroglucinol (1.57 g) and ethyl 2-(4-chlorobenzyl)acetoacetate (3.18 g) in ethanol (25 ml) was treated with dry HOI at 0 00 for hours and the solution was kept at that temperature overnight. Solvent was evaporated and the precipitate triturated with water. Yield 3.87 g (98 Melting point 270-278 00.

1 H.NMR (DMSO-d6, 300 MHz): 2.52 3H, OH- 3 3.87 2H, CH2), 6.17 1 H, J 2.4 Hz), 6.28 1 H, J 2.4 Hz), 7.1 8-7.34 (in, 4H, Ph), 10.21 1 H, OH), 10.48 1 H, OH).

b) 5,7-Bis(cyanomethoxy)-3-(4-chlorobenzyl)-4-methyl-21-1 benzopyran-2-one WO 99/15523 PCT/F198/00755 37 The product from the previous example (1 .00 chioroacetonitrile (0.50 g) and potassium carbonate (2.18 g) were heated in DMF (5 ml) at 100 00 for 30 minutes. The product was isolated as described in example l b.

Yield 0.90 g (72 1 HNMR (DMSO-d6, 300 MHz): 2.52 3H, CH3), 3.95 2H, 0H2), 5.308 2H, OCH2CN), 5.312 2H, OCH2CN), 6.81 1lH, J 2.5 Hz), 6.94 1 H, J 2.5 Hz), 7.22-7.33 (in, 4H, Ph).

c) 5,7-Bis[( 1 H-tetrazol-5-yl)methoxy]-3-(4-chlorobenzyl)-4-methyl-2H- 1 -benzopyran-2-one 0

SN

NN

0:i1I~ 0 0 C The product from the previous example (0.40 sodium azide (0.14 g) and ammonium chloride 11 g) were heated in DMF (2 ml) at 100 00 for 2 hours. The product was isolated as in example 1c. Yield 0.40 g (82 1 HNMR (DMSO-d6, 300 MHz): 2.46 3H, 0H3), 3.92 2H, CH2), 5.602 2H, OCH2Tet), 5.609 2H, OCH2Tet), 6.83 1 H, J 2.5 Hz), 6.85 1 H, J 2.5 Hz), 7.20-7.33 (in, 4H, Ph).

Example 15. Preparation of 5,7-Bis[(1 H-tetrazol-5-yI)methoxy]-3-(4nitrobenzyl)-4-methyl-2H-1 -benzopyran-2-one a) 5,7-Dihydroxy-4-methyl-3-(4-nitrobenzyl)-2 H-i -benzopyran-2-one WO 99/15523 PCT/FI98/00755 38 HO 0 0

NO

I-

0 A solution of phloroglucinol (0.48 g) and ethyl 2-(4-nitrobenzyl)acetoacetate (1.00 g) in ethanol (150 ml) was treated with dry HCI at 0 °C for hours and the solution was kept at that temperature overnight. Solvent was evaporated and the precipitate triturated with water. Yield 0.63 g (51 Melting point 280-285 oC.

1 H-NMR (DMSO-d6, 300 MHz): 2.53 3H, CH3), 4.03 2H, CH2), 6.19 1H, J 2.4 Hz), 6.29 1H, J 2.4 Hz), 7.40-7.51 and 8.11-8.17 4 H, Ph), 10.25 1 H, OH), 10.52 1 H, OH).

b) 5,7-Bis(cyanomethoxy)-3-(4-nitrobenzyl)-4-methyl-2H-1benzopyran-2-one

N

N O

I--

0 The product from the previous example (0.57 chloroacetonitrile (0.27 g) and potassium carbonate (1.20 g) were heated in DMF (2 ml) at 100 °C for 50 minutes. The product was isolated as described in example lb.

Yield 0.47 g (67 Melting point 178-185 oC.

1 H-NMR (DMSO-d6, 400 MHz): 2.53 3H, CH3), 4.11 2H, CH2), 5.319 2H, OCH2CN), 5.323 2H, OCH2CN), 6.83 1H, J 2.4 Hz), 6.96 1H, J 2.4 Hz), 7.48-7.53 and 8.12-8.16 4H, Ph).

