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To a solution of 4-bromobenzene-l,2-diamine (2.5 g, 13.37 mmol) in DCM (30 mL) was added (S)-2-((tert-butoxycarbonyl)amino)-3,3-dimethylbutanoic acid (3.09 g, 13.37 mmol), DIPEA (2.334 mL, 13.37 mmol) and HATU (5.08 g, 13.37 mmol). The reaction mixture was stirred at room temperature for 18 h. The reaction mixture was diluted with water and extracted with DCM. The organic phase was washed with brine, dried over Na2S04, filtered and concentrated. The crude material was purified by ISCO using 40 g Redisep silica column, CHCl3/MeOH as eluant to obtain (S)-tert-butyl ( 1 -((2-amino-4-bromophenyl)amino)-3 ,3 -dimethyl- 1 -oxobutan-2-yl) carbamate (1.82 g) as yellow solid. LC (Condition 1): Rt = 2.13 min. LC/MS: Anal. Calcd. for [M+H20]+ Ci7H27BrN204 : 402.12; found 402.2. 1H NMR (DMSO-d6, δ = 2.50 ppm, 400 MHz): δ 9.35 – 9.21 (m, 1 H), 7.07 (d, J= 8.5 Hz, 1 H), 6.91 (d, J= 2.0 Hz, 1 H), 6.80 – 6.60 (m, 1 H), 5.25 – 5.01 (m, 2 H), 4.07 – 3.89 (m, 1 H), 1.52 – 1.34 (m, 9 H), 1.02 – 0.86 (m, 9 H).
Example B-l, Step b
Acetic acid (15 mL) was added to (S)-tert-butyl (l-((2-amino-4-bromo phenyl)amino)-3,3-dimethyl-l-oxobutan-2-yl)carbamate (1.8 g, 4.50 mmol) and the reaction mixture was heated to 65 °C for overnight. The volatile component was removed in vacuo, and the residue was co-evaporated with dry CH2C12 (2 x 15 mL). The organic phase was washed with saturated NaHC03 solution, brine, dried over Na2S04 and concentrated to obtain (S)-tert-butyl (l-(6-bromo-lH-benzo[d] imidazol-2-yl)-2,2-dimethyl propyl)carbamate (1.68 g) as yellow solid. LC (Condition 1): Rt = 2.19 min. LC/MS: Anal. Calcd. for [M+H]+ Ci7H25BrN302 : 381.11; found 382.2. 1H NMR (DMSO-dg, δ = 2.50 ppm, 300 MHz): δ 12.46 – 12.27 (m, 1 H), 7.82 – 7.65 (m, 1 H), 7.59 – 7.41 (m, 1 H), 7.29 (dt, J= 1.9, 8.5 Hz, 1 H), 7.12 – 6.90 (m, 1 H), 4.64 (d, J= 9.8 Hz, 1 H), 1.44 – 1.27 (m, 9 H), 0.88 (br. s., 9 H).
-1 Step c
To a solution of (S)-tert-butyl (l-(6-bromo-lH-benzo[d]imidazol-2-yl)-2,2-dimethyl propyl) carbamate (1.57 g, 4.11 mmol) in dioxane (25 mL) was added bis (pinacolato)diboron (1.564 g, 6.16 mmol) and potassium acetate (1.209 g, 12.32 mmol). The reaction mixture was purged with argon for 10 min then PdCl2(dppf) (0.150 g, 0.205 mmol) was added to the above reaction mixture and again purged with argon for 5 min. The reaction mixture was heated to 90 °C for overnight. The reaction mixture was diluted with water (15 ml) and extracted with EtOAc (2 x 25 ml). The combined organic phase was washed with brine, dried over Na2S04 and concentrated in vacuo. The crude material was purified by ISCO using 40 g Redisep column, hexane/ethyl acetate as eluant to afford (S)-tert-butyl (2,2-dimethyl-l-(6-(4,4,5 ,5-tetramethyl- 1 ,3 ,2-dioxaborolan-2-yl)- 1 H-benzo[d]imidazol-2-yl)propyl) carbamate (1.35 g) as yellow solid. LC (Condition 1): Rt = 2.21 min. LC/MS: Anal. Calcd. for [M+H]+ C23H37BN304 : 430.29; found 430.4. 1H NMR (CD3OD, δ = 3.34 ppm, 400 MHz): δ 7.98 (s, 1 H), 7.65 (dd, J= 1.0, 8.5 Hz, 1 H), 7.53(d, J= 8.5 Hz, 1 H), 4.73 (br. s., 1 H), 1.37 (s, 12 H), 1.24 (m, 9 H), 1.01 (s, 9 H).
-1 Step d
To a solution of (S)-tert-butyl (2,2-dimethyl-l-(6-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)-lH-benzo[d]imidazol-2-yl)propyl)carbamate (1.114 g, 2.59 mmol) and 4,16-dibromo[2,2]paracyclophane (0.38g, 1.038 mmol) in dioxane (10 mL) was added Cs2C03 (0.845 g, 2.59 mmol) in water (2 mL) and degassed for 10 min.
PdCl2(dppf) (0.038 g, 0.052 mmol) was added to the above reaction mixture and again degassed for 5 min. The reaction mixture was heated to 90 °C for 12 h. Then the reaction mixture was filtered to get Example B-1 Step d which was taken for next step without further purification. LC (Condition 1): Rt = 2.54 min. LC/MS: Anal. Calcd. for [M+H]+ ^0Η63Ν6Ο4 : 811.49; found 811.6. 1H NMR (DMSO-d6, δ = 2.50 ppm, 300 MHz): δ 12.36 (br. s., 2 H), 7.85 – 7.52 (m, 4 H), 7.32 (d, J= 7.9 Hz, 2 H), 7.05 (br. s., 2 H), 6.89 – 6.67 (m, 4 H), 6.54 (br. s., 2 H), 4.72 (d, J= 8.7 Hz, 2 H), 3.57 – 3.44 (m, 2 H), 3.07 (br. s., 2 H), 2.83 (br. s., 2 H), 2.65 (br. s., 2 H), 1.36 (s, 18 H), 1.08 – 0.91 (m, 18 H).
-1 Step e
HC1 in dioxane (4 mL, 24.00 mmol) was added to Example B-1 Step d (0.1 g,
0.102 mmol), and the reaction mixture was allowed to stir at RT for 2 h. Completion of the reaction was monitored by LCMS. The volatile component was removed in vacuo and the residue was washed with diethyl ether and dried to afford Example B-1 Step e (0.07 g) as yellow solid. LC (Condition 1): R, = 2.54 min. LC/MS: Anal.
To a solution of Example B-1 Step e (0.04 g, 0.053 mmol) in DMF (5 mL) was added 4,4-difluorocyclohexanecarboxylic acid (0.017 g, 0.106 mmol), DIPEA (0.055 mL, 0.317 mmol) and HATU (0.030 g, 0.079 mmol). After being stirred for 2 h at room temperature, the volatile component was removed in vacuo and the residue was dissolved in DCM (10 mL), washed with saturated solution of NH4C1, 10% NaHC03 solution, brine, dried over Na2S04 and concentrated in vacuo. The crude was purified by reverse phase HPLC purification to give Example B-1 as a white solid. LC (Condition 1): R, = 2.37 min. LC/MS: Anal. Calcd. for [M+H]+
Starting materials can be obtained from commercial sources or prepared by well-established literature methods known to those of ordinary skill in the art. Acid precursors for the final step can be prepared according to the methods described in U.S. Patent Application Serial No. 13/933495, filed July 2, 2013.
LC/MS Condition 1
Column = Ascentis Express C18, 2.1 X 50 mm, 2.7 um
Solvent A = CH3CN (2%) + 10 mM NH4COOH in H20 (98%)
Solvent B = CH3CN (98%) + 10 mM NH4COOH in H20 (2%)
Start %B = 0; Final %B = 100
Gradient time = 1.4 min; Stop time = 4 min
Stop time = 4 min
Flow Rate = 1 mL/min; Wavelength = 220 nm
LC/MS Condition 2
Column = Waters BEH CI 8, 2.0 x 50 mm, 1.7 μιη
Slovent A = ACN (5%) + H20 (95%) containing 10 mM NH4OAc
Solvent B = ACN (95%) + H20 (5%) containing 10 mM NH4OAc
Start %B = 0; Final %B = 100
Gradient time = 3 min
Flow Rate = 1 mL/min
Wavelength = 220 nm
Temperature = 50 °C
LC/MS Condition 3
Column: Waters Phenomenex CI 8, 2.0 x 30 mm, 3 μιη particle
Mobile Phase A: 10% MeOH:90% Water :0.1%TFA
Mobile Phase B: 90% MeOH: 10% Water :0.1%TFA
Gradient: 0%B, 0-100% B over 3 minutes, then a 1 -minute hold at 100% B Flow: 0.8mL/min
Detection: 220 nm
Temperature: 40 °C
LC/MS Condition 4
Column: Waters BEH CI 8, 2.0 x 50 mm, 1.7 μιη particle
Mobile Phase A: 5:95 acetonitrile: water with 10 mM ammonium acetate Mobile Phase B: 95:5 acetonitrile: water with 10 mM ammonium acetate Gradient: 0%B, 0-100% B over 3 minutes, then a 0.5-minute hold at 100% B Flow: 1 mL/min
Phase II clinical development at AbbVie (previously Abbott) for the treatment of chronic hepatitis C infection in combination with ABT-450/ritonavir and, in combination with peginterferon alpha-2a/ribavirin (pegIFN/RBV) in treatment naïve Hepatitis C virus (HCV) genotype 1 infected patients.
Ombitasvir is Dimethyl ([(2S,5S)-1-(4-tert-butylphenyl) pyrrolidine-2,5diyl]bis{benzene-4,1-diylcarbamoyl(2S)pyrrolidine-2,1-diyl[(2S)-3-methyl-1-oxobutane-1,2diyl]})biscarbamate hydrate. The molecular formula is C50H67N7O8•4.5H2O (hydrate) and the molecular weight for the drug substance is 975.20 (hydrate).
Ombitasvir is in phase II clinical development at AbbVie (previously Abbott) for the treatment of chronic hepatitis C infection in combination with ABT-450/ritonavir and, in combination with peginterferon alpha-2a/ribavirin (pegIFN/RBV) in treatment naïve Hepatitis C virus (HCV) genotype 1 infected patients.
Ombitasvir is part of a fixed-dose formulation with ABT-450/ritonavir that is approved in the U.S. and the E.U.
In January 2013, Abbott spun-off its research-based pharmaceutical business into a newly-formed company AbbVie. In 2013, breakthrough therapy designation was assigned in the U.S. for the treatment of genotype 1 hepatitis C in combination with ABT-450, ritonavir and ABT-333, with and without ribavirin.
Ombitasvir acts by inhibiting the HCV protein NS5A.[5]
Ombitasvir is an orally available inhibitor of the hepatitis C virus (HCV) non-structural protein 5A (NS5A) replication complex, with potential activity against HCV. Upon oral administration and after intracellular uptake, ombitasvir binds to and blocks the activity of the NS5A protein. This results in the disruption of the viral RNA replication complex, blockage of HCV RNA production, and inhibition of viral replication. NS5A, a zinc-binding and proline-rich hydrophilic phosphoprotein, plays a crucial role in HCV RNA replication. HCV is a small, enveloped, single-stranded RNA virus belonging to the Flaviviridae family; HCV infection is associated with the development of hepatocellular carcinoma (HCC).
Ombitasvir hydrate is a NS5A non-nucleoside polymerase inhibitor which is approved as part of a four drug combination for the
treatment of adults with genotype 1 hepatitis C virus infection including those with compensated cirrhosis.REF 6,7
The four drug combination treatment consists of ombitasvir, paritaprevir (XXVII), ritonavir, and dasabuvir (X). This combination treatment is marketed as Viekira PakTM and was developed by Abbvie as an all oral treatment that eliminates the need for pegylated interferon-a injections.
The synthesis of ombitasvir hydrate is shown in Scheme 34.REF 8 Alkylation of 1-(4-nitrophenyl)ethanone (209)
with 2-bromo-1-(4-nitrophenyl)ethanone (208) in the presence of zinc chloride produced diketone 210 in 61% yield.
Asymmetric reduction of the diketone using N,N-diethylaniline borane with (S)-()-a,a-diphenyl-2-pyrrolidinemethanol (211) and trimethoxyborate gave diol 212 in 61% yield and 99.3% ee.
The diol was then treated with methanesulfonic anhydride to generate the corresponding bis-mesylate which was reacted with 4-tert-butylaniline to give pyrrolidine 213 in 51% yield over the two steps.
Hydrogenolysis of the nitro groups was accomplished using Raney nickel catalyst to give bis-aniline 214.
Separately, (L)-valine (216,Scheme 35) was reacted with methyl chloroformate to give the corresponding methyl carbamate in 90% yield which was coupled to L-proline benzyl ester in the presence of EDC and HOBt to give the corresponding dipeptide in 90% yield.
Hydrogenolysis of the benzyl ester group of the protected dipeptide using Pd/alumina catalyst produced dipeptide acid 215. Aniline 214 was treated with two equivalents of acid 215 in the presence of 1-propanephosphonic acid cyclic anhydride (T3P). The crude product was recrystallized from ethanol and heptane to give ombitasvir hydrate (XXV). No yields were provided to the final steps of this synthesis.
7. DeGoey, D. A.; Randolph, J. T.; Liu, D.; Pratt, J.; Hutchins, C.; Donner, P.;Krueger, A. C.; Matulenko, M.; Patel, S.; Motter, C. E.; Nelson, L.; Keddy, R.;Tufano, M.; Caspi, D. D.; Krishnan, P.; Mistry, N.; Koev, G.; Reisch, T. J.;Mondal, R.; Pilot-Matias, T.; Gao, Y.; Beno, D. W.; Maring, C. J.; Molla, A.;Dumas, E.; Campbell, A.; Williams, L.; Collins, C.; Wagner, R.; Kati, W. M. J.
