Deuterated CFTR potentiators

Abstract

This invention relates to compounds of Formula I: ##STR00001##
and pharmaceutically acceptable salts thereof, wherein each X and each R is defined within. This invention also provides compositions comprising a compound of this invention and the use of such compositions in methods of treating diseases and conditions that are beneficially treated by administering a CFTR potentiator.

Claims

1. A process for the synthesis of compound 28: ##STR00015## comprising converting compound 27: ##STR00016## into compound 28 in the presence of CD.sub.3MgI, diethyl ether, and THF.

Description

EXAMPLES

Example 1. N-(3-Hydroxy-4-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)phenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide (Compound 100)

(1) ##STR00010## ##STR00011##

Step 1. Methyl 2-((tert-butoxycarbonyl)oxy)benzoate (27)

(2) A solution of di-tert-butyldicarbonate (89.5 g, 410 mmol) in THF (50 mL) was added to a solution of methyl salicylate 26 (25.2 g, 165.6 mmol) in THF (300 mL) at 0 C. Sodium hydride (60% dispersion in mineral oil, 16.4 g, 410 mmol) was added over 5 minutes. The mixture was stirred at 0 C. for 30 minutes then at room temperature for 6 hours. The mixture was cooled to 0 C., diluted with MTBE (300 mL) and water (400 mL) was added. The layers were separated and the aqueous layer was extracted with MTBE (2150 mL). The combined organic layer was washed with saturated sodium chloride (200 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting product was purified using an Analogix chromatography system (SiO.sub.2, 0-10% ethyl acetate/heptanes) to afford 27 (39.2 g, 94% yield) as colorless oil.

Step 2. 2-(2-(Methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)phenol (28)

(3) A solution of 1.0M methyl-d.sub.3-magnesium iodide in diethyl ether (260 mL, 260.0 mmol, 99 atom % D Sigma-Aldrich) was added over 15 minutes to a solution of 27 (18.77 g, 74.4 mmol) in THF (700 mL) at 0 C. with the reaction temperature rising to 22 C. during the addition. The mixture was stirred at 0 C. for 4 hours, then at room temperature overnight. The mixture was cooled to 0 C., quenched with IM aqueous hydrochloric acid (350 mL) and extracted with MTBE (500 mL, 300 mL, 200 mL). The combined organic layers were washed with saturated sodium chloride (300 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure. The resulting product was purified using an Analogix chromatography system (SiO.sub.2, 0-10% ethyl acetate/heptanes) to afford 28 (7.19 g, 61% yield, 70% purity) as yellow oil.

Step 3. 4-Bromo-2-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)phenol (29)

(4) Bromine (2.3 mL, 44.9 mmol) was added dropwise to a solution of 28 (7.17 g, 70% purity) in dichloromethane (70 mL) at 0 C. and the mixture was stirred at 0 C. for 40 minutes. Water (80 mL) was added and the mixture was extracted with dichloromethane (70 mL). The organic layer was washed with saturated aqueous sodium thiosulfate solution (250 mL) and saturated sodium chloride (230 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting product was purified using an Analogix chromatography system (SiO.sub.2, 0-10% ethyl acetate/heptanes) to afford 29 (8.00 g, 75% yield, 80% purity) as yellow oil.

Step 4. 4-Bromo-2-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)phenyl Methyl Carbonate (30)

(5) Methylchloroformate (5.2 mL, 67.3 mmol) was added dropwise to a solution of 29 (8.00 g, 80% purity) and triethylamine (6.6 mL, 47.3 mmol) in dichloromethane (150 mL) at 0 C. and the mixture was stirred at 0 C. for 2 hours. Saturated aqueous ammonium chloride (70 mL) was added, the layers were separated and the aqueous layer was extracted with dichloromethane (250 mL). The combined organic layer was washed with saturated sodium chloride (50 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting product was purified using an Analogix chromatography system (SiO.sub.2, 0-10% ethyl acetate/heptanes) to afford 30 (7.36 g, 74% yield) as yellow oil.