WO 99/15523 PCT/F198/00755 39 c) 5,7-Bis( H-tetrazol-5-yI)methoxyl-3-(4-nitrobenzyl)-4-methyl-2 H-i benzopyran-2-one

/N-

N/II 0

NI-

H

f 0 0 0 N N I N 0

H

The product from the previous example (0.38 sodium azide (0.12 g) and ammonium chloride 11 g) were heated in DMVF (3 ml) at 100 00C for 2 hours. The product was isolated as described in example 1c. Yield 0.25 g (54 ).Melting point 240-244 00.

1 H.NMR (DMSO-d6, 400 MHz): 2.47 3H, CH3), 4.08 2H, 0H2), 5.611 2H, OCH2Tet), 5.623 2H, OCH2Tet), 6.85 1 H, J 2.4 Hz), 6.87 1 H, J 2.4 Hz), 7.46-7.50 and 8.12-8.16 (in, 4H, Ph).

Example 16. Preparation of 5,7-Bis[(1 H-tetrazol-5-yl)methoxy]-3cyclopentyl-4-methyl-2H-1 -benzopyran-2-one a) 3-Cyclopentyl-5 ,7-dihydroxy-4-methyl-2 H-i -benzopyran-2-one

HO

HO 0 0.

A solution of phloroglucinol (2.00 g) and ethyl 2-cyclopentylacetoacetate (3.14 g) in ethanol (40 ml) was treated with dry HOI at 0 00 for hours and the solution kept at that temperature overnight. Solvent was evaporated and the precipitate purified with flash chromatography eluting with toluene-EtOAc-AcOH Yield 1 .22 g (29%) 1 HNMR (DMSO-d6, 300 MHz): 1.50-1.88 (in, 8H, 2.57 (s, 3H, 0H3), 3.25 (in, 1 H, OH), 6.11 1 H, J 2.4 Hz), 6.25 1 H, J 2.4 Hz), 10.25 2H, OH).

b) 5,7-Bis(cyanoinethoxy)-3-cyclopentyl-4-methyl-2H-1 -benzopyran-2one WO 99/15523 PCT/F198/00755 <0 The product from the previous example (0.50 chioroacetonitrile (0.31 g) and potassium carbonate (0.61 g) were heated in DMF (2 ml) at 00 for 40 minutes. The product was isolated as described in example l b.

Yield 0.56 g (86%) 1 HNMR (DMSO-d6, 300 MHz): 1.55-1.90 (in, 8H, 2.56 (s, 3H, CH3), 3.37 (in, 1 H, CH), 5.29 2H, OCH2CN), 5.31 2H, OCH2CN), 6.75 1 H, J 2.5 Hz), 6.88 1 H, J 2.5 Hz) c) 5,7-Bis[( 1 H-tetrazol-5-yl)methoxy]-3-cyclopentyl-4-methyl-2 H-i benzopyran-2-one 0

N

H

N -N N

N

H

The product from the previous example (0.30 sodium azide 13 g) and ammonium chloride 11 g) were heated in DMF (1 ml) at 100 OC for hours. The product was isolated as described in example 1 c. Yield 0.30 g (80 Melting point 248-252 00.

1 HNMR (DMSO-d6, 400 MHz): 1.53-1 .89 (mn, 8H, 2.51 (s, 3H, CH3), 3.34 (in, 1 H, OH), 5.59 2H, OOH2Tet), 5.61 2H, OCH2Tet), 6.80 2H).

Example 17. Preparation of 5,7-Bis[(1 H-tetrazol-5-yl)methoxy]-4methyl-3-( 1 -naphtylm ethyl) -2 H- 1 -benzopyran-2-one a) 5 ,7-d ihyd roxy-4-m ethyl (l-naphtylmethyl)-2H-l -benzopyran-2one WO 99/15523 PCTIFI98/00755 41

HO

HO O O A solution of phloroglucinol (0.47 g) and ethyl 2-(1-naphtylmethyl)acetoacetate (1.00 g) in ethanol (20 ml) was treated with dry HCI at 0 °C for 3 hours and the solution kept at that temperature overnight. Solvent was evaporated and the precipitate triturated with water and recrystallized from isopropanol-water Yield 0,96 g (78 Melting point 275-280 oC.