Med. Chem. 2014, 57, 2047. 8. DeGoey, D. A.; Kati, W. M.; Hutchins, C. W.; Donner, P. L.; Krueger, A. C.;Randolph, J. T.; Motter, C. E.; Nelson, L. T.; Patel, S. V.; Matulenko, M. A.;Keddy, R. G.; Jinkerson, T. K.; Soltwedel, T. N.; Liu, D.; Pratt, J. K.; Rockway, T.W.; Maring, C. J.; Hutchinson, D. K.; Flentge, C. A.; Wagner, R.; Tufano, M. D.;Betebenner, D. A.; Lavin, M. J.; Sarris, K.; Woller, K. R.; Wagaw, S. H.; Califano,
J. C.; Li, W.; Caspi, D. D.; Bellizzi, M. E. US Patent 2010317568A1, 2010.
We describe here N-phenylpyrrolidine-based inhibitors of HCV NS5A with excellent potency, metabolic stability, and pharmacokinetics. Compounds with 2S,5S stereochemistry at the pyrrolidine ring provided improved genotype 1 (GT1) potency compared to the 2R,5Ranalogues. Furthermore, the attachment of substituents at the 4-position of the central N-phenyl group resulted in compounds with improved potency. Substitution with tert-butyl, as in compound 38 (ABT-267), provided compounds with low-picomolar EC50 values and superior pharmacokinetics. It was discovered that compound 38 was a pan-genotypic HCV inhibitor, with an EC50 range of 1.7–19.3 pM against GT1a, -1b, -2a, -2b, -3a, -4a, and -5a and 366 pM against GT6a. Compound 38 decreased HCV RNA up to 3.10 log10 IU/mL during 3-day monotherapy in treatment-naive HCV GT1-infected subjects and is currently in phase 3 clinical trials in combination with an NS3 protease inhibitor with ritonavir (r) (ABT-450/r) and an NS5B non-nucleoside polymerase inhibitor (ABT-333), with and without ribavirin.
Dimethyl (2S,2′S)-1,1′-((2S,2′S)-2,2′-(4,4′-((2S,5S)-1-(4-tert-Butylphenyl)pyrrolidine-2,5-diyl)bis(4,1-phenylene))bis(azanediyl)bis(oxomethylene)bis(pyrrolidine-2,1-diyl))bis(3-methyl-1-oxobutane-2,1-diyl)dicarbamate (38)…desired and Dimethyl (2S,2′S)-1,1′-((2S,2′S)-2,2′-(4,4′-((2R,5R)-1-(4-tert-Butylphenyl)pyrrolidine-2,5-diyl)bis(4,1-phenylene))bis(azanediyl)bis(oxomethylene)bis(pyrrolidine-2,1-diyl))bis(3-methyl-1-oxobutane-2,1-diyl)dicarbamate (39)…….undesired
…………….. The resulting mixture was stirred at room temperature for 16 h. The mixture was partitioned between ethyl acetate and water, and the organic layer was washed with saturated aqueous NaHCO3, brine (2×) and dried with Na2SO4. The drying agent was filtered off and the solution was concentrated in vacuo to give a crude product that was purified by column chromatography on silica gel, eluting with a solvent gradient of 2–8% methanol in dichloromethane to give a 1:1 mixture of trans-pyrrolidine isomers (290 mg, 96%). The mixture was separated on a Chiralpak AD-H column, eluting with a mixture of 1 part (2:1 isopropanol/ethanol) and 2 parts hexanes (0.1% TFA).
Example 34A l-(4-fer?-butylphenyl)-2,5-bis(4-nitrophenyl)pyrrolidine The product from Example 1C (3.67 g, 7.51 mmol) and 4-tert-butylaniline (11.86 ml, 75 mmol) in DMF (40 ml) was stirred under nitrogen at 50 °C for 4 h. The resulting mixture was diluted into ethyl acetate, treated with IM HCl, stirred for 10 minutes and filtered to remove solids. The filtrate organic layer was washed twice with brine, dried with sodium sulfate, filtered and evaporated. The residue was purified by chromatography on silica gel eluting with ethyl acetate in hexane (5% to 30%) to give a solid. The solid was triturated in a minimal volume of 1 :9 ethyl acetate/hexane to give a light yellow solid as a mixture of trans and cis isomers (1.21 g, 36%).
Example 34B 4,4′-((2S,5S)-1-(4-tert-butylphenyl)pyrrolidine-2,5-diyl)dianiline and 4,4′-((2R,5R)-1-(4-fert- butylphenyl)pyrrolidine-2,5-diyl)dianiline To a solution of the product from Example 34A (1.1 g, 2.47 mmol) in ethanol (20 ml) and
THF (20 ml) was added PtC>2 (0.22 g, 0.97 mmol) in a 50 ml pressure bottle and stirred under 30 psi hydrogen at room temperature for 1 h. The mixture was filtered through a nylon membrane and evaporated. The residue was purified by chromatography on silica gel eluting with ethyl acetate in hexane (20% to 60%). The title compound eluted as the first of 2 stereoisomers (trans isomer, 0.51 g, 54%).
Example 34C
(2S,2’S)-tert-Butyl 2,2′-(4,4′-((2S,5S)-1-(4-fer/’-butylphenyl)pyrrolidine-2,5-diyl)bis(4,l- phenylene))bis(azanediyl)bis(oxomethylene)dipyrrolidine- 1 -carboxylate and (2S,2’S)-tert-Butyl 2,2′- (4,4′-((2R,5R)-1-(4-tert-butylphenyl)pyrrolidine-2,5-diyl)bis(4,l- phenylene))bis(azanediyl)bis(oxomethylene)dipyrrolidine-1-carboxylate To a mixture of the product from Example 34B (250 mg, 0.648 mmol), (S)-1-(tert- butoxycarbonyl)pyrrolidine-2-carboxylic acid (307 mg, 1.427 mmol) and HATU (542 mg, 1.427 mmol) in DMSO (10 ml) was added Hunig’s base (0.453 ml, 2.59 mmol). The reaction mixture was stirred at room temperature for 1 h. The mixture was partitioned with ethyl acetate and water. The organic layer was washed with brine, dried with sodium sulfate, filtered and evaporated. The residue was purified by chromatography on silica gel eluting with ethyl acetate in hexane (10% to 50%) to give the title compound (500 mg, 99%).
Example 34D
(2S,2’S)-N,N’-(4,4′-((2S,5S)-1-(4-ter/’-butylphenyl)pyrrolidine-2,5-diyl)bis(4,l- phenylene))dipyrrolidine-2-carboxamide and (2S,2’S)-N,N’-(4,4′-((2R,5R)-1-(4-tert- butylphenyl)pyrrolidine-2,5-diyl)bis(4,l-phenylene))dipyrrolidine-2-carboxamide To the product from Example 34C (498 mg, 0.638 mmol) in dichloromethane (4 ml) was added TFA (6 ml). The reaction mixture was stirred at room temperature for 1 h and concentrated in vacuo. The residue was partitioned between 3: 1 CHCl3dsopropyl alcohol and saturated aq. NaHCO3. The aqueous layer was extracted by 3: 1 CHCl3:isopropyl alcohol again. The combined organic layers were dried over
filtered and concentrated to give the title compound (345 mg, 93%).
Example 34E Dimethyl (2S,2’S)-l,r-((2S,2’S)-2,2′-(4,4′-((2S,5S)-1-(4-fert-butylphenyl)pyrrolidine-2,5-diyl)bis(4,l- phenylene))bis(azanediyl)bis(oxomethylene)bis(pyrrolidine-2, 1 -diyl))bis(3-methyl- 1 -oxobutane-2, 1 – diyl)dicarbamate and
The product from Example 34D (29.0 mg, 0.050 mmol), (S)-2-(methoxycarbonylamino)-3- methylbutanoic acid (19.27 mg, 0.110 mmol), EDAC (21.09 mg, 0.110 mmol), HOBT (16.85 mg,
0.110 mmol) and N-methylmorpholine (0.027 ml, 0.250 mmol) were combined in DMF (2 ml). The reaction mixture was stirred at room temperature for 3 h. The mixture was partitioned with ethyl acetate and water. The organic layer was washed with brine twice, dried with sodium sulfate, filtered and evaporated. The residue was purified by chromatography on silica gel eluting with ethyl acetate in hexane (50% to 80%) to give a solid. The solid was triturated with ethyl acetate/hexane to give the title compound (13 mg, 29%). 1H NMR (400 MHz, DMSO-D6) δ ppm 0.85 – 0.95 (m, 12 H) 1.11 (s, 9 H) 1.59 – 1.65 (m, 2 H) 1.79 – 2.04 (m, 8 H) 2.10 – 2.18 (m, 2 H) 2.41-2.46 (m, 2H) 3.52 (s, 6 H)
Hz, 2 H) 7.50 (d, J=8.24 Hz, 4 H) 9.98 (s, 2 H); (ESI+) m/z 895 (M+H)+. The title compound showed an EC50 value of less than about 0.1 nM in HCV lb-Conl replicon assays in the presence of 5% FBS.
The product from Example 34E was purified by chiral chromatography on a Chiralpak AD-H semi-prep column eluting with a 2:1 mixture of hexane:(2: l isopropyl alcohol: EtOH). The title compound was the second of 2 diastereomers to elute. 1H NMR (400 MHz, DMSO-D6) δ ppm 0.87
Example 37A (S)-2,5-dioxopyrrolidin-1-yl 2-(methoxycarbonylamino)-3-methylbutanoate To a mixture of (S)-2-(methoxycarbonylamino)-3-methylbutanoic acid (19.66 g, 112 mmol) and N-hydroxysuccinimide (13.29g, 116 mmol) was added ethyl acetate (250 ml), and the mixture was cooled to 0-5 °C. Diisopropylcarbodiimide (13.88 g, 110 mmol) was added and the reaction mixture was stirred at 0-5 °C for about 1 hour. The reaction mixture was warmed to room temperature. The solids (diisopropylurea by-product) were filtered and rinsed with ethyl acetate. The filtrate was concentrated in vacuo to an oil. Isopropyl alcohol (200 ml) was added to the oil and the mixture was heated to about 50 °C to obtain a homogeneous solution. Upon cooling, crystalline solids formed. The solids were filtered and washed with isopropyl alcohol (3 x 20 ml) and dried to give the title compound as a white solid (23.2 g, 77% yield).
Example 37B
(S)- 1 -((S)-2-(methoxycarbonylamino)-3-methylbutanoyl)pyrrolidine-2-carboxylic acid To a mixture of L-proline (4.44g, 38.6 mmol), water (20 ml), acetonitrile (20 ml) and DIEA (9.5 g, 73.5 mmol) was added a solution of the product from Example 37A (1Og, 36.7 mmol) in acetonitrile (20 inL) over 10 minutes. The reaction mixture was stirred overnight at room temperature. The solution was concentrated under vacuum to remove the acetonitrile. To the resulting clear water solution was added 6N HCl (9 ml) until pH ~ 2 .The solution was transferred to a separatory funnel and 25% NaCl (10 ml) was added and the mixture was extracted with ethyl acetate (75 ml), and then again with ethyl acetate (6 x 20 ml), and the combined extracts were washed with 25% NaCl (2 x 10ml). The solvent was evaporated to give a thick oil. Heptane was added and the solvent was evaporated to give a foam, which was dried under high vacuum. Diethyl ether was added and the solvent was evaporated to give a foam, which was dried under high vacuum to give the title compound (10.67g) as a white solid.
The compound of Example 37B can also be prepreared according to the following procedure: To a flask was charged L- valine (35 g, 299 mmol), IN sodium hydroxide solution (526 ml,
526 mmol) and sodium carbonate (17.42 g, 164 mmol). The mixture was stirred for 15 min to dissolve solids and then cooled to 15 °C. Methyl chloroformate (29.6 g, 314 mmol) was added slowly to the reaction mixture. The mixture was then stirred at rt for 30 min. The mixture was cooled to 15 °C and pH adjusted to -5.0 with concentrated HCl solution. 100 inL of 2-methytetrahydrofuran (2- MeTHF) was added and the adjustment of pH continued until the pH reached ~ 2.0. 150 mL of 2- MeTHF was added and the mixture was stirred for 15 min. Layers were separated and the aqueous layer extracted with 100 mL of 2-MeTHF. The combined organic layer was dried over anhyd Na2SC^ and filtered, and Na2SC^ cake was washed with 50 mL of 2-MeTHF. The product solution was concentrated to ~ 100 mL, chased with 120 mL of IPAc twice. 250 mL of heptanes was charged slowly and then the volume of the mixture was concentrated to 300 mL. The mixture was heated to 45 °C and 160 mL of heptanes charged. The mixture was cooled to rt in 2h, stirred for 30 min, filtered and washed with 2-MeTHF/heptanes mixture (1:7, 80 inL). The wetcake was dried at 55 °C for 24 h to give 47.1 g of Moc-L- VaI-OH product as a white solid (90%).
Moc-L- VaI-OH (15O g, 856 mmol), HOBt hydrate (138 g, 899 mmol) and DMF (1500 ml) were charged to a flask. The mixture was stirred for 15 min to give a clear solution. EDC hydrochloride (172 g, 899 mmol) was charged and mixed for 20 min. The mixture was cooled to 13
°C and (L)-proline benzyl ester hydrochloride (207 g, 856 mmol) charged. Triethylamine (109 g,
1079 mmol) was then charged in 30 min. The resulting suspension was mixed at rt for 1.5 h. The reaction mixture was cooled to 15 °C and 1500 mL of 6.7% NaHCO3 charged in 1.5 h, followed by the addition of 1200 mL of water over 60 min. The mixture was stirred at rt for 30 min, filtered and washed with water/DMF mixture (1 :2, 250 mL) and then with water (1500 mL). The wetcake was dried at 55 °C for 24 h to give 282 g of product as a white solid (90%).