Step 5. 4-Bromo-2-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)-5-nitrophenyl Methyl Carbonate (31)

(6) A solution of 30 (12.90 g, 43.5 mmol) in dichloromethane (10 mL) was added to concentrated sulfuric acid (30 mL) at 0 C. Potassium nitrate (7.05 g, 69.7 mmol) was added in portions over 15 minutes at 0 C. The mixture was stirred at room temperature for 2 hours then cooled to 0 C. A mixture of ice and water (70 mL) was added and the mixture was extracted with dichloromethane (60 mL, 60 mL, 40 mL). The combined organic layer was dried over sodium sulfate, filtered and concentrated under reduced pressure. The resulting product was purified using an Analogix chromatography system (SiO.sub.2, 0-15% ethyl acetate/heptanes) to give 31 (10.40 g, 70% yield) as a yellow solid.

Step 6. 4-Bromo-2-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)-5-nitrophenol (18c)

(7) A solution of potassium hydroxide (2.56 g, 45.7 mmol) in methanol (50 mL) was added to a solution of 31 (10.40 g, 30.5 mmol) in dichloromethane (60 mL) and methanol (100 mL). The mixture was stirred at room temperature for 30 minutes then quenched with IM aqueous hydrochloric acid to pH 3. The mixture was concentrated under reduced pressure to remove methanol, and extracted with dichloromethane (370 mL). The combined organic layer was dried over sodium sulfate, filtered and concentrated under reduced pressure. The crude product was adsorbed onto Celite and the resulting product was purified using an Analogix chromatography system (SiO.sub.2, 0-20% ethyl acetate/heptanes) to give 18c (8.00 g, 93% yield) as a bright yellow solid.

Step 7. 5-Amino-2-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)pheno(33)

(8) Compound 18 (0.75 g, 2.65 mmol) was hydrogenated at 50 psi over 10% palladium on carbon (50% wet) (0.40 g) in ethanol (25 mL) overnight. The mixture was filtered through a pad of Celite, concentrated under reduced pressure and dry-loaded onto Celite. The resulting product was purified using an Analogix chromatography system (SiO.sub.2, 0-50% ethyl acetate/heptanes, then 10% methanol/dichloromethane) to give 33 (0.46 g, quantitative yield) as an off-white solid.

Step 8. N-(3-Hydroxy-4-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)phenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide (Compound 100)

(9) A mixture of 33 (0.26 g, 1.49 mmol), commercially available 34 (0.30 g, 1.58 mmol), HATU (0.62 g, 1.64 mmol) and triethylamine (0.46 mL, 3.28 mmol) in 2-methyltetrahydrofuran (20 mL) was heated at 70 C. for 6 hours. The mixture was diluted with THF (50 mL) and water (20 mL). The layers were separated and the aqueous layer was extracted with THF (220 mL). The combined organic layer was washed with saturated sodium chloride (10 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The crude product was adsorbed onto Celite and purified using an Analogix chromatography system (SiO.sub.2, 10-90% ethyl acetate/heptanes) to give titled compound (0.47 g). The material was triturated with diethyl ether (10 mL) for 30 minutes and filtered, washed with diethyl ether (20 mL), triturated with water (20 mL) for 30 minutes, filtered, washed with water (20 mL) and dried in a vacuum oven at 50 C. for 3 days to give Compound 100 (173 mg, 33% yield). 1H NMR (DMSO-d.sub.6, 400 MHz) 12.8-12.9 (br s, 1H), 12.2-12.3 (s, 1H), 9.35-9.40 (s, 1H), 8.8-8.85 (s, 1H), 8.25-8.35 (d, 1H), 7.70-7.90 (m, 2H), 7.50-7.60 (m, 1H), 7.35-7.40 (s, 1H), 7.10-7.15 (m, 1H), 6.90-6.98 (m, 1H); 13C NMR (DMSO-d.sub.6, 100 MHz) 176.8, 162.9, 156.5, 144.5, 139.6, 137.9, 131.2, 126.95, 126.4, 125.95, 125.7, 119.6, 111.2, 110.3, 108.1, 33.7; MS (ESI) 346.2 [(M+H).sup.+].