1 H-NMR (DMSO-d6, 400 MHz): 2.45 3H, CH3), 4.32 2H, CH2), 6.23 1 H, J 2.5 Hz), 6.32 1 H, J 2.5 Hz), 6.97-8.25 7H, Naph), 10.26 1H, OH), 10.53 1H, OH).

b) 5,7-Bis(cyanomethoxy)-4-methyl-3-(1-naphtylmethyl)-2H-1benzopyran-2-one 0 0 The product from the previous example (0.80 chloroacetonitrile (0.36 g) and potassium carbonate (0.66 g) were heated in DMF (4 ml) at 100 °C for 1 hour. The product was isolated as in example lb. Yield 0.30 g (30 1 H-NMR (DMSO-d6, 300 MHz): 2.45 3H, CH3), 4.40 2H, CH2), 5.34 2H, OCH2CN), 5.36 2H, OCH2CN), 6.86 1H, J 2.5 Hz), 7.010 1H, J 2.5 Hz), 7.016-8.27 7H, Naph).

c) 5,7-Bis[(1 H-tetrazol-5-yl)methoxy]-4-methyl-3-(1 -naphtylmethyl)-2H- 1 -benzopyran-2-one

N

N-NH

N-H 0 0 0

N-NH

WO 99/15523 PCT/FI98/00755 42 The product from the previous example (0.25 sodium azide (0.080 g) and ammonium chloride (0.072 g) were heated in DMF (2 ml) at 100 °C for 2.5 hours. The product was isolated as described in example c. Yield 0.11 g (36 Melting point 164-174 °C.

1 H-NMR (DMSO-d6, 300 MHz): 2.40 3H, CH3), 4.37 2H, CH2), 5.63 2H, OCH2Tet), 5.65 2H, OCH2Tet), 6.87 1H, J 2.5 Hz), 6.92 1 H, J 2.5 Hz), 6.98-8.26 7H, Naph).

Example 18. Preparation of 1-Benzyl-5,7-bis-[(1H-tetrazol-5-yl)methoxy]-4-methyl-2(1H)-quinolinone a) 5,7-Dimethoxy-4-methyl-2(1 H)-quinolinone 0 N 0

H

tert-Butyl acetoacetate (1.58 g) was heated to 120 °C and dimethoxyaniline (1.53 g) dissolved in xylene (4 ml) was added. The mixture was heated at 120-130 °C for 20 minutes and then cooled to room temperature. Methanesulfonic acid (2 ml) was added and the mixture was stirred at ambient temperature for 10 minutes. Water (40 ml) was added and the precipitate filtered and dried. Yield 1.31 g (60 1 H-NMR (DMSO-d6, 300 MHz): 2.50 3H, CH3), 3.79 3H, OCH3), 3.83 3H, OCH3), 6.03 1H, CH=C), 6.31 1 H, J 2.3 Hz), 6.45 1H, J 2.3 Hz), 11.4 1H, NH).

b) 1 -Benzyl-5,7-dimethoxy-4-methyl-2(1 H)-quinolinone WO 99/15523 PCT/FI98/00755 43 The product from the previous example (1.20 g) was suspended to DMSO (15 ml) and t-BuOK (0.68 g) and benzylbromide (1.03 g) were added.

Reaction mixture was stirred at ambient temperature overnight. Water was added and the product extracted to EtOAc. EtOAc was dried and evaporated to dryness. The product was recrystallized from toluene. Yield 0.80 g (47 1 H-NMR (DMSO-d6, 300 MHz): 2.55 3H, J 1.1 Hz, CH3), 3.71 3H, OCH3), 3.84 3H, OCH3), 5.48 2H, NCH2), 6.29 1H, J 1.1 Hz, CH=C), 6.4 2H), 7.18-7.33 5H, Ph).