The resulting solids (40 g) and 5% Pd/ Alumina were charged to a Parr reactor followed by THF (160 mL). The reactor was sealed and purged with nitrogen (6 x 20 psig) followed by a hydrogen purge (6 x 30 psig). The reactor was pressurized to 30 psig with hydrogen and agitated at room temperature for approximately 15 hours. The resulting slurry was filtered through a GF/F filter and concentrated to approximately 135 g solution. Heptane was added (120 mL), and the solution was stirred until solids formed. After an addition 2 – 3 hours additional heptane was added drop-wise (240 mL), the slurry was stirred for approximately 1 hour, then filtered. The solids were dried to afford the title compound.
The product from Example 32 (5.01 g, 13.39 mmol) was combined with 2- methyltetrahydrofuran (70 mL) and cooled to -5 °C, and N,N-diisopropylethylamine (6.81 g, 52.7 mmol) was added over 30 seconds. Separately, a solution of methanesulfonic anhydride (6.01 g, 34.5 mmol) in 2-methyltetrahydrofuran (30 mL) was prepared and added to the diol slurry over 3 min., maintaining the internal temperature between -15 °C and -25 °C. After mixing for 5 min at -15 °C, the cooling bath was removed and the reaction was allowed to warm slowly to 23 °C and mixed for 30 minutes. After reaction completion, the crude slurry was carried immediately into the next step.
To the crude product solution from Example 37C (7.35 g, 13.39 mmol) was added 4-tert- butylaniline (13.4 g, 90 mmol) at 23 °C over 1 minute. The reaction was heated to 65 °C for 2 h. After completion, the reaction mixture was cooled to 23 °C and diluted with 2-methyltetrahydrofuran (100 mL) and 1 M HCl (150 mL). After partitioning the phases, the organic phase was treated with 1 M HCl (140 mL), 2-methyltetrahydrofuran (50 mL), and 25 wt% aq. NaCl (100 mL), and the phases were partitioned. The organic phase was washed with 25 wt% aq. NaCl (50 mL), dried over MgSO/t, filtered, and concentrated in vacuo to approximately 20 mL. Heptane (30 mL) and additional 2- methyltetrahydrofuran were added in order to induce crystallization. The slurry was concentrated further, and additional heptane (40 mL) was slowly added and the slurry was filtered, washing with 2- methyltetrahydrofuran:heptane (1:4, 20 mL). The solids were suspended in MeOH (46 mL) for 3 h, filtered, and the wet solid was washed with additional MeOH (18 mL). The solid was dried at 45 °C in a vacuum oven for 16 h to provide the title compound (3.08 g, 51% 2-step yield).
To a 160 ml Parr stirrer hydrogenation vessel was added the product from Example 37D (2 g, 4.49 mmol), followed by 60 ml of THF, and Raney Nickel Grace 2800 (1 g, 50 wt% (dry basis)) under a stream of nitrogen. The reactor was assembled and purged with nitrogen (8 x 20 psig) followed by purging with hydrogen (8 x 30 psig). The reactor was then pressurized to 30 psig with hydrogen and agitation (700 rpm) began and continued for a total of 16 h at room temperature. The slurry was filtered by vacuum filtration using a GF/F Whatman glass fiber filter. Evaporation of the filtrate to afford a slurry followed by the addition heptane and filtration gave the crude title compound, which was dried and used directly in the next step.
Example 37F dimethyl (2S,2’S)-l,r-((2S,2’S)-2,2′-(4,4′-((2S,5S)-1-(4-tert-butylphenyl)pyrrolidine-2,5-diyl)bis(4, l- phenylene)bis(azanediyl)bis(oxomethylene))bis(pyrrolidine-2, 1 -diyl))bis(3-methyl- 1 -oxobutane-2, 1 – diy 1) die arb amate To a solution of the product from Example 37E (1.64 g, 4.25 mmol) in DMF (20 ml), the product from Example 37B (2.89 g, 10.63 mmol), and HATU (4.04 g, 10.63 mmol) in DMF (15OmL) was added triethylamine (1.07 g, 10.63 mmol), and the solution was stirred at room temperature for 90 min. To the reaction mixture was poured 20 mL of water, and the white precipitate obtained was filtered, and the solid was washed with water (3×5 mL). The solid was blow dried for Ih. The crude material was loaded on a silica gel column and eluted with a gradient starting with ethyl acetate/ heptane (3/7), and ending with pure ethyl acetate. The desired fractions were combined and solvent distilled off to give a very light yellow solid, which was dried at 45 °C in a vacuum oven with nitrogen purge for 15 h to give the title compound (2.3 g, 61% yield). 1H NMR (400 MHz, DMSO- D6) δ ppm 0.88 (d, J=6.61 Hz, 6 H) 0.93 (d, J=6.72 Hz, 6 H) 1.11 (s, 9 H) 1.63 (d, J=5.42 Hz, 2 H) 1.80 – 2.04 (m, 8 H) 2.09 – 2.19 (m, 2 H) 2.44 – 2.47 (m, 2 H) 3.52 (s, 6 H) 3.59 – 3.66 (m, 2 H) 3.77 – 3.84 (m, 2 H) 4.02 (t, J=8.40 Hz, 2 H) 4.42 (dd, J=7.86, 4.83 Hz, 2 H) 5.14 (d, J=6.18 Hz, 2 H) 6.17 (d, J=8.67 Hz, 2 H) 6.94 (d, J=8.78 Hz, 2 H) 7.13 (d, J=8.46 Hz, 4 H) 7.31 (d, J=8.35 Hz, 2 H) 7.50 (d, J=8.35 Hz, 4 H) 9.98 (s, 2 H).
Alternately, the product from example 37E (11.7 g, 85 wt%, 25.8 mmol) and the product from example 37B (15.45 g, 56.7 mmol) are suspended in EtOAc (117 mL), diisopropylethylamine (18.67 g, 144 mmol) is added and the solution is cooled to 0 °C. In a separate flask, 1-propanephosphonic acid cyclic anhydride (T3P®) (46.0 g, 50 wt% in EtOAc, 72.2 mmol) was dissolved in EtOAc (58.5 mL), and charged to an addition funnel. The T3P solution is added to the reaction mixture drop-wise over 3-4 h and stirred until the reaction is complete. The reaction is warmed to room temperature,and washed with IM HCl/7.5 wt% NaCl (100 mL), then washed with 5% NaHCO3 (100 mL), then washed with 5% NaCl solution (100 mL). The solution was concentrated to approximately 60 mL, EtOH (300 mL) was added, and the solution was concentrated to 84 g solution.
A portion of the EtOH solution of product (29 g) was heated to 40 °C, and added 134 g 40 w% EtOH in H2O. A slurry of seeds in 58 wt/wt% EtOH/H2O was added, allowed to stir at 40 °C for several hours, then cooled to 0 °C. The slurry is then filtered, and washed with 58wt/wt% EtOH/H2O. The product is dried at 40 – 60 °C under vacuum, and then rehydrated by placing a tray of water in the vacuum oven to give the title compound. The title compound showed an EC50 value of less than about 0.1 nM in HCV lb-Conl replicon assays in the presence of 5% FBS.
Example 34A l-(4-fer?-butylphenyl)-2,5-bis(4-nitrophenyl)pyrrolidine The product from Example 1C (3.67 g, 7.51 mmol) and 4-tert-butylaniline (11.86 ml, 75 mmol) in DMF (40 ml) was stirred under nitrogen at 50 °C for 4 h. The resulting mixture was diluted into ethyl acetate, treated with IM HCl, stirred for 10 minutes and filtered to remove solids. The filtrate organic layer was washed twice with brine, dried with sodium sulfate, filtered and evaporated. The residue was purified by chromatography on silica gel eluting with ethyl acetate in hexane (5% to 30%) to give a solid. The solid was triturated in a minimal volume of 1 :9 ethyl acetate/hexane to give a light yellow solid as a mixture of trans and cis isomers (1.21 g, 36%).
Example 34B 4,4′-((2S,5S)-1-(4-tert-butylphenyl)pyrrolidine-2,5-diyl)dianiline and 4,4′-((2R,5R)-1-(4-fert- butylphenyl)pyrrolidine-2,5-diyl)dianiline To a solution of the product from Example 34A (1.1 g, 2.47 mmol) in ethanol (20 ml) and
THF (20 ml) was added PtC>2 (0.22 g, 0.97 mmol) in a 50 ml pressure bottle and stirred under 30 psi hydrogen at room temperature for 1 h. The mixture was filtered through a nylon membrane and evaporated. The residue was purified by chromatography on silica gel eluting with ethyl acetate in hexane (20% to 60%). The title compound eluted as the first of 2 stereoisomers (trans isomer, 0.51 g, 54%).
Example 34C
(2S,2’S)-tert-Butyl 2,2′-(4,4′-((2S,5S)-1-(4-fer/’-butylphenyl)pyrrolidine-2,5-diyl)bis(4,l- phenylene))bis(azanediyl)bis(oxomethylene)dipyrrolidine- 1 -carboxylate and (2S,2’S)-tert-Butyl 2,2′- (4,4′-((2R,5R)-1-(4-tert-butylphenyl)pyrrolidine-2,5-diyl)bis(4,l- phenylene))bis(azanediyl)bis(oxomethylene)dipyrrolidine-1-carboxylate To a mixture of the product from Example 34B (250 mg, 0.648 mmol), (S)-1-(tert- butoxycarbonyl)pyrrolidine-2-carboxylic acid (307 mg, 1.427 mmol) and HATU (542 mg, 1.427 mmol) in DMSO (10 ml) was added Hunig’s base (0.453 ml, 2.59 mmol). The reaction mixture was stirred at room temperature for 1 h. The mixture was partitioned with ethyl acetate and water. The organic layer was washed with brine, dried with sodium sulfate, filtered and evaporated. The residue was purified by chromatography on silica gel eluting with ethyl acetate in hexane (10% to 50%) to give the title compound (500 mg, 99%).
Example 34D
(2S,2’S)-N,N’-(4,4′-((2S,5S)-1-(4-ter/’-butylphenyl)pyrrolidine-2,5-diyl)bis(4,l- phenylene))dipyrrolidine-2-carboxamide and (2S,2’S)-N,N’-(4,4′-((2R,5R)-1-(4-tert- butylphenyl)pyrrolidine-2,5-diyl)bis(4,l-phenylene))dipyrrolidine-2-carboxamide To the product from Example 34C (498 mg, 0.638 mmol) in dichloromethane (4 ml) was added TFA (6 ml). The reaction mixture was stirred at room temperature for 1 h and concentrated in vacuo. The residue was partitioned between 3: 1 CHCl3dsopropyl alcohol and saturated aq. NaHCO3. The aqueous layer was extracted by 3: 1 CHCl3:isopropyl alcohol again. The combined organic layers were dried over
filtered and concentrated to give the title compound (345 mg, 93%).
Example 34E Dimethyl (2S,2’S)-l,r-((2S,2’S)-2,2′-(4,4′-((2S,5S)-1-(4-fert-butylphenyl)pyrrolidine-2,5-diyl)bis(4,l- phenylene))bis(azanediyl)bis(oxomethylene)bis(pyrrolidine-2, 1 -diyl))bis(3-methyl- 1 -oxobutane-2, 1 – diyl)dicarbamate and
The product from Example 34D (29.0 mg, 0.050 mmol), (S)-2-(methoxycarbonylamino)-3- methylbutanoic acid (19.27 mg, 0.110 mmol), EDAC (21.09 mg, 0.110 mmol), HOBT (16.85 mg,
0.110 mmol) and N-methylmorpholine (0.027 ml, 0.250 mmol) were combined in DMF (2 ml). The reaction mixture was stirred at room temperature for 3 h. The mixture was partitioned with ethyl acetate and water. The organic layer was washed with brine twice, dried with sodium sulfate, filtered and evaporated. The residue was purified by chromatography on silica gel eluting with ethyl acetate in hexane (50% to 80%) to give a solid. The solid was triturated with ethyl acetate/hexane to give the title compound (13 mg, 29%). 1H NMR (400 MHz, DMSO-D6) δ ppm 0.85 – 0.95 (m, 12 H) 1.11 (s, 9 H) 1.59 – 1.65 (m, 2 H) 1.79 – 2.04 (m, 8 H) 2.10 – 2.18 (m, 2 H) 2.41-2.46 (m, 2H) 3.52 (s, 6 H)
Hz, 2 H) 7.50 (d, J=8.24 Hz, 4 H) 9.98 (s, 2 H); (ESI+) m/z 895 (M+H)+. The title compound showed an EC50 value of less than about 0.1 nM in HCV lb-Conl replicon assays in the presence of 5% FBS.
The product from Example 34E was purified by chiral chromatography on a Chiralpak AD-H semi-prep column eluting with a 2:1 mixture of hexane:(2: l isopropyl alcohol: EtOH). The title compound was the second of 2 diastereomers to elute. 1H NMR (400 MHz, DMSO-D6) δ ppm 0.87
Example 37A (S)-2,5-dioxopyrrolidin-1-yl 2-(methoxycarbonylamino)-3-methylbutanoate To a mixture of (S)-2-(methoxycarbonylamino)-3-methylbutanoic acid (19.66 g, 112 mmol) and N-hydroxysuccinimide (13.29g, 116 mmol) was added ethyl acetate (250 ml), and the mixture was cooled to 0-5 °C. Diisopropylcarbodiimide (13.88 g, 110 mmol) was added and the reaction mixture was stirred at 0-5 °C for about 1 hour. The reaction mixture was warmed to room temperature. The solids (diisopropylurea by-product) were filtered and rinsed with ethyl acetate. The filtrate was concentrated in vacuo to an oil. Isopropyl alcohol (200 ml) was added to the oil and the mixture was heated to about 50 °C to obtain a homogeneous solution. Upon cooling, crystalline solids formed. The solids were filtered and washed with isopropyl alcohol (3 x 20 ml) and dried to give the title compound as a white solid (23.2 g, 77% yield).