Example 2. N-(5-hydroxy-4-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)-2-(trifluoromethyl)phenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide (Compound 120)

(10) ##STR00012##

Step 1. 1-(Benzyloxy)-4-bromo-2-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,3,3,3-d.SUB.6.)-5-nitrobenzene (19c)

(11) Cesium carbonate (11.06 g, 33.9 mmol) was added in one portion to a solution of 18c (8.00 g, 28.3 mmol) in DMF (100 mL) followed by benzyl bromide (3.6 mL, 29.7 mmol). The mixture was stirred at room temperature overnight at which time LC-MS analysis indicated complete reaction. Water (100 mL) was added and the mixture was extracted with ethyl acetate (2300 mL, 200 mL). The combined organic layer was washed with saturated sodium chloride (100 mL), dried over sodium sulfate, filtered, concentrated under reduced pressure, dry-loaded onto silica gel and the resulting product was purified using an Analogix chromatography system (SiO.sub.2, 0-10% ethyl acetate/heptanes) to afford 19c (9.84 g, 93% yield) as a light yellow solid.

Step 2. 1-(Benzyloxy)-2-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)-5-nitro-4-(trifluoromethyl)benzene (20c)

(12) A mixture of 19c (6.56 g, 17.6 mmol), potassium fluoride (2.04 g, 35.1 mmol), potassium bromide (4.18 g, 35.1 mmol), methyl chlorodifluoroacetate (16.0 mL, 151.7 mmol), and copper (I) iodide (4.00 g, 21.0 mmol) in DMF (40 mL) was heated overnight in a sealed tube at 120 C. (oil bath temperature). This reaction was repeated using 3.28 g (8.8 mmol) of 19c. The crude material was combined and water (100 mL) was added. The resulting mixture was extracted with ethyl acetate (2300 mL, 200 mL), and the combined organic layer was washed with saturated sodium chloride (100 mL), dried over sodium sulfate, filtered, concentrated under reduced pressure, dry-loaded onto silica gel and was purified using an Interchim chromatography system (SiO.sub.2, 0-10% ethyl acetate/heptanes) to give 20c (6.90 g, 72% yield, 80% purity) as a light yellow solid.

Step 3. 5-Amino-2-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)-4-(trifluoromethyl)-phenol (2cc)

(13) Compound 20c (6.90 g, 19.0 mmol, 80% purity) was hydrogenated at 25 psi over 10% palladium on carbon (50 wt. % wet) (2.07 g)) in ethanol for 5 hours. The mixture was filtered through a Celite pad, concentrated under reduced pressure, dry-loaded onto Celite and was purified using an Analogix chromatography system (SiO.sub.2, 0-20% ethyl acetate/heptanes) to give 2cc (1.46 g, 32% yield) as an off-white solid.

Step 4. N-(5-Hydroxy-4-(2-(methyl-d.SUB.3.)propan-2-yl-1,1,1,3,3,3-d.SUB.6.)-2-(trifluoromethyl)-phenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide (Compound 120)