c) 1-Benzyl-5,7-dihydroxy-4-methyl-2(1H)-quinolinone

OH

H N 0 The product from the previous example (0.69 g) was dissolved to

CH

2 CI2 (14 ml) and the reaction mixture cooled to -20 BBr 3 (2.4 g) in CH2CI2 (1M solution) was added and the mixture was allowed to warm to ambient temperature during the night. The precipitate was filtered, washed with CH 2 C2 and dissolved to EtOAc. EtOAc was washed with dilute HCI, dryed and evapotated to dryness. Yield 0.34 g (54 1 H-NMR (DMSO-d6, 300 MHz): 2.56 3H, J 1.0 Hz, CH3), 5.33 2H, NCH2), 6.11 1H, J 2.1 Hz), 6.13 1H, J 1.0 Hz, CH=C), 6.17 1 H, J 2.1 Hz), 7.12-7.34 5H, Ph), 9.90 1H, OH), 10.22 1H, OH).

d) 1-Benzyl-5,7-bis(cyanomethoxy)-4-methyl-2(1 H)-quinolinone

N

WO 99/15523 PCT/FI98/00755 44 The product from the previous example (0.34 chloroacetonitrile (0.13 g) and potassium carbonate (0.34 g) were heated in DMF (2 ml) at 100 oC for 1.5 hours. Water was added and the precipitate filtered and dried.

The product was recrystallized from isopropanol. Yield 0.20 g (46 1 H-NMR (DMSO-d6, 400 MHz): 2.57 3H, CH3), 5.22 2H, OCH2CN), 5.30 2H, OCH2CN), 5.50 2H, NCH2), 6.42 1H, CH=C), 6.70 1 H, J 2.1 Hz), 6.73 1 H, J 2.1 Hz), 7.21-7.32 5H, Ph).

e) 1 -Benzyl-5,7-bis-[(1 H-tetrazol-5-yl)methoxy]-4-methyl-2(1 H)quinolinone N I

I

The product from the previous example (0.20 sodium azide (0.072 g) and ammonium chloride (0.060 g) were heated in DMF (2 ml) at 100 °C for 3 hours. The product was isolated as described in example 1c. Yield 0.21 g (85 Melting point 246-249 oC.

1 H-NMR (DMSO-d6, 400 MHz): 2.50 3H, CH3), 5.48 4H, OCH2Tet, NCH2), 5.60 2H, OCH2Tet), 6.34 1 H, CH=C), 6.64 1 H, J 1.9 Hz), 6.77 1H, J 1.9 Hz), 7.18-7.32 5H, Ph).

Example 19. Preparation of 1-Benzyl-5,7-bis[1 yl)methoxy]-3-(2-fluorobenzyl)-4-methyl-2(1 H)-quinolinone a) 5,7-Dimethoxy-3-(2-fluorobenzyl)-4-methyl-2(1 H)-quinolinone

F

H O

H

WO 99/15523 PCT/FI98/00755 Ethyl 2-(2-fluorobenzyl)acetoacetate (2.5 g) in xylene (1 ml) was heated to 150 °C and 3,5- dimethoxyaniline (1.46 g) in xylene (4 ml) was added in small portions during 30 minutes. The reaction mixture was heated at 160 °C for 3 hours and then cooled to room temperature. Methanesulfonic acid (1.7 ml) was added and the mixture was stirred at ambient temperature for 30 minutes. Water was added and the precipitate filtered and dried. The product was triturated with warm ethanol. Yield 0.64 g (21 1 H-NMR (DMSO-d6, 300 MHz): 2.45 3H), 3.79 3H), 3.82 (s, 3H), 3.97 2H), 6.33 1 H, J 2.4 Hz), 6.48 1 H, J 2.4 Hz), 6.90-7.25 4H), 11.61 1H).

b) 1-Benzyl-5,7-dimethoxy-3-(2-fluorobenzyl)-4-methyl-2(1

H)-

quinolinone O F 0 The product from the previous example (0.62 g) was treated with t-BuOK (0.23 g) and benzylbromide (0.36 g) in DMSO (12 ml) at 60 °C for 2.5 hours.