Example 37B
(S)- 1 -((S)-2-(methoxycarbonylamino)-3-methylbutanoyl)pyrrolidine-2-carboxylic acid To a mixture of L-proline (4.44g, 38.6 mmol), water (20 ml), acetonitrile (20 ml) and DIEA (9.5 g, 73.5 mmol) was added a solution of the product from Example 37A (1Og, 36.7 mmol) in acetonitrile (20 inL) over 10 minutes. The reaction mixture was stirred overnight at room temperature. The solution was concentrated under vacuum to remove the acetonitrile. To the resulting clear water solution was added 6N HCl (9 ml) until pH ~ 2 .The solution was transferred to a separatory funnel and 25% NaCl (10 ml) was added and the mixture was extracted with ethyl acetate (75 ml), and then again with ethyl acetate (6 x 20 ml), and the combined extracts were washed with 25% NaCl (2 x 10ml). The solvent was evaporated to give a thick oil. Heptane was added and the solvent was evaporated to give a foam, which was dried under high vacuum. Diethyl ether was added and the solvent was evaporated to give a foam, which was dried under high vacuum to give the title compound (10.67g) as a white solid.
The compound of Example 37B can also be prepreared according to the following procedure: To a flask was charged L- valine (35 g, 299 mmol), IN sodium hydroxide solution (526 ml,
526 mmol) and sodium carbonate (17.42 g, 164 mmol). The mixture was stirred for 15 min to dissolve solids and then cooled to 15 °C. Methyl chloroformate (29.6 g, 314 mmol) was added slowly to the reaction mixture. The mixture was then stirred at rt for 30 min. The mixture was cooled to 15 °C and pH adjusted to -5.0 with concentrated HCl solution. 100 inL of 2-methytetrahydrofuran (2- MeTHF) was added and the adjustment of pH continued until the pH reached ~ 2.0. 150 mL of 2- MeTHF was added and the mixture was stirred for 15 min. Layers were separated and the aqueous layer extracted with 100 mL of 2-MeTHF. The combined organic layer was dried over anhyd Na2SC^ and filtered, and Na2SC^ cake was washed with 50 mL of 2-MeTHF. The product solution was concentrated to ~ 100 mL, chased with 120 mL of IPAc twice. 250 mL of heptanes was charged slowly and then the volume of the mixture was concentrated to 300 mL. The mixture was heated to 45 °C and 160 mL of heptanes charged. The mixture was cooled to rt in 2h, stirred for 30 min, filtered and washed with 2-MeTHF/heptanes mixture (1:7, 80 inL). The wetcake was dried at 55 °C for 24 h to give 47.1 g of Moc-L- VaI-OH product as a white solid (90%).
Moc-L- VaI-OH (15O g, 856 mmol), HOBt hydrate (138 g, 899 mmol) and DMF (1500 ml) were charged to a flask. The mixture was stirred for 15 min to give a clear solution. EDC hydrochloride (172 g, 899 mmol) was charged and mixed for 20 min. The mixture was cooled to 13
°C and (L)-proline benzyl ester hydrochloride (207 g, 856 mmol) charged. Triethylamine (109 g,
1079 mmol) was then charged in 30 min. The resulting suspension was mixed at rt for 1.5 h. The reaction mixture was cooled to 15 °C and 1500 mL of 6.7% NaHCO3 charged in 1.5 h, followed by the addition of 1200 mL of water over 60 min. The mixture was stirred at rt for 30 min, filtered and washed with water/DMF mixture (1 :2, 250 mL) and then with water (1500 mL). The wetcake was dried at 55 °C for 24 h to give 282 g of product as a white solid (90%).
The resulting solids (40 g) and 5% Pd/ Alumina were charged to a Parr reactor followed by THF (160 mL). The reactor was sealed and purged with nitrogen (6 x 20 psig) followed by a hydrogen purge (6 x 30 psig). The reactor was pressurized to 30 psig with hydrogen and agitated at room temperature for approximately 15 hours. The resulting slurry was filtered through a GF/F filter and concentrated to approximately 135 g solution. Heptane was added (120 mL), and the solution was stirred until solids formed. After an addition 2 – 3 hours additional heptane was added drop-wise (240 mL), the slurry was stirred for approximately 1 hour, then filtered. The solids were dried to afford the title compound.
The product from Example 32 (5.01 g, 13.39 mmol) was combined with 2- methyltetrahydrofuran (70 mL) and cooled to -5 °C, and N,N-diisopropylethylamine (6.81 g, 52.7 mmol) was added over 30 seconds. Separately, a solution of methanesulfonic anhydride (6.01 g, 34.5 mmol) in 2-methyltetrahydrofuran (30 mL) was prepared and added to the diol slurry over 3 min., maintaining the internal temperature between -15 °C and -25 °C. After mixing for 5 min at -15 °C, the cooling bath was removed and the reaction was allowed to warm slowly to 23 °C and mixed for 30 minutes. After reaction completion, the crude slurry was carried immediately into the next step.
To the crude product solution from Example 37C (7.35 g, 13.39 mmol) was added 4-tert- butylaniline (13.4 g, 90 mmol) at 23 °C over 1 minute. The reaction was heated to 65 °C for 2 h. After completion, the reaction mixture was cooled to 23 °C and diluted with 2-methyltetrahydrofuran (100 mL) and 1 M HCl (150 mL). After partitioning the phases, the organic phase was treated with 1 M HCl (140 mL), 2-methyltetrahydrofuran (50 mL), and 25 wt% aq. NaCl (100 mL), and the phases were partitioned. The organic phase was washed with 25 wt% aq. NaCl (50 mL), dried over MgSO/t, filtered, and concentrated in vacuo to approximately 20 mL. Heptane (30 mL) and additional 2- methyltetrahydrofuran were added in order to induce crystallization. The slurry was concentrated further, and additional heptane (40 mL) was slowly added and the slurry was filtered, washing with 2- methyltetrahydrofuran:heptane (1:4, 20 mL). The solids were suspended in MeOH (46 mL) for 3 h, filtered, and the wet solid was washed with additional MeOH (18 mL). The solid was dried at 45 °C in a vacuum oven for 16 h to provide the title compound (3.08 g, 51% 2-step yield).
To a 160 ml Parr stirrer hydrogenation vessel was added the product from Example 37D (2 g, 4.49 mmol), followed by 60 ml of THF, and Raney Nickel Grace 2800 (1 g, 50 wt% (dry basis)) under a stream of nitrogen. The reactor was assembled and purged with nitrogen (8 x 20 psig) followed by purging with hydrogen (8 x 30 psig). The reactor was then pressurized to 30 psig with hydrogen and agitation (700 rpm) began and continued for a total of 16 h at room temperature. The slurry was filtered by vacuum filtration using a GF/F Whatman glass fiber filter. Evaporation of the filtrate to afford a slurry followed by the addition heptane and filtration gave the crude title compound, which was dried and used directly in the next step.
Example 37F dimethyl (2S,2’S)-l,r-((2S,2’S)-2,2′-(4,4′-((2S,5S)-1-(4-tert-butylphenyl)pyrrolidine-2,5-diyl)bis(4, l- phenylene)bis(azanediyl)bis(oxomethylene))bis(pyrrolidine-2, 1 -diyl))bis(3-methyl- 1 -oxobutane-2, 1 – diy 1) die arb amate To a solution of the product from Example 37E (1.64 g, 4.25 mmol) in DMF (20 ml), the product from Example 37B (2.89 g, 10.63 mmol), and HATU (4.04 g, 10.63 mmol) in DMF (15OmL) was added triethylamine (1.07 g, 10.63 mmol), and the solution was stirred at room temperature for 90 min. To the reaction mixture was poured 20 mL of water, and the white precipitate obtained was filtered, and the solid was washed with water (3×5 mL). The solid was blow dried for Ih. The crude material was loaded on a silica gel column and eluted with a gradient starting with ethyl acetate/ heptane (3/7), and ending with pure ethyl acetate. The desired fractions were combined and solvent distilled off to give a very light yellow solid, which was dried at 45 °C in a vacuum oven with nitrogen purge for 15 h to give the title compound (2.3 g, 61% yield). 1H NMR (400 MHz, DMSO- D6) δ ppm 0.88 (d, J=6.61 Hz, 6 H) 0.93 (d, J=6.72 Hz, 6 H) 1.11 (s, 9 H) 1.63 (d, J=5.42 Hz, 2 H) 1.80 – 2.04 (m, 8 H) 2.09 – 2.19 (m, 2 H) 2.44 – 2.47 (m, 2 H) 3.52 (s, 6 H) 3.59 – 3.66 (m, 2 H) 3.77 – 3.84 (m, 2 H) 4.02 (t, J=8.40 Hz, 2 H) 4.42 (dd, J=7.86, 4.83 Hz, 2 H) 5.14 (d, J=6.18 Hz, 2 H) 6.17 (d, J=8.67 Hz, 2 H) 6.94 (d, J=8.78 Hz, 2 H) 7.13 (d, J=8.46 Hz, 4 H) 7.31 (d, J=8.35 Hz, 2 H) 7.50 (d, J=8.35 Hz, 4 H) 9.98 (s, 2 H).
Alternately, the product from example 37E (11.7 g, 85 wt%, 25.8 mmol) and the product from example 37B (15.45 g, 56.7 mmol) are suspended in EtOAc (117 mL), diisopropylethylamine (18.67 g, 144 mmol) is added and the solution is cooled to 0 °C. In a separate flask, 1-propanephosphonic acid cyclic anhydride (T3P®) (46.0 g, 50 wt% in EtOAc, 72.2 mmol) was dissolved in EtOAc (58.5 mL), and charged to an addition funnel. The T3P solution is added to the reaction mixture drop-wise over 3-4 h and stirred until the reaction is complete. The reaction is warmed to room temperature,and washed with IM HCl/7.5 wt% NaCl (100 mL), then washed with 5% NaHCO3 (100 mL), then washed with 5% NaCl solution (100 mL). The solution was concentrated to approximately 60 mL, EtOH (300 mL) was added, and the solution was concentrated to 84 g solution.
A portion of the EtOH solution of product (29 g) was heated to 40 °C, and added 134 g 40 w% EtOH in H2O. A slurry of seeds in 58 wt/wt% EtOH/H2O was added, allowed to stir at 40 °C for several hours, then cooled to 0 °C. The slurry is then filtered, and washed with 58wt/wt% EtOH/H2O. The product is dried at 40 – 60 °C under vacuum, and then rehydrated by placing a tray of water in the vacuum oven to give the title compound. The title compound showed an EC50 value of less than about 0.1 nM in HCV lb-Conl replicon assays in the presence of 5% FBS.
To (S)-(-)-α,α-diphenyl-2-pyrrohdinemethanol (2 71 g, 10 70 mmol) was added THF (80 mL) at 23 °C The very thin suspension was treated with t11methyl borate (1 44 g, 13 86 mmol) over 30 seconds, and the resulting solution was mixed at 23 °C for 1 h The solution was cooled to 16-19 °C, and N,N-diethylanilme borane (21 45 g, 132 mmol) was added dropwise via syringe over 3-5 mm (caution vigorous H2 evolution), while the internal temperature was maintained at 16-19 °C After 15 mm, the H2 evolution had ceased To a separate vessel was added the product from Example IA (22 04 g, 95 wt%, 63 8 mmol), followed by THF (80 mL), to form an orange slurry After cooling the slurry to 11 °C, the borane solution was transferred via cannula into the dione slurry over 3-5 min During this period, the internal temperature of the slurry rose to 16 °C After the addition was complete, the reaction was maintained at 20-27 °C for an additional 2 5 h After reaction completion, the mixture was cooled to 5 °C and methanol (16 7 g, 521 mmol) was added dropwise over 5-10 mm, maintaining an internal temperature <20 °C (note vigorous H2 evolution) After the exotherm had ceased (ca 10 mm), the temperature was adjusted to 23 °C, and the reaction was mixed until complete dissolution of the solids had occurred Ethyl acetate (300 mL) and 1 M HCl (120 mL) were added, and the phases were partitioned The organic phase was then washed successively with 1 M HCl (2 x 120 mL), H2O (65 mL), and 10% aq NaCl (65 mL) The orgamcs were dried over MgSO4, filtered, and concentrated in vacuo Crystallization of the product occurred during the concentration The slurry was warmed to 50 °C, and heptane (250 inL) was added over 15 min. The slurry was then allowed to mix at 23 °C for 30 min and filtered. The wet cake was washed with 3: 1 heptane:ethyl acetate (75 mL), and the orange, crystalline solids were dried at 45 °C for 24 h to provide the title compound (15.35 g, 99.3% ee, 61% yield), which was contaminated with 11% of the meso isomer (vs. dl isomer).
Jordan J. Feld; Kris V. Kowdley; Eoin Coakley; Samuel Sigal; David R. Nelson; Darrell Crawford; Ola Weiland; Humberto Aguilar; Junyuan Xiong; Tami Pilot-Matias; Barbara DaSilva-Tillmann; Lois Larsen; Thomas Podsadecki & Barry Bernstein (2014). “Treatment of HCV with ABT-450/r–Ombitasvir and Dasabuvir with Ribavirin”. N Engl J Med370: 1594–1603. doi:10.1056/NEJMoa1315722.