(14) A mixture of 2cc (0.79 g, 3.27 mmol), pyridine (0.53 mL, 6.5 mmol), commercially available 34 (0.65 g, 3.44 mmol), and 50 wt. % propylphosphonic anhydride solution in ethyl acetate (9.7 mL, 16.3 mmol) in 2-methyltetrahydrofuran (75 mL) was heated overnight at 50 C. The mixture was diluted with THF (50 mL), washed with saturated sodium bicarbonate solution (350 mL), saturated sodium chloride (30 mL), dried over sodium sulfate, filtered, concentrated under reduced pressure, and dry-loaded onto silica gel. The resulting product was purified using an an Analogix chromatography system (SiO.sub.2, 0-60% ethyl acetate/heptanes) to give Compound 120 (186 mg, 14% yield) as a white solid which was lyophilized from a mixture of methanol and benzene to remove residual solvents. 1H NMR (DMSO-d.sub.6, 400 MHz) 12.85-13.0 (br s, 1H), 12.50-12.55 (s, 1H), 10.25-10.35 (s, 1H), 8.80-8.90 (s, 1H), 8.25-8.35 (d, 1H), 7.95-8.0 (s, 1H), 7.70-7.85 (m, 2H), 7.45-7.55 (s, 1H), 7.30-7.35 (m, 1H); 13C NMR (DMSO-d.sub.6, 100 MHz) 176.8, 163.7, 159.8, 144.98, 139.6, 133.5, 131.5, 126.4, 126.1, 125.8, 119.7, 112.4, 110.7, 33.7; 19F (DMSO-d.sub.6, 376 MHz) 57.99; MS (ESI) 414.2 [(M+H).sup.+].

(15) Compound A and Compound B, for use in Example 3 and 4 respectively, are prepared as described in U.S. Pat. No. 8,354,427 and Hadida, S. et al., Journal of Medicinal Chemistry, 57(23), 9776-9795; 2014.

Compound A. N-(4-(tert-butyl)-3-hydroxyphenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide

(16) ##STR00013##

Compound B. N-(4-(tert-butyl)-5-hydroxy-2-(trifluoromethyl)phenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide

(17) ##STR00014##

Example 3. Evaluation of Metabolic Stability of Compound 100 in Human CYP3A4 Supersomes

(18) SUPERSOMES Assay.

(19) Stock solutions (7.5 mM) of Compound 100 and Compound A were prepared in DMSO. The 7.5 mM stock solutions were diluted to 12.5 M in acetonitrile (ACN). The human CYP3A4 Supersomes (2000 pmol/mL, purchased from BD Biosciences) were diluted to 62.5 pmol/mL in 0.1 M potassium phosphate buffer, pH 7.4, containing 3 mM MgCl.sub.2. The diluted supersomes were added to wells of a 96-well polypropylene plate in triplicate. A 10 L aliquot of the 12.5 M test compound was added to the supersomes and the mixture was pre-warmed for 10 minutes. Reactions were initiated by addition of pre-warmed NADPH solution. The final reaction volume was 0.5 mL and contained 50 pmol/mL CYP3A4 Supersomes, 0.25 M test compound, and 2 mM NADPH in 0.1 M potassium phosphate buffer, pH 7.4, and 3 mM MgCl.sub.2. The reaction mixtures were incubated at 37 C., and 50 L aliquots were removed at 0, 3, 7, 10, 20 and 30 minutes and added to 96-well plates which contained 125 L of ice-cold ACN with internal standard to stop the reactions. The plates were stored at 4 C. for 20 minutes after which 75 L of water was added to the wells of the plate before centrifugation to pellet precipitated proteins. Supernatants were transferred to another 96-well plate and analyzed for amounts of parent remaining by LC-MS/MS using an AB Sciex QTrap 5500 mass spectrometer.

(20) Data Analysis:

(21) The in vitro half-lives (t.sub.1/2 values) for test compounds were calculated from the slopes of the linear regression of LN (% parent remaining) vs incubation time relationship:
in vitro t.sub.1/2=0.693/k, where k=[slope of linear regression of LN (% parent remaining) vs incubation time].

(22) The results of this experiment are shown in Table 2, below. Compound 100 was shown to be more stable in Human CYP3A4 Supersomes relative to Compound A, with a calculated average half-life of 26.5 minutes for Compound 100 and 15.1 minutes for Compound A. This represents an average 75% increase in t.sub.1/2 for Compound 100 and an apparent intrinsic clearance ratio of 0.57 L/min*pmol.