The product was isolated as described in example 18b. Yield 0.39 g (49 1 H-NMR (DMSO-d 6 400 MHz): 2.51 3H), 3.72 3H), 3.84 (s, 3H), 4.11 2H), 5.55 2H), 6.433 1 H, J 2.1 Hz), 6.443 1 H, J 2.1 Hz), 6.97-7.33 9H).

c) 1-Benzyl-5,7-dihydroxy-3-(2-fluorobenzyl)-4-methyl-2(1H)quinolinone WO 99/15523 PCT/FI98/00755 46 The product from the previous example (0.34 g) was treated with BBr3 (8.48 g) in CH2C12 (7 ml) as described in example 18c. Yield 0.30 g (82 1 H-NMR (DMSO-d6, 400 MHz): 2.55 3H), 4.06 2H), 5.40 2H), 6.13 1H, J 2.1 Hz), 6.22 1H, J 2.1 Hz), 6.97-7.33 9H), 10.3 2H).

d) 1 -Benzyl-5,7-bis(cyanomethoxy)-3-(2-fluorobenzyl)-4-methyl-2(1H)quinolinone N- 0

N

The product from the previous example (0.21 chloroacetonitrile (0.086 g) and potassium carbonate (0.37 g) were heated in DMF (2 ml) at 100 °C for 2 hours. The product was isolated as described in example lb.

Yield 0.18 g (71 1 H-NMR (DMSO-d6, 400 MHz): 2.53 3H), 4.13 2H), 5.23 (s, 2H), 5.29 2H), 5.57 2H), 6.746 1H, J 2.3 Hz), 6.756 1 H, J 2.3 Hz), 7.00-7.32 9H).

e) 1 -Benzyl-5,7-bis[1 H-tetrazol-5-yl)methoxy]-3-(2-fluorobenzyl)-4methyl-2(1 H)-quinolinone N 0 F

N-NH

0 N 0

N-NH

The product from the previous example (0.17 sodium azide (0.051 g) and ammonium chloride (0.042 g) were heated in DMF at 100 °C for 3 hours. The product was isolated as described in example 1c. Yield 0.17 g Melting point 135-140 oC.

WO 99/15523 PCT/FI98/00755 47 1 H-NMR (DMSO-d6, 400 MHz): 2.46 3H), 4.10 2H), 5.48 (s, 2H), 5.51 2H), 5.59 2H), 6.68 1 H, J 2.2 Hz), 6.79 1 H, J 2.2 Hz), 6.99-7.32 9H).

Example 20. Preparation of 1-Benzyl-5,7-bis[1 methoxy]-4-methyl-3-(2-phenylethyl)-2(1 H)-quinolinone a) 5,7-Dimethoxy-4-methyl-3-(2-phenylethyl)-2(1 H)-quinolinone 0 N 0

H

Ethyl 2-(2-phenylethyl)acetoacetate (2.70 g) in xylene (5 ml) was treated with 3,5-dimethoxyaniline (1.60 g) at 150 oC as described in example 19a. Methanesulfonic acid (4.0 ml) was added at room temperature and the mixture heated at 80 oC for 1 hour. The product was isolated as described in example 19a. Yield 1.38 g (41 1 H-NMR (DMSO-d6, 400 MHz): 2.45 3H), 2.64-2.68 2H), 2.82- 2.86 2H), 3.78 3H), 3.81 3H), 6.30 1 H, J 2.3 Hz), 6.45 1 H, J= 2.3 Hz), 7.18-7.30 5H), 11.45 1H).

b) 1 -Benzyl-5,7-dimethoxy-4-methyl-3-(2-phenylethyl)-2(1

H)-

quinolinone The product from the previous example (0.61 t-BuOK (0.24 g) and benzylbromide (0.36 g) were heated in DMSO (12 ml) at 60 °C for 2 hours.