. Mass spectrometry was performed for this isolated impurity by ESI technique in positive mode. The positive DI-MS spectrum of isolated impurity exhibited molecular ion peak as [M + H]+ at m/z 592.29 and as sodium adduct [M + Na]+ at m/z 614.28. The MS/MS data displayed a dominant fragment at m/z 574.28 which is 17 amu less than the molecular ion peak indicating a removal of the hydroxyl group.
This indicates that the impurity is Metoprolol-lactose adduct as proposed. The high resolution mass proposed
the probable molecular formula C27H45NO13.
The 1H NMR spectrum of this impurity displayed signals at δ = 1.27–1.30(6H), δ = 2.73–2.76 (2H), δ = 2.95–2.98 (1H), δ = 3.12–3.18(1H), δ = 3.23 (3H), δ = 3.44–3.68(11H), δ = 3.73–3.77(2H), δ = 3.83–3.84(2H), δ = 4.15–4.17(3H), δ = 4.41–4.43(1H), δ = 4.64–4.67 (1H), δ = 6.68–6.90 (2H), δ = 7.15–7.17(2H)
corresponding to 37 protons, indicating the Metoprolol adduct impurity possibility as it contains total 45 protons out of which 8 protons are of hydroxyl groups of lactose.
The 1H and 13C NMR spectra of Metoprolol adduct impurity and Metoprolol tartrate was compared and significant changes were observed. In 1H NMR spectrum of impurity additional 13 protons in aliphatic region were observed. While in 13C NMR, additional 12 carbon signals can be seen. Methylene carbons C21 and C16 were observed at 60.5 and 61.8 ppm respectively and 10 carbon signals were observed between 68.5 ppm to 103 ppm. These signals
confirmed the presence of both lactose as well as Metoprolol moieties in the impurity.
Further to confirm the exact structure of Metoprolol adduct impurity, the 2D NMR HSQC has also been reviewed (see
Supplementary Fig. S-5). It was observed that the proton in aliphatic region showing doublet at (4.41–4.43) ppm corresponds to C22 which appeared at 103 ppm. This confirms the presence of anomeric carbon of pyranose ring. Also C17 appeared at 95.4 ppm found to be quaternary carbon which confirms the presence of furanose anomeric carbon. Proton corresponding to C20 found to shown multiplet in the region of (4.15–4.17) ppm which confirms that C20 is from furanose ring. Apart from these interactions, carbon signals appeared at 70.7, 72.2, 74.5, 61.8 ppm confirming the arabinosyl moiety.
Based on the above observations it has been confirmed that the impurity is Metoprolol lactose adduct and the ‘glucose moiety’ of lactose present in adduct exists in furanose form
The 1H Metoprolol tartrate
The 13C NMR spectra Metoprolol tartrate
.
The 1H spectra of Metoprolol adduct impurity
The13C NMR spectra of Metoprolol adduct impurity
HSQC spectra of Metoprolol adduct impurity
Identification, synthesis, isolation and characterization of new impurity in metoprolol tartrate tablets
Ataluren, formerly known as PTC124, is a pharmaceutical drug for the treatment of Duchenne muscular dystrophy and potentially other genetic disorders. It was designed by PTC Therapeutics and is sold under the trade name Translarna in the European Union.
Ataluren was approved by European Medicine Agency (EMA) on July 31, 2014. It was developed and marketed as Translarna® by PTC Therapeutics.
Ataluren was regulator of nonsense mutations indicated for the treatment of Duchenne muscular dystrophy resulting from a nonsense mutation in the dystrophin gene, in ambulatory patients aged 5 years and older.
Translarna® is available as granules for oral use, containing 125 mg, 250 mg or 1000 mg of free Ataluren. The recommended dose is 10 mg/kg body weight in the morning, 10 mg/kg body weight at midday, and 20 mg/kg body weight in the evening.
Medical uses
Ataluren has been tested on healthy humans and humans carrying genetic disorders caused by nonsense mutations,[1][2] such as some people with cystic fibrosis and Duchenne muscular dystrophy. It is approved for the use in Duchenne in the European Union.
Mechanism of action
Ataluren makes ribosomes less sensitive to premature stop codons (referred to as “read-through”). This may be beneficial in diseases such as Duchenne muscular dystrophy where the mRNA contains a mutation causing premature stop codons or nonsense codons. Studies have demonstrated that PTC124 treatment increases expression of full-length dystrophin protein in human and mouse primary muscle cells containing the premature stop codon mutation for Duchenne muscular dystrophy and rescues striated muscle function.[3] Studies in mice with the premature stop codon mutation for cystic fibrosis demonstrated increased CFTR protein production and function.[4] The European Medicines Agency review on the approval of ataluren concluded that “the non-clinical data available were considered sufficient to support the proposed mechanism of action and to alleviate earlier concerns on the selectivity of ataluren for premature stop codons.” [5]
In cystic fibrosis, early studies of ataluren show that it improves nasal potential difference.[6] Ataluren appears to be most effective for the stop codon ‘UGA’.[1]
History
Clinical trials
In 2010, PTC Therapeutics released preliminary results of its phase 2b clinical trial for Duchenne muscular dystrophy, with participants not showing a significant improvement in the six minute walk distance after the 48 weeks of the trial.[7] This failure resulted in the termination of a $100 million deal with Genzyme to pursue the drug.
Phase 2 clinical trials were successful for cystic fibrosis in Israel, France and Belgium.[8] Multicountry phase 3 clinical trials are currently in progress for cystic fibrosis in Europe and the USA.[9]
Approval
On 23 May 2014 ataluren received a positive opinion from the Committee for Medicinal Products for Human Use (CHMP) of the European Medicines Agency (EMA).[10]Translarna was first available in Germany, the first EU country to launch the new medicine.[11]
In August 2014, ataluren received market authorization from the European Commission to treat patients with nonsense mutation Duchenne muscular dystrophy. A confirmatory phase III clinical trial is ongoing.[11] The drug does not yet have approval by the US Food and Drug Administration.
In October 2015, NICE asked for further evidence of benefit to justify the “very high cost”.[12] NICE estimated that for a typical patient, treatment would cost £220,256 per year.
In February 2016, FDA declined to approve or even discuss PTC Therapeutics application for ataluren because it deemed the data presented by the developer “insufficient to warrant a review”.[13]
Samples were analyzed for purity on an Agilent 1200 series LC/MS equipped with a Luna® C18 reverse phase (3 micron, 3 x 75 mm) column having a flow rate of 0.8-1.0 mL/min. The mobile phase was a mixture of acetonitrile (0.025% TFA) and H2O (0.05% TFA), and temperature was maintained at 50 °C. A gradient of 4% to 100% acetonitrile over 7 minutes was used during analytical analysis. Purity of final compounds was determined to be >95%, using a 5 μL injection with quantitation by AUC at 220 and 254 nM. High resolution mass spectra were obtained with an Agilent 6210 Time-of-Flight LC/MS with a 3.5 um Zorbax SB-C18 column (2.1 x 30 mm) (solvents are Water and ACN with 0.1% Formic Acid). A 3 minute gradient at 1 mL/min from 5% to 100% acetonitrile was used.
Ataluren (Translarna) Ataluren is a drug marketed under the trade name Translarna which was developed by PTC Therapeutics and approved by the European Union in May 2014 for the treatment of Duchenne’s muscular dystrophy (DMD) and potentially other genetic disorders.50
Ataluren renders ribosomes less sensitive to premature stop or ‘read-through’ codons, which are thought to be beneficial in diseases such as DMD and cystic fibrosis.51 Of the reported synthetic approaches to ataluren,52–55 the most likely process-scale approach consists of the sequence described in Scheme 7, which reportedly has been exemplified on kilogram scale.56
The sequence to construct ataluren, which was described by the authors at PTC Therapeutics, commenced with commercially available methyl 3-cyanobenzoate (38).56 This ester was exposed to hydroxylamine in aqueous tert-butanol and warmed gently until the reaction was deemed complete.
Then this mixture was treated with 2-fluorobenzoyl chloride dropwise and subsequently triethylamine dropwise. To minimize exotherm and undesired side products, careful control of the addition of reagents was achieved through slow dropwise addition of these liquid reagents.
Upon complete consumption of starting materials and formation of amidooxime 39, the aqueous reaction mixture was then heated to 85 C to facilitate 1,2,4-oxadiazole formation, resulting in the tricyclic ester 40 in excellent yield across the three steps.
Finally,saponification of ester 40 through the use of sodium hydroxide followed by acidic quench gave ataluren (V) in 96% over the two-step sequence.57
50. Welch, E. M.; Barton, E. R.; Zhuo, J.; Tomizawa, Y.; Friesen, W. J.; Trifillis, P.;Paushkin, S.; Patel, M.; Trotta, C. R.; Hwang, S.; Wilde, R. G.; Karp, G.; Takasugi,J.; Chen, G.; Jones, S.; Ren, H.; Moon, Y. C.; Corson, D.; Turpoff, A. A.; Campbell,J. A.; Conn, M. M.; Khan, A.; Almstead, N. G.; Hedrick, J.; Mollin, A.; Risher, N.;Weetall, M.; Yeh, S.; Branstrom, A. A.; Colacino, J. M.; Babiak, J.; Ju, W. D.;Hirawat, S.; Northcutt, V. J.; Miller, L. L.; Spatrick, P.; He, F.; Kawana, M.; Feng,H.; Jacobson, A.; Peltz, S. W.; Sweeney, H. L. Nature 2007, 447, 87.
51. Hirawat, S.; Welch, E. M.; Elfring, G. L.; Northcutt, V. J.; Paushkin, S.; Hwang,S.; Leonard, E. M.; Almstead, N. G.; Ju, W.; Peltz, S. W.; Miller, L. L. J. Clin.Pharmacol. 2007, 47, 430.
52Karp, G. M.; Hwang, S.; Chen, G.; Almstead, N. G. US Patent 2004204461A1,2004.
53. Andersen, T. L.; Caneschi, W.; Ayoub, A.; Lindhardt, A. T.; Couri, M. R. C.;Skrydstrup, T. Adv. Synth. Catal. 2014, 356, 3074.
54. Gupta, P. K.; Hussain, M. K.; Asad, M.; Kant, R.; Mahar, R.; Shukla, S. K.; Hajela,K. New J. Chem. 2014, 38, 3062.
55. Lentini, L.; Melfi, R.; Di Leonardo, A.; Spinello, A.; Barone, G.; Pace, A.; PalumboPiccionello, A.; Pibiri, I. Mol. Pharm. 2014, 11, 653.
56. Almstead, N. G.; Hwang, P. S.; Pines, S.; Moon, Y. -C.; Takasugi, J. J. WO Patent2008030570A1, 2008.
57. Almstead, N. G.; Chen, G.; Hirawat, S.; Hwang, S.; Karp, G. M.; Miller, L.; Moon,Y. C.; Ren, H.; Takasugi, J. J.; Welch, E. M.; Wilde, R. G. WO Patent2007117438A2, 2007.
Last week the newspaper NRC Handelsblad reported on a court case in which the parents of two young boys sued a pharmaceutical company over access to one of their developmental drugs. The drug in question wasAtaluren, the pharmaceutical companyPTC Therapeutics. The boys suffer from Duchenne muscular dystrophyand had taken part in a clinical trial. Whereas the results of this trial on the whole were inconclusive the boys did seriously benefit from the drug. Hardly any wonder the parents took action when the whole development program was canceled.
And the judge? He threw the case out arguing that doctors do not make the compound themselves and arguing that the compound is not commercially available. Are these arguments valid? and do the boys have options?
It is not that ataluren is a complex molecule. To judge from one of the patents, synthesis is straightforward starting from 2-cyanobenoic acid and 2-fluorobenzoyl chloride, both commercially available. The synthetic steps are methylation of 2-cyanobenoic acid (iodomethane), nitrile hydrolysis with hydroxylamine, esterification with the fluoro acid chloride using DIPEA, high-temperature dehydration to the oxadiazole and finally ester hydrolysis (NaOH).
Except for the fluorine atom in it the compound is unremarkable. If you have to believe the Internet many Chinese companies produce and sell it. Ataluren is also still in the running as a potential treatment for some other diseases. So if need be the compound will be around for some time to come.
CLIP
Ataluren [3-[5-(2-Fluorophenyl)-1,2,4-oxadiazol-3-yl]benzoic acid] is an orally available, small molecule compound that targets nonsense mutation. It is the first drug in its class and appears to allow cellular machinery to read through premature stop codons in mRNA, and thus enables the translation process to produce full-length, functional proteins.
Ataluren is developed and approved for the treatment of nonsense mutation Duchenne muscular dystrophy (nmDMD) by EU in July 2014 [1].
Ataluren: 2D and 3D Structure
Nonsense Mutations as Target for DMD
A single nucleotide change in the DNA sequence that introduces a premature stop codon is known as a nonsense mutation, a subset of a major class of premature termination codon (PTC) mutations. Nonsense mutations cause premature termination of translation resulting in the production of truncated polypeptides, which in turn halts the ribosomal translation process at an earlier site than normal, producing a truncated, non-functional protein [1].
Nonsense mutations are implicated in 5-70 % of individual cases of most inherited diseases, including Duchenne muscular dystrophy (DMD) and cystic fibrosis. Ataluren appears to allow cellular machinery to read through premature stop codons in mRNA, enabling the translation process to produce full length, functional proteins.
Ataluren Synthesis
New J Chem 2014, 38, 3062-3070: The text reports one pot synthesis of Ataluren with an overall yield of 40%. It also reports few interesting and potent derivatives too.