(23) TABLE-US-00002 TABLE 2 Metabolic Stability of Compound 100 versus Compound A in Human CYP3A4 Supersomes **Cl.sub.int, app rCYP (L/min*pmol) Compound *t.sub.1/2 (minutes) Ratio (100/A) Compound A 15.1 0.69 0.46 0.02 Compound 100 26.5 1.4 0.26 0.00 *t.sub.1/2 was calculated based on 10 min data; ** CL int = (0.693/In Vitro T) (mL IncubationVolume/pmol P450)

Example 4. Evaluation of Metabolic Stability of Compound 120 in Human CYP3A4. Supersomes

(24) SUPERSOMES Assay.

(25) Stock solutions (7.5 mM) of Compound 120 and Compound B were prepared in DMSO. The 7.5 mM stock solutions were diluted to 12.5 M in acetonitrile (ACN). The human CYP3A4 Supersomes (2000 pmol/mL, purchased from BD Biosciences) were diluted to 62.5 pmol/mL in 0.1 M potassium phosphate buffer, pH 7.4, containing 3 mM MgCl.sub.2. The diluted supersomes were added to wells of a 96-well polypropylene plate in triplicate. A 10 L aliquot of the 12.5 M test compound was added to the supersomes and the mixture was pre-warmed for 10 minutes. Reactions were initiated by addition of pre-warmed NADPH solution. The final reaction volume was 0.5 mL and contained 50 pmol/mL CYP3A4 Supersomes, 0.25 M test compound, and 2 mM NADPH in 0.1 M potassium phosphate buffer, pH 7.4, and 3 mM MgCl.sub.2. The reaction mixtures were incubated at 37 C., and 50 L aliquots were removed at 0, 3, 7, 10, 20 and 30 minutes and added to 96-well plates which contained 125 L of ice-cold ACN with internal standard to stop the reactions. The plates were stored at 4 C. for 20 minutes after which 75 L of water was added to the wells of the plate before centrifugation to pellet precipitated proteins. Supernatants were transferred to another 96-well plate and analyzed for amounts of parent remaining by LC-MS/MS using an AB Sciex QTrap 5500 mass spectrometer.

(26) Data Analysis:

(27) The in vitro half-lives (t.sub.1/2 values) for test compounds were calculated from the slopes of the linear regression of LN (% parent remaining) vs incubation time relationship:
in vitro t.sub.1/2=0.693/k, where k=[slope of linear regression of LN (% parent remaining) vs incubation time].

(28) The results of this experiment are shown in Table 3. Compound 120 was shown to be more stable in Human CYP3A4 Supersomes relative to Compound B, with a calculated average half-life of 20.6 minutes for Compound 120 and 14.5 minutes for Compound B. This represents an average 42% increase in t.sub.1/2 for compound 120 and an apparent intrinsic clearance ratio of 0.71 mL/min*pmol.

(29) TABLE-US-00003 TABLE 3 Metabolic Stability of Compound 120 versus Compound B in Human CYP3A4 Supersomes **Cl.sub.int, app rCYP *t.sub.1/2 (minutes) (L/min*pmol) Compound (% ) Ratio (120/B) Compound B 14.5 1.8 0.48 0.06 Compound 120 20.6 2.7 0.34 0.00 *t.sub.1/2 was calculated based on 10 min data; **CL int = (0.693 / In Vitro T) (mL Incubation Volume/pmol P450)

(30) Data analysis was performed using Microsoft Excel Software.

(31) Without further description, it is believed that one of ordinary skill in the art can, using the preceding description and the illustrative examples, make and utilize the compounds of the present invention and practice the claimed methods. It should be understood that the foregoing discussion and examples merely present a detailed description of certain preferred embodiments. It will be apparent to those of ordinary skill in the art that various modifications and equivalents can be made without departing from the spirit and scope of the invention.