The product was isolated as described in example 18b. Yield 0.31 g (40 WO 99/15523 PCT/F198/00755 48 1 H-NMR (DMSO-d6, 400 MHz): 2.51 3H), 2.73-2.77 (in, 2H), 2.96- 3.00 (in, 2H), 3.70 3.83 3H), 5.55 2H), 6.40 2H), 7.17-7.33

H).

c) 1 -Benzyl-5,7-dihydroxy-4-methyl-3-(2-phenylethyl)-2(1

H)-

quinolinone The product from the previous example (0.31 g) was treated with BBr3 (0.75 g) in CH2012 (5 ml) as in example 18c. Yield 0.26 g (89 1 H-NMR (DMSO-d6, 300 MHz): 2.56 3H), 2.69-2.75 (in, 2H), 2.90- 2.95 (in, 2H), 5.39 2H), 6.08 1 H, J 2.0 Hz), 6.19 1 H, J 2.0 Hz), 7.11-7.33 (mn, 10H), 10.2 2H).

d) 1 -Benzyl-5,7-bis(cyanoinethoxy)-4-methyl-3-(2-phenylethyl)-2( 1 H)quinolinone

I-Z

The product from the previous example (0.22 chioroacetonitrile (0.09 1 g) and potassium carbonate (0.39 g) were heated at 100 00 for 2 hours. The product was isolated as in example 1 b. Yield 0.20 g (76 WO 99/15523 PCT/F198/00755 49 1 H-NMR (DMSO-d6, 400 MHz): 2.50 3H), 2.73-2.77 (in, 2H), 2.98- 3.02 (in, 2H), 5.21 2H), 5.29 2H), 5.56 6.70 1 H, J 2.1 Hz), 6.72 1 H, J 2.1 Hz), 7.18-7.33 (in, 1 OH).

e) 1 -Benzyl-5,7-bis[1 H-tetrazol-5-yl)methoxyl-4-methyl-3-(2phenylethyl)- 2(1 H)-quinolinone The product from the previous example (0.19 sodium azide (0.057 g) and ammonium chloride (0.047 g) were heated in DMIF at 100 00 for 3 hours. The product was isolated as described in example 1c. Yield 0.18 g (78 ).Melting point 215-218 00.

1 H-NMR (DMSO-d6, 400 MHz): 2.46 2.70-2.74 (in, 2H), 2.95- 2.99 (in, 2 5.47 2 5.54 2 5.57 2 6.64 1 H, J 2. 0 Hz), 6.77 1 H, J 2.0 Hz), 7.16-7.33 (in, 1 OH).

Example 21. Preparation of 5,7-Bis(aininocarbonylmethoxy)-1 ,3dibenzyl-4-methyl-2(1 H)-quinolinone.

H 2Nyr 0 H 2

N

The mixture of 5,7-dihydroxy- 1,3-dibenzyl-4-methyl-2( 1 H)-quinolinone potassium carbonate (0.9 g) and 2-chloroacetamide 0.25 g) in DMIF ml) were reacted at 100 00 for two hours. The reaction mixture was WO 99/15523 PCT/FI98/00755 treated with ice water and filtered. The product was triturated with hot ethanol. Yield: 0.32 g. Melting point 252-253oC.

1 H-NMR (400 MHz, DMSO-d6): 2.63 3H, CH3), 4.13 2H, PhCH2), 4.37 2H, OCH2), 4.55 2H, OCH2), 5.54 2H, NCH2Ph), 6.40 1H, J =2 Hz, ArH), 6.53 1H, J 2 Hz, ArH), 7.13-7.33 10 H, Ph), 7.44 2H, J 65 Hz, CONH2), 7.47 2H, J 68 Hz, CONH2).

Example 22. Preparation of 5,7-Bis(ethoxycarbonylmethoxy)-1,3dibenzyl-4-methyl-2(1 H)-quinolinone.

O*NO O 0N 0 The mixture of 5,7-dihydroxy-1,3-dibenzyl-4-methyl-2(1 H)-quinolinone (1 ethyl 2-bromoacetate (0.63 ml) and potassium carbonate (1.49 g) in DMF (5 ml) was heated under nitrogen at 110 °C for three hours, poured into ice water and filtered. The resulting solid material was triturated with ether and filtered again. Yield: 1.03 g, melting point 113-116 oC.

1 H-NMR (400 MHz, DMSO-d6): 1.15 3H, CH3CH2, J 7.1 Hz), 1.20 3H, CH3CH2, J 7.1 Hz), 2.63 3H, CH3), 4.03 2H, CH2CH3, J 7.1 Hz), 4.13 2H, CH2Ph), 4.17 2H, CH2CH3, J 7.1 Hz), 4.78 (s, 2H, OCH2), 4.90 2H, OCH2), 6.41 1 H, J 2.2 Hz), 6.44 1 H, J 2.2 Hz), 7.13-7.33 10 H, Ph).