WO 2007117438A2: It appears to be the industrial process. The patent also reports various pharmaceutically relevant assay and their results wrt Ataluren. Identifications:
It is not that ataluren is a complex molecule. To judge from one of the patents, synthesis is straightforward starting from 2-cyanobenoic acid and 2-fluorobenzoyl chloride, both commercially available. The synthetic steps are methylation of 2-cyanobenoic acid (iodomethane), nitrile hydrolysis with hydroxylamine, esterification with the fluoro acid chloride using DIPEA, high-temperature dehydration to the oxadiazole and finally ester hydrolysis (NaOH).
Carcinogenicity bioassays in transgenic mice (26 weeks) and in rats (24 months):
● For Tg.rasH2 mouse: Ataluren did not increase the incidence of tumors up to the HDs in males (600 mg/kg/day) and in females (300 mg/kg/day). The non-neoplastic findings included endometrial hyperplasia and nephropathy in females.
● For rats: Urinary bladder tumors (benign urothelial cell papilloma [2 rats] and malignant urothelial cell carcinoma [1 rat]) were observed in 3/60 female rats dosed at 300 mg/kg/day. In addition, one case of malignant hibernoma was observed in 1/60 male rats at the dose of 300 mg/kg/day. The non-neoplastic toxicity consisted of a decrease of body weight.
Method ataluren according to Patent Document 2 is described in Example W02004091502A2 prepared.
Specific methods of preparation:
To a solution of 0.6 l of DMF was 44. 14g3- cyano acid 62.19 g of potassium carbonate was added, followed by stirring at room temperature for 30 minutes. 20 minutes To the suspension was added 28 ml of methyl iodide (450mmol), and the reaction mixture was stirred at room temperature for 4 hours. The reaction mixture was poured into 1.2 l of ice water, stirred for 30 minutes, the precipitate was filtered out thereof. The white cake was dissolved in 70 ml of methanol, and then reprecipitated in cold water. To give 79% yield of 3-cyano-benzoic acid methyl ester.
50 g of 3-cyano-benzoic acid methyl ester was dissolved in 500 ml of ethanol, to which was added 41 ml of 50% aqueous hydroxylamine (620mmol). 100 ° C and the reaction mixture was stirred for 1 hour, the solvent was removed under reduced pressure. So that the oily residue is dissolved in 100 ml of 20/80 ethanol / toluene, concentrated again. To give 61 g 3- (N- hydroxy amidino (carbamimidoyl)) – benzoic acid methyl ester.
60 g of 3- (N- hydroxy amidino (carbamimidoyl)) – benzoic acid methyl ester was dissolved in 200 ml of anhydrous tetrahydrofuran, followed by adding thereto 75 ml of diisopropylethylamine (434 mmol), and then 20 minutes this mixture was added 48.1 ml 2- fluorobenzoyl chloride (403mmol). The reaction mixture was stirred at room temperature for 1 hour. The precipitate was filtered off, the filtrate was concentrated under reduced pressure. The residue was dissolved in 400 ml of ethyl acetate, washed with 400 ml of water and then twice. The solvent was removed under reduced pressure, containing 60% ethyl acetate in hexane to give the desired product, generating 81 g 3- (N-2- amidino-fluorobenzoyl) – benzoate.
at 130 ° C with a Dean-Stark apparatus was dissolved in 500 ml of toluene was heated under reflux in 44 g of 3- (N-2- fluorobenzoyl) -1,2,3,4-_ benzoate 4 hours. 5 ° C and the reaction mixture was stirred for 18 hours. The white precipitate was filtered off, the filtrate was concentrated, recrystallized in toluene. To give 38 g of 3- [5- (2-fluorophenyl) – [1,2,4] oxadiazol-3-yl] – benzoic acid methyl ester.
33 g of 3- [5- (2-fluorophenyl) – [1,2,4] oxadiazol-3-yl] – benzoic acid methyl ester was dissolved in 400 ml of tetrahydrofuran, to which was added 100 ml of 1. 5M aqueous sodium hydroxide solution. At 100 ° C and the reaction mixture was heated at reflux for 2 hours. The solvent was removed under reduced pressure at 5 ° C the solution was stirred for 2 hours. The organic solvent was removed, washed with 50 mL of water. The aqueous solution was then acidified with hydrochloric acid to pH 1. The white precipitate was filtered off, the filter cake washed with cold water, then dried with a freeze dryer. To give 3.0 g of 3- [5- (2-fluorophenyl) – [1,2,4] oxadiazol-3-yl] benzoic acid. 1H-NMR (500MHz, d6-DMS0): 8. 31 (1H), 8 18 (2H), 8 08 (1H), 7 88 (2H), 7 51 (2H)….. Display: ataluren- Sample Preparation Example 1 prepared in Preparation Example 2 and TO2004091502A2 induced.
Each prepared in Example 2 (prepared according to known Form A)
Method [0084] A known polymorph according to Patent Document W02008039431A2 Example 5. 1. 1.1 prepared as described. Specifically: ataluren be prepared 1 100 mg Preparation Example, 60 ° C add 16.2 ml of isopropanol ultrasound clear solution, the solution by 2 square micron filter and the filtrate was kept covered with aluminum foil having a small hole. vial, 60 ° C and evaporated. The solid formed was isolated to give ataluren the A polymorph.
as needles.
its XRPD shown in Figure 1, the display ataluren polymorph A disclosed in Patent Document W02008039431A2 consistent.
SEE
European Journal of Organic Chemistry (2016), 2016(3), 438-442
Russian Chemical Bulletin (2015), 64(1), 142-145.
European Journal of Medicinal Chemistry (2015), 101, 236-244.
[l,2,4]oxadiazol-3-yl]-benzoic acid, which has the following chemical structure (I):
(I)
In particular, crystalline forms of 3-[5-(2-fluorophenyl)-[l,2,4]oxadiazol-3-yl]- benzoic acid are useful for the treatment, prevention or management of diseases ameliorated by modulation of premature translation termination or nonsense-mediated mRNA decay, as described in U.S. Patent No. 6,992,096 B2, issued January 31, 2006, which is incorporated herein by reference in its entirety. In addition, the present provides a crystalline form of 3-[5-(2-fluorophenyl)-[l,2,4]oxadiazol-3-yl]-benzoic acid which is substantially pure, i.e., its purity greater than about 90%.
Processes for the preparation of 3-[5-(2-fluorophenyl)-[l,2,4]oxadiazol-3-yl]- benzoic acid are described in U.S. Patent No. 6,992,096 B2, issued January 31, 2006, and U.S. patent application no. 1 1/899,813, filed September 9, 2007, both of which are incorporated by reference in their entirety.
The 3-cyano-benzoic acid methyl ester (105 kg) and t-butanol was added molten drying reactor.Under an inert atmosphere for about 2 hours 48 minutes, 50./.Aqueous hydroxylamine (43L, 47.4 kg) was added to a clear solution of 3-cyano benzoic acid methyl ester in a molten in t-butanol.The addition of a 50% aqueous solution of hydroxylamine period, the maximum temperature batch of about 43 ° C.50% aqueous solution of hydroxylamine addition rate of from about 9L / h when the changes start adding to about 30L / hr.To maintain the temperature of the batch by varying the reactor jacket set point.In particular, the set value is about 40.5 ° C, with the addition of a rate increase at the beginning join, change the setting to about 29.6 ° C.After about 40-45t stirred for about 4 hours, the reaction was deemed complete (i.e., less than about 0.5% ester).
The batch was transferred to a drying reactor, additional (chased through) approximately 10L molten tert-butanol.Jacket setpoint from about 33 when the batch was received when dried reactor.C is reduced to about 27 after the completion of the transfer.C.Batch crystallization was observed part, which does not adversely affect stirring.The batch was cooled to about 34.4 ° C, triethylamine (72.6 kg, IOOL) added to the reactor.The jacket temperature set value from about 20.4.C is increased to about 31.0 ° C, in order to maintain the batch temperature in the range of about 30-35t.With molten tert-butanol (IO L) was washed with a linear (line rinse) After the batch was added to the 2-fluorobenzoyl chloride (113.7 kg, 86.0L).Charge is added to the first third of the rate of about 25L / hr.In the meantime, the jacket inlet temperature was lowered to about 15 ° C, the batch temperature is maintained at about 34.6 ° C.In about 5.5 hours after the addition was complete.During the addition, the maximum temperature of the batch was about 38.8 ° C.Near the end of the addition, the addition rate slowed to about 11L / hr was added last 27 liters of 2-fluoro-benzoyl chloride.30-35.C After stirring for about 2 hours, that the reaction was complete (i.e., less than about 0.5% of methyl 3-amidinophenoxy).Then, after about 1 hour 42 minutes, the batch was heated to reflux temperature (about 82 ° C), and then stirred for about 18 hours.During the stirring, a number of product partially crystallized to form a slurry.The slurry was cooled to about 40.C thus sampled, during which complete crystallization occurs.The batch was then heated to reflux temperature and stirred for about 1 hour 50 minutes.Then, after about two hours, the batch was cooled to about 69 ° C, and after about four hours and 15 minutes, slowly added 630L of pure water, while maintaining the batch temperature at about 66-69 ° C.After about 3 hours 14 minutes, the slurry was cooled to about 22.4 ° C, and transferred to 2x200L ceramic filter, the ceramic filter equipped 25-30n polypropylene mesh filter cloth.In about 55 minutes after the completion of material from the container to the filter transfer.With 50n /.The tert-butanol solution (210L) was washed cake was washed for about 10 minutes so that the cleaning liquid can penetrate into each cake.Then, the cake was dried in a vacuum for about 5-10 minutes.The purified water as a second washing (158L / cake) applied to the filter cake to remove residual t-butanol and triethylammonium chloride salt.Dried in a vacuum for about 5 minutes, the solution was removed.In vacuo and then the cake was dried for about 2 hours, and then sampled using liquid chromatography.The filter cake was measured by liquid chromatography purity of about 99.6%.
The filter cake was dried in vacuo for about 8 hours 25 minutes later, the wet cake (207.4kg) is transferred to an air oven.At about 50-55.C, the oven dried in air for about 52 hours.The product was isolated in a total yield of about 89.9% (174.65kg), in the calculation of cost of materials sampling, you can adjust the overall yield of about 90.7%.
Batch 2
The 3-cyano-benzoic acid methyl ester (105 kg) and t-butanol was added molten drying reactor.Under an inert atmosphere for about 3 hours 29 minutes, 50% aqueous solution of hydroxylamine (47.85 kg) was added to the reactor.During the addition, the temperature is maintained at about 40-45 ° C.At about 40-45.C After stirring for about 3 hours 16 minutes, that the reaction was complete (i.e., less than about 0.5% ester).As for the drying reactor, the batch was transferred to one of the batch in.The batch was cooled
To about 34.4 ° C, and triethylamine (72.6 kg, 100 L).During about 45 minutes was added, while maintaining the batch temperature between about 30-35 ° C.During the addition, the jacket inlet temperature of from about 31.4.C increased to about 32.6.C.After the molten tert-butanol linear washed, was added to the batch 2- fluorobenzoyl chloride (l 13.7 kg, 86.0 L).After about 3 hours, 27 minutes, add the acid chloride.35.C under stirring for about 8 hours, that the reaction is not complete (i.e., more than about 0.5% residual 3-amidino-benzoyl ester).Then, 1.5% by weight of the original charge of triethylamine and 2-fluorobenzoyl chloride was added to the batch.Linear washed with tert-butanol (IO L) associated with each additional charge.During the addition of the acid chloride, no additional cooling.The batch was maintained at a temperature of about 30-35 ° C, the jacket inlet temperature range was maintained at about 30.3.C to about 33.0 ° C.After stirring for about 2 hours at 30-35t, that reaction was complete (i.e., less than 0.5% of methyl 3-amidinophenoxy).
After about 1 hour and 44 minutes, the batch was heated to reflux temperature (about 83 ° C), and stirred for about 18 hours.The same batch 1, during cooling the sample, the solid was completely crystallized.The batch was then heated to reflux temperature and stirred for about 1 hour and 2 minutes.Then, after about 2 hours and 20 minutes, the batch was cooled to about 69.2 ° C, and after about four hours and 30 minutes, slowly added 630 L of pure water, while the temperature of the batch was maintained at about 65.6-69.2 ° C.After about 3 hours and 30 minutes, the slurry was cooled to about 23.4 ° C, and, as for, the contents were transferred to one of the double batch of the ceramic filter.About 5 hours and 6 minutes, to complete the transfer of the material.With about 50% of t-butanol (2 volumes / cake) was washed filter cake was washed with 10 minutes to allow the cleaning liquid to penetrate into each cake, then dried in vacuo.About 1 hour and 40 minutes, the filter is completed.The purified water was added to a final wash the filter cake.The liquid was removed by drying under vacuum for about 10 minutes.In vacuo and then the cake was dried for about 2 hours and 5 minutes, and then sampled using liquid chromatography.The cake purity liquid chromatography were about 99.5% and 99.6%.After the cake was then dried in vacuo for about 2 hours and 5 minutes, the wet cake (191.5 kg) is transferred to an air oven.At about 50-55.C under dry in an air oven for about 48 hours.The product was isolated in a total yield of about 92.5% (179.7 kg).
Lot 3
The 3-cyano-benzoic acid methyl ester (52.5 kg) and molten tert-butanol (228 kg) added to the reaction vessel.The vessel was sealed, the batch temperature set of about 40-45 ° C, and the stirrer is started.Under an inert atmosphere, after 2 hours 40 minutes, 50% of the shoes amine solution (24 kg) was added to the reactor.During the addition, the temperature is maintained at about 40-45 ° C.In about 42.Under C, then further stirred for about 5 hours to complete the reaction.
The batch was cooled to 30-35 ° C, and after 15 minutes, was added triethylamine (36 kg).After about 2 hours 44 minutes, was added 2-fluorobenzoyl chloride (57 kg).During the addition, batch temperature was maintained at about 30-35 ° C.Under the 32t, the batch was stirred for 2 hours 10 minutes to complete the reaction.
After about 50 minutes, the batch was heated to reflux temperature (about 83-86 ° C), at about 8rc, stirred for about 18 hours.Then, over about two hours, the batch was cooled to about 65-70 ° C, and after about 6 hours 25 minutes, slowly added to purified water (315 L), while the batch temperature was maintained at about 65- 70 ° C.After about 2 hours and 15 minutes, the slurry was cooled to about 22 ° C, and the contents were transferred to a centrifuge filter (2 batches).About 1 hour and 40 minutes, the filter is completed.After about 20 minutes, with about 50% aqueous solution of tert-butyl alcohol (90 kg / cake), dried cake.The purified water (79 kg / cake) as the last added to the filter cake washed.At about 900 rpm drying the cake for about 1 hour and 5 minutes, then filled cylinder.Liquid chromatography wet cake (91.5 kg, LOD = 5% w / w) of a purity of about 99.75% area.
3- [5- (2-fluorophenyl) – [l, 2,4] oxadiazol-3-yl] – benzoic acid methyl ester (74.0kg) added to the reaction vessel, the vessel is sealed, evacuated and purification.Jacket set value of about 35.C, start the stirrer in the container.Molten tert-butanol (222 L, 3 volumes) and purified water (355 L, 4.8 vol) was added to the vessel.After the addition was added 25.1% w / w aqueous sodium hydroxide solution (43.5 kg, 1.1 molar equivalents), and with additional purified water (100L, 1.35 mol) was washed linear.During the addition, the batch temperature from about 39.0t reduced to about 38.8 ° C.After about 1 hour and 54 minutes, the batch temperature to about 63-67.C, and then, after about 30 minutes, which was adjusted to about 68-72.C.About 68-72t, stirring the mixture for about 3 hours.Then, after about five hours 11 minutes, the solution was cooled to about 40-45 ° C.Then, after the above process, after about three hours 33 minutes, the solution was then heated to about 68-72.C.
Jacket temperature of the reaction vessel was set to about 60 ° C, the stirrer started, and at about 70 ° C, a slightly positive pressure of nitrogen (1.5 to 5.6 psig), the heat transfer liquid through a micron filter .During the transfer, the product temperature is reduced to about 64.3 ° C, the transfer is completed in about 45 minutes.Was added to the purified water container (61 L, 0.82 vol) and the contents were heated to about 68-72.C.
The batch temperature was adjusted to about 69.4 ° C, and after about four hours and 18 minutes, with 13.9% w / w sulfuric acid (100.7 kg, 1.15 mol equiv.).During the addition, batch temperature was maintained at about 68.0-70.8 ° C.After the addition of the acid, with purified water (50 L, 0.68 vol) line wash at about 68-72 ° C, the stirring was continued for 31 minutes.
After about 4 hours and 10 minutes, the batch in a linear fashion from about 69.2t cooled to about 41.2 ° C.The stirrer Rosenmund filter / dryer was elevated to the highest position and jacket set value is set at about 40 ° C.The slurry was transferred to the two portions of the filter / drier.Applying a constant nitrogen pressure to the first portion (less than about 15 psig).During the transfer, a pressure of about 23.9 to about 28.8 psi, the transfer is complete in about 1 hour and 5 minutes.The second part of the slurry was transferred onto the filter cake, and the composite was stirred briefly to homogenize the batch.Use about 26.1 to about 29.1 psi nitrogen pressure filters the second part, after about three hours, squeeze the cake so that it does not contain liquid.With about 38-42 ° C hot tert-butanol solution (352 kg, 5 volumes) and about 65-70 ° C in 3x hot purified water (370 L, 5 volumes) and the filter 々.
Said filter / dryer jacket temperature was set to about 43 ° C, the product was dried under vacuum for about 26 hours while stirring periodically.Determination of purity of about 99.7%.The product was isolated in a total yield of about 74.4% (52.45 kg).
Batch 2
Was added to the reactor vessel 3- [5- (2-fluorophenyl) – [1,2,4] oxadiazol-3-yl] – benzoic acid methyl ester (47 kg, wet cake) and melt-hyun tert-butyl alcohol (111.4 kg).A sealed container, and the batch temperature was set at 30-40t, and start the stirrer.The purified water (51.6 kg) was added to the vessel.After the addition was added 3.47% w / w aqueous sodium hydroxide solution (202.4 kg).After about l hour, the batch temperature to about 67-73.C, then, at about 7 (under TC, stirred for about three hours.
Under a slight positive pressure of nitrogen, with a 1 micron polypropylene bag filter the batch, and then transferred to the new reactor.Was added to the vessel pure water (146 kg), and heating the batch to about 68-72.C.
After about four hours, the 10.7% aqueous hydrochloric acid was added to the batch.During the addition, batch temperature was maintained at about 68-72 ° C.PH was measured by using the batch pH of about 2.2, and then stirring was continued at about 7 (under TC about 1 hour.
After about two hours, the batch in a linear fashion from about 70.C is cooled to about 60 ° C.After about two hours, about 60.C of the batch in a linear fashion from about 6 (TC was cooled to about 40 ° C. In 40t, the batch was stirred for 2 hours, and the slurry was transferred to a centrifuge filter. After about 30 minutes, filtered completion . After about 30 minutes, with about 42Mw / w in t-butanol solution (165kg) cake was washed. The purified water (118kg, 4 (TC) as the last added to the filter cake was washed. The filter cake was dried at about 900rpm about 1 hour, then filled cylinder.
The wet cake was transferred to a paddle dryer (a double cone drier also suitable for this step), the jacket temperature was set to about 70.C.At about 70.C, the product was dried under vacuum for about 48 hours while stirring periodically.Determination of purity of about 99.8%.The product was isolated overall yield of about 74% (68.5 kg).
Lot 3
To the reaction vessel was added 3- [5- (2-fluorophenyl) – [1,2,4] oxadiazol-3-yl] – benzoic acid methyl ester (10 g) and t-butanol fused (128mL ).The batch temperature was set to 30-40 ° C, and the stirrer is started.After about 30 minutes, the aqueous sodium hydroxide solution 4.48% w / w of (32.5 g) was added to the vessel.The batch was maintained at a temperature of about 40-50 ° C.After about l hour, the batch temperature is raised to about 78-82 ° C, and then, at about 78-82t, and then stirred for about one hour.Under positive pressure of nitrogen, a polyethylene bag with a 5 micron filter the batch, and then transferred to a new reaction vessel.The batch was maintained at a temperature of about 78-82 ° C.
It was added to a new vessel 37% aqueous hydrochloric acid (4 mL) and tert-butanol molten (8 mL).The temperature was maintained at about 30-40.Under C, and stirring the mixture for about 30 minutes.
After about four hours, using a metering pump was added to the batch of hydrochloric acid in tert-butanol.After about SO-SO minutes before adding half filled.The stirrer speed is set at about 200rpm.After about 3.5 hours, add the remaining charge.The stirrer speed is set at about 100 rpm.During the addition, batch temperature was maintained at about 78-82 ° C.PH meter with a final batch pH was adjusted to about 1.2, at about 78-82t, then continue stirring for about l hour.After about one hour, the batch in a linear fashion from about 78-82.C is cooled to about 70 ° C.After about four hours, about 7 (TC batches in a linear fashion from 70.C cooled to about 50 ° C, and the stirrer speed was set at about 80 rpm. After about four hours, about 50 ° C Batch linearly cooled from 50 ° C to about 40t, and stirrer speed was set at approximately 60rpm. In 40.C, the batch was stirred for a further 4 hours.
The temperature of the filter is set to about 40-45 ° C.The slurry was transferred to the filter.After about one minute to complete filtration.After about two minutes, with tert-butanol (50 mL, 50.C) washing the filter cake.The pure water (IOO mLx2, 60.C) as the last wash was added to the cake.Under vacuum at about 60-70 ° C the cake was dried for about 12 hours, and then loaded into the container.
Determination of HPLC purity of about 99.9% of the area.The yield of isolated product was about 94% (9.0g).
The methyl 3-cyanophenyl Yue (7.35 g) and tert-butanol molten (100 mL) added to the reactor vessel.Sealed containers, the batch temperature was set to 60 ° C, and the stirrer is started.The suspension was stirred for 1 hour and then the batch temperature was set to 40.C.Under an inert atmosphere, after three hours, 50% aqueous solution of hydroxylamine (3.63 g) was added to the reactor.During the addition, batch temperature was maintained at 38-41 ° C.40.C After stirring for 18 hours, to complete the reaction.
The batch was cooled to 27 ° C, and after two minutes, triethylamine (5.56 g).After 3 hours, was added 2-fluorobenzoyl chloride (7.82 g).During the addition, batch temperature was maintained at 24-27 ° C.40.C, the batch was stirred for a further 4 hours.
After 30 minutes, the batch was heated to 79 ° C, and at about 79.C was stirred for 16 hours.After 3 hours, the white suspension was added to the water (IOO mL), while the batch temperature was maintained at 70 ° C.After 20 minutes, a 37% aqueous hydrochloric acid were added to the batch.PH was measured by using the batch pH of about 2.2, stirring was continued at about 70t for about 1 hour.
After three hours, the batch in a linear manner from 7 (TC cooled to 30 ° C, and the slurry is transferred to the filter. After 5 minutes, the filtering is done. After five minutes, with tert-butanol (50mL, 40 .C) filter cake was washed. The purified water (IOO mL, 60.C) is added to a final wash the filter cake. In 70.C of the filter cake was dried in a vacuum oven for 18 hours and then removed. Determination of purity approximately 98.68%. The total yield of isolated product of about 76% (10.8g).
PICS
A large-scale, multinational, phase 3 trial of the experimental drug ataluren has opened its first trial site, in Cincinnati, Ohio.
The trial is recruiting boys with Duchenne muscular dystrophy (DMD) or Becker muscular dystrophy (BMD) caused by anonsense mutation — also known as a premature stop codon — in the dystrophin gene. This type of mutation causes cells to stop synthesizing a protein before the process is complete, resulting in a short, nonfunctional protein. Nonsense mutations are believed to cause DMD or BMD in approximately 10 to 15 percent of boys with these disorders.
Ataluren — sometimes referred to as a stop codon read-through drug — has the potential to overcome the effects of a nonsense mutation and allow functional dystrophin — the muscle protein that’s missing in Duchenne MD and deficient in Becker MD — to be produced.
The orally delivered drug is being developed by PTC Therapeutics, a South Plainfield, N.J., biotechnology company, to whichMDA gave a $1.5 million grant in 2005.
Welch EM, Barton ER, Zhuo J, Tomizawa Y, Friesen WJ, Trifillis P, Paushkin S, Patel M, Trotta CR, Hwang S, Wilde RG, Karp G, Takasugi J, Chen G, Jones S, Ren H, Moon YC, Corson D, Turpoff AA, Campbell JA, Conn MM, Khan A, Almstead NG, Hedrick J, Mollin A, Risher N, Weetall M, Yeh S, Branstrom AA, Colacino JM, Babiak J, Ju WD, Hirawat S, Northcutt VJ, Miller LL, Spatrick P, He F, Kawana M, Feng H, Jacobson A, Peltz SW, Sweeney HL (May 2007). “PTC124 targets genetic disorders caused by nonsense mutations”. Nature447 (7140): 87–91. Bibcode:2007Natur.447…87W.doi:10.1038/nature05756. PMID17450125.
Hirawat S, Welch EM, Elfring GL, Northcutt VJ, Paushkin S, Hwang S, Leonard EM, Almstead NG, Ju W, Peltz SW, Miller LL (Apr 2007). “Safety, tolerability, and pharmacokinetics of PTC124, a nonaminoglycoside nonsense mutation suppressor, following single- and multiple-dose administration to healthy male and female adult volunteers”. Journal of clinical pharmacology47 (4): 430–444.doi:10.1177/0091270006297140. PMID17389552.
Nature. 2007 May 3;447(7140):87-91.
Proc Natl Acad Sci U S A. 2008 Feb 12;105(6):2064-9.
Neuromuscul Disord. 2015 Jan;25(1):5-13.
Wilschanski, M. (2013). “Novel therapeutic approaches for cystic fibrosis”. Discovery Medicine15 (81): 127–133. PMID23449115.
Wilschanski, M.; Miller, L. L.; Shoseyov, D.; Blau, H.; Rivlin, J.; Aviram, M.; Cohen, M.; Armoni, S.; Yaakov, Y.; Pugatsch, T.; Cohen-Cymberknoh, M.; Miller, N. L.; Reha, A.; Northcutt, V. J.; Hirawat, S.; Donnelly, K.; Elfring, G. L.; Ajayi, T.; Kerem, E. (2011). “Chronic ataluren (PTC124) treatment of nonsense mutation cystic fibrosis”. European Respiratory Journal38 (1): 59–69. doi:10.1183/09031936.00120910. PMID21233271.Sermet-Gaudelus, I.; Boeck, K. D.; Casimir, G. J.; Vermeulen, F.; Leal, T.; Mogenet, A.; Roussel, D.; Fritsch, J.; Hanssens, L.; Hirawat, S.; Miller, N. L.; Constantine, S.; Reha, A.; Ajayi, T.; Elfring, G. L.; Miller, L. L. (November 2010). “Ataluren (PTC124) induces cystic fibrosis transmembrane conductance regulator protein expression and activity in children with nonsense mutation cystic fibrosis”. American Journal of Respiratory and Critical Care Medicine182 (10): 1262–1272. doi:10.1164/rccm.201001-0137OC. PMID20622033.