WO 99/15523 PCT/F198/00755 51 Example 23. Preparation of 5 ,7-Bis(hydroxyaminocarbonyimethoxy)- 1, 3-d ibenzyl -4-m ethyl -2(1 H)-quinolinone

OH

0 O- N 0 HN,

OHN

The product from the previous example (0.3 hydroxylamine hydrochloride (0.32 g) and 5 N NaOH (1.05 ml) were reacted in ethanol (8 ml) at 50 00 for six hours. The reaction mixture was treated with water and made basic (pH 10) and filtered. The filtrate was acidified to pH 2 and filtered. Yield: 0.2 g, melting point 121-1 2700.

1 H.NMR (400 MHz, DMSO-d6): the tautomeric forms of hydroxamic acid are seen in 00H2-signals: 2.63 CH-3), 4.13 2H, CH2Ph), 4.41 2H, 00H2), 4.54 2H,00H2), 4.64 2H-, HON=C(OH)CH2O), 4.65 2H, HON=C(OH)0H20), 4.77 2H, HON=C(OH)CH2O), 4.78 2H, HON=C(OH)0H20), 5.54 2H, NCH2Ph), 6.38-6.54 (in, 2H, ArH), 7.14- 7.34 (in, 10 H, Ph), 9.05 2H, NOH), 10.84 1 H, HONHCO), 10.88 (s, 1 H, HONHCO).

Example 24. Preparation of 5,7-Bis -[1-(6-hydroxypyridazinyl)]oxy-1 ,3dibenzyl-4-methyl-2( 1 H)-quinolinone a) 5,7-Bis (6-chlIoropy rid azinyl)]oxy-1, ,3d i benzyl-4-methyl-2 (1 H)quinolinone Ck,

N

WO 99/15523 PCT/FI98/00755 52 A mixture of 1,3-dibenzyl-5,7-dihydroxy-4-methyl-2(1 H)-quinolinone 3,6-dichloropyridazine (0.83 g) and potassium carbonate (0.75 g) in DMF (12,5 ml) was stirred at 80 °C for 4 hours. The reaction mixture was treated with water at pH 8 and filtered. The solids were recrystallized from ethanol-DMF Yield 0.5 g. Melting point 208-218°C.

1 H-NMR (DMSO-d6, 300 MHz): 2.43 3H,CH3), 4.16 2H, CH2Ph), 5.58 2H, NCH2Ph), 7.09-7.33 12H, ArH Ph), 7.55 1H, PyridH, J 9,2 Hz), 7.70 1H, PyridH, J 9,2 Hz),7.93 1H, PyridH, J 9,2 Hz), 7.98 1H, PyridH, J 9,2 Hz).

b) 5,7-Bis -(6-hydroxypyridazinyl)]oxy-1,3-dibenzyl-4-methyl-2(1

H)-

quinolinone

H

N

N

The product from the previous example (0.2 g) and potassium acetate (0.13 g) in acetic acid (5 ml) were refluxed for 4 hours. The mixture was evaporated, treated with water at pH 10 and filtered. The filtrate was acidified to pH 6 and filtered. Yield 70 mg.

1 H-NMR (DMSO-d6, 300 MHz): 2.47 3H, CH3), 4.15 2H, CH2Ph), 5.55 2H, NCH2), 6.93- 7.34 15 H, PyridH ArH Ph), 7.47 1H, J 10 Hz), 12.25 1H, NH), 12.38 1H NH).

Claims (17)

1. 2. A compound of claim 1 wherein said compound has formula and R2 is hydrogen.
2. 3. A compound of claim 2 wherein R1 is hydrogen, C1-6 alkyl, C2.6 alkenyl, C6-10 aryl, C7-12 arylalkyl, C1- 6 hydroxyalkyl, C1-6 halogenalkyl or C1-6 alkoxy.
3. 4. A compound of claim 3 wherein Y is O or S, and X is O. A compound of claim 3 wherein Y is O or S; and X is NR11, where R11 is hydrogen, C1 6 alkyl, C6- 10 aryl, C7-12 arylalkyl, C1 6 alkoxy, C6-10 aryloxy, hydroxy, C1 6 alkanoyl or C1 6 carboxyalkyl.
4. 6. A compound of claim 5, wherein R3 is hydrogen, C1-6 alkyl, C6.10 aryl, or C7-12 arylalkyl.
5. 7. A compound of claim 6, wherein R3 is C1-6 alkyl.
6. 8. A compound of claim 7, wherein A is straight-chain or branched C 1 4 alkylene and R4 and R5 are each WO 99/1 5523 PTF9/05 PCT/F198/00755 N N-..N H ,N N 0; iH ,~N O NH H 0 HN'- 0 HCNN IN\/\ 0 N-N H or where X is NR1I 1, then R4 and R5 can also be HOOC-, Ri 2000-, H2NCO- or HOHNCO-, where R12 is 01.6 alkyl, C 6 io aryl, or C7-12 arylalkyl.
7. 9. A compound of claim 1, wherein said compound has formula (11) and R7 is hydrogen. A compound of claim 9, wherein R6 is hydrogen, C 1 6 alkyl, 06.1 0 aryl, C7-12 arylalkyl; and n is 1, 2 or 3.
8. 11. A compound of claim 10, wherein Y is 0 or S, and X is 0.
9. 12. A compound of claim 10, wherein Y is 0 or S; and X is Nl R 1, where Ri11 s hydrogen, C01 -6 alkyl, C6-.1 0 aryl, C7-1 2 arylalkyl, Ci1 -6 alkoxy, 06-1 0 aryloxy, hydroxy, C01-6 alkanoyt o r C1-6 carboxyalkyl.
10. 13. A compound of claim 11, wherein A is straight-chain or branched Ci alkylene.
11. 14. A compound of claim 13, wherein R4 and R5 are each WN NN H -N 0 NH* N HN' 0 0, H 3 CN N N N N-N H WO 99/15523 PCT/FI98/00755 56 or where X is NR11, then R4 and R5 can also be HOOC-, R1200C-, H2NCO- or HOHNCO-, where R12 is C1-6 alkyl, C6-10 aryl, or C7-12 arylalkyl.
12. 15. A pharmaceutical composition comprising a compound of claim 1 as an active ingredient together with pharmaceutically acceptable carrier.
13. 16. A method of treating heart failure comprising administering to a subject in need thereof a therapeutically effective amount of a compound of claim 1.
14. 17. A method for the treatment and prevention of stunned myocardium comprising administering to a subject in need thereof a therapeutically effective amount of a compound of claim 1.
15. 18. A compound of formula (XXV): OH R3 (XXV) HO N O R1l wherein R1 is hydrogen, alkyl, alkenyl, aryl, arylalkyl, hydroxyalkyl, halogenalkyl, alkoxy, COR10, CONR10R11, OR10, NR10COR11 or NR10R11, where R10 is hydrogen, alkyi, alkenyl, aryl, arylalkyl, hydroxyalkyl, halogenalkyl, alkoxy or hydroxy and R11 is hydrogen, alkyl, aryl, arylalkyl, alkoxy, aryloxy, hydroxy or acyl, or carboxylalkyl, R3 is hydrogen, alkyl, aryl or arylalkyl, R11 is hydrogen, alkyl, aryl, arylalkyl, alkoxy, aryloxy, hydroxy or acyl, or carboxylalkyl, and wherein each aryl residue defined above by itself or as part of another group may be substituted. -57-
16. 19. Use of a compound according to claim 1 in the preparation of a medicament for the treatment of heart failure. Use of a compound according to claim 1 in the preparation of a medicament for the treatment of stunned myocardium.
17. 21. A compound according to claim 1 or claim 18 substantially as hereinbefore described with reference to any of the examples. io DATED: 19 April 2000 PHILLIPS ORMONDE FITZPATRICK Attorneys for: ORION CORPORATION 9 9 9*9 o**