References:
1. Ryan, N. J. Ataluren: first global approval. Drugs 2014, 74(14), 1709-14. (FMO only)
2. Gupta, P. K.; et. al. A metal-free tandem approach to prepare structurally diverse N-heterocycles: synthesis of 1,2,4-oxadiazoles and pyrimidinones. New J Chem 2014, 38, 3062-3070 (FMO only)
3. Almstead, N. G.; et. al. Methods for the production of functional protein from dna having a nonsense mutation and the treatment of disorders associated therewith. WO2007117438A2
SHENZHEN CHIPSCREEN BIOSCIENCES LTD. [CN/CN]; Research Institute of Tsinghua University, Suite C301, P.O. Box 28, High-Tech Industrial Park Nanshan District, Shenzhen, Guangdong 518057
As described for Example 2 according to the patent ZL03139760.3 obtained chidamide poor purity (about 95%).LC / MS analysis results shown in Figure 1, show that the product contains N- (2- amino-5-fluorophenyl) -4- (N- (3- pyridin-acryloyl group of 4.7% of the structure shown in formula II) aminomethyl) benzamide.1H NMR analysis of the results shown in Figure 2, show that the product contains 1.80% of tetrahydrofuran, far beyond the technical requirements for people with drug registration International Conference on Harmonization (ICH, International Conference of Harmonizition) provided 0.072% residual solvent limits.Therefore, the solid
Is NOT approved for the treatment of pancreatic cancer.
Chidamide drug administration and clinical milestone
November 2005: China declared IND
November 2006: eligible for Phase I clinical documents of approval
November 2006: completion of the International Patent Licensing, China entered the international fray original new drug development
May 2008: completed Phase I clinical, showing international mechanism similar drugs have the potential to become the best
February 2009: eligible lymphoma indications II / III of this document
March 2009: Start of the Phase II clinical trial for the NDA to ①CTCL goal of clinical trials and ②PTCL
March 2009: IND by the FDA application is eligible to start Phase I clinical in the United States
July 2009: eligible for non-small cell lung cancer, breast cancer and prostate cancer clinical documents of approval
December 2010: of PTCL by a conventional phase II directly into Phase II clinical trial registered drug trial center and by recognition
March 2011: combination chemotherapy for non-small cell lung cancer clinical trials enter phase Ib
September 2012: of PTCL indication test deadline
December 2012: of PTCL clinical summary will be held
January 2013: Chidamide declare China NDA
December 2014: the State Food and Drug Administration (CFDA) approved the listing
Chidamide overview, location and clinical significance
Chidamide (Chidamide, love spectrum sand ® / Epidaza®) Shenzhen microchip biotechnology limited liability company developed a new subtype selective histone having a chemical structure and is eligible for a global patent licensing deacetylase inhibitor, belong to the new mechanisms of epigenetic regulation new class of targeted anticancer drugs, has now completed with relapsed or refractory peripheral T-cell lymphoma clinical trial study registered indications, was in March 2013 to the SFDA reporting new drug certificate (NDA) and the marketing authorization (MAA). While a number of Chinese Cancer clinical trials undertaken Chidamide is also China’s first approved by the US FDA clinical studies in the United States of Chinese chemical original new drug trials in the United States Phase I has been completed. Chidamide has won the national “Eleventh Five-Year” 863 major projects (project number: 2006AA020603) and the national “Eleventh Five-Year”, “significant Drug Discovery” science and technology and other major projects funded project (project number: 2009ZX09401-003), was chosen the Ministry of Science and one of the “Eleventh five-Year” major national scientific and technological achievements.
Relapsed or refractory peripheral T-cell lymphoma (PTCL) is Chidamide first approvedclinical indications, PTCL belongs to the category of rare diseases, the lack of standard drug currently recommended clinical treatment, conventional chemotherapy response rate is low, recur, 5-year overall survival rate was about 25%. The world’s first PTCL treatment Folotyn (intravenous drug use) is eligible for FDA clearance to market in 2009, the second drugs Istodax (intravenous drug use) approved by the FDA in 2011. Add a new drug information for these drugs is very expensive, and were listed in China. Chidamide album clinical trial results showed that the primary endpoint of objective response rate was 28%, reaching the intended target research and development; sustained remission rate of 24% three months; drug safety was significantly better than the international similar drugs, and oral medication.
Chidamide is a completely independent intellectual property rights China originator of innovative medicines, has been multi-national patent. In China, for patients with relapsed or refractory PTCL to carry out effective drug treatment is urgent clinical need, Chidamide expected to bring new treatment options for patients with PTCL, prolong survival and improve quality of life of patients.
In China, for the effective treatment of patients with relapsed or refractory PTCL has undertaken urgent clinical need
Chidamide is a completely independent intellectual property rights China originator of innovative medicines
Chidamide (Chidamide) has been multi-national invention patents
In October 2006, the US HUYA biological microchip company formally signed the International Patent Chidamide licensing and international clinical cooperative development agreement; the United States in the ongoing Phase I clinical
Chidamide (Epidaza), a class I HDAC inhibitor, was discovered and developed by ChipScreen and approved by the CFDA in December 2014 for the treatment of recurrent of refractory peripheral T-cell lymphoma. Chidamide, also known as CS055 and HBI- 8000, is an orally bioavailable benzamide type inhibitor of HDAC isoenzymes class I , as well as class IIb 10, with potential antineoplastic activity. It selectively binds to and inhibits HDAC, leading to an increase in acetylation levels of histone protein H3.
Chidamide, the English called Chidamide, by the Shenzhen-core biotechnology limited liability company independent design and synthesis of a novel anti-cancer drugs with new chemical structures and global intellectual property, and its chemical name N- (2-amino-_4_ fluorophenyl) -4_ (N- (3- topiramate Li acryloyl) aminomethyl) benzamide, its chemical structure of the structural formula I
The patent ZL03139760.3 and said US7,244,751, Chidamide have histone deacetylase inhibitory activity can be used to treat the differentiation and proliferation-related diseases such as cancer and psoriasis, especially for leukemia and solid tumors with excellent results.
Patent No. ZL03139760.3 and US7,244,751 discloses a method for preparing chidamide, but did not specify whether the resulting product is a crystalline material, nor did the presence or absence of the compound polymorphism.In the above patent, the activity of the compound for evaluation is not conducted in a solid state and, therefore, does not disclose any description about characteristics of the crystal.
Chipscreen grabs CFDA approval for chidamide
Posted on
Chipscreen BioSciences announced that the CFDA had approved chidamide for the treatment of relapsed or refractory peripheral T-cell lymphoma (PTCL) in December 2014. The drug and Hengrui’s apatinib were the only two NCEs launched by domestic drug makers last year.
Chidamide (CS055/HBI-8000) is a HDAC1/2/3/10 inhibitor derived from entinostat (MS-27-275)[1] which was first discoved by Mitsui Pharmaceuticals in 1999. Chipscreen holds worldwide IP rights to chidamide (patents: WO2004071400, WO2014082354).
Syndax Pharmaceuticals (NASDAQ: SNDX) is testing entinostat in breast cancer and NSCLC in pivotal trials. The FDA granted Breakthrough Therapy Designation to entinostat for advanced breast cancer in 2013. Eddingpharm in-licensed China rights to entinostat from Syndax in September 2013.
Chipscreen disclosed positive results from Phase II study of chidamide in relapsed or refractory PTCL at 2013 ASCO Annual Meeting[2]. Out of 79 evaluable patients in the trial, 23 patients (29.1%) had confirmed responses (8 CR, 3 CRu, and 12 PR). The most common grade 3/4 AEs were thrombocytopenia (24%), leucocytopenia (13%), neutropenia(10%).
The FDA has approved three HDAC inhibitors, known as Zolinza (vorinostat), Istodax (romidepsin) and Beleodaq (belinostat), for the treatment of PTCL. Celgene priced Istodax at $12000-18000/month and reported annual sales of $54 million in 2013. The efficacy and safety profile of chidamide compares favorably with romidepsin.
Although a dozen of companies are developing generic vorinostat and romidepsin, no chemical 3.1 NDA has been submitted to the CFDA so far. Chipscreen will be the only domestic maker of HDAC inhibitor in the coming two years. Moreover, the company is testing chidamide in NSCLC and breast cancer in early clinical studies.
CLIP
Chiamide synthesis: US7244751B2
Procedure:
Step a: To a suspension of 0.33 g (2.01 mmol) of N,N’-carbonyldiimidazole in tetrahydrofunan (10 ml) is added drop-wise a solution of 0.30 g (2.01 mmol) of 3-pyridineacrylic acid at 0 °C. Then, the mixture is stirred at room temperature for 3 hours and added drop-wise to a separately prepared 2.0 ml (2.00 mmol) of 1N aqueous sodium hydroxide solution including 0.30 g (2.00 mmol) of 4-aminomethylbenzoic acid, followed by stirring at room temperature for 8 hours. The reaction mixture is evaporated under vacuum. To the residue is added a saturated solution of sodium chloride (2 ml), then the mixture is neutralized with concentrated hydrochloric acid to pH 5. The deposited white solid is collected by filtration, washed with ice-water, and then dried to give 4-[N-(Pyridin-3-ylacryloyl)aminomethyl]benzoic acid (0.46 g, 82%). HRMS calcd for C16H14N2O3: 282.2988. Found: 282.2990. MA calcd for: C16H14N2O3: C, 68.07%; H, 5.00%; N, 9.92%. Found: C, 68.21%; H, 5.03%; N, 9.90%.
Step b: To a suspension of 0.29 g (1.78 mmol) of N,N’-carbonyldiimidazole in tetrahydrofunan (15 ml) is added 0.50 g (1.78 mmol) of 4-[N-(Pyridin-3-ylacryloyl)aminomethyl]benzoic acid, followed by stirring at 45 °C. for 1 hour. After cooling, the reaction mixture is added to a separately prepared tetrahydrofiman (10 ml) solution including 0.28 g (2.22 mmol) of 4-fluoro-1,2-phenylenediamine and 0.20 g (1.78 mmol) of trifluoroacetic acid at room temperature. After reaction at room temperature for 24 hours, the deposited white solid is collected by filtration, washed with tetrahydrofunan, and then dried to give N-(2-amino-4-fluorophenyl)-4-[N-(Pyridin-3-ylacryloyl)aminomethyl]benzamide (0.40 g, 57%). 1H NMR (300 MHz, DMSO-d6): dppm: 4.49 (2H, d), 4.84 (2H, br.s), 6.60 (1H, t), 6.80 (2H, m),696 (1H, t), 7.18 (1H, d), 7.42 (2H, d), 7.52 (1H, d), 7.95 (2H, d), 8.02 (1H, d), 8.56 (1H, d), 8.72 (1H, br. t), 8.78 (1H, s), 9.60 (1H, br.s). IR (KBr) cm1: 3310, 1655, 1631, 1524, 1305, 750. HRMS calcd for C22H19N4O2F: 390.4170. Found: 390.4172. MA calcd for C22H19N4O2F: C, 67.68%; H, 4.40%; N, 14.35%. Found: C, 67.52%; H, 4.38%; N, 14.42%.
Preparation of 4-[N-(Pyridin-3-ylacryloyl)aminomethyl]benzoic acid
To a suspension of 0.33 g (2.01 mmol) of N,N′-carbonyldiimidazole in tetrahydrofunan (10 ml) is added drop-wise a solution of 0.30 g (2.01 mmol) of 3-pyridineacrylic acid at 0° C. Then, the mixture is stirred at room temperature for 3 hours and added drop-wise to a separately prepared 2.0 ml (2.00 mmol) of 1N aqueous sodium hydroxide solution including 0.30 g (2.00 mmol) of 4-aminomethylbenzoic acid, followed by stirring at room temperature for 8 hours. The reaction mixture is evaporated under vacuum. To the residue is added a saturated solution of sodium chloride (2 ml), then the mixture is neutralized with concentrated hydrochloric acid to pH 5. The deposited white solid is collected by filtration, washed with ice-water, and then dried to give the title compound (0.46 g, 82%). HRMS calcd for C16H14N2O3: 282.2988. Found: 282.2990. MA calcd for: C16H14N2O3: C, 68.07%; H, 5.00%; N, 9.92%. Found: C, 68.21%; H, 5.03%; N, 9.90%.EXAMPLE 2
Preparation of N-(2-amino-4-fluorophenyl)-4-[N-(Pyridn-3-ylacryloyl)aminomethyl]benzamide
To a suspension of 0.29 g (1.78 mmol) of N,N′-carbonyldiimidazole in tetrahydrofunan (15 ml) is added 0.50 g (1.78 mmol) of 4-[N-(Pyridn-3-ylacryloyl)aminomethyl]benzoic acid, followed by stirring at 45° C. for 1 hour. After cooling, the reaction mixture is added to a separately prepared tetrahydrofiman (10 ml) solution including 0.28 g (2.22 mmol) of 4-fluoro-1,2-phenylenediamine and 0.20 g (1.78 mmol) of trifluoroacetic acid at room temperature. After reaction at room temperature for 24 hours, the deposited white solid is collected by filtration, washed with tetrahydrofunan, and then dried to give the title compound (0.40 g, 57%). 1H NMR (300 MHz, DMSO-d6): δppm: 4.49 (2H, d), 4.84 (2H, br.s), 6.60 (1H, t), 6.80 (2H, m),696 (1H, t), 7.18 (1H, d), 7.42 (2H, d), 7.52 (1H, d), 7.95 (2H, d), 8.02 (1H, d), 8.56 (1H, d), 8.72 (1H, br. t), 8.78 (1H, s), 9.60 (1H, br.s). IR (KBr) cm1: 3310, 1655, 1631, 1524, 1305, 750. HRMS calcd for C22H19N4O2F: 390.4170. Found: 390.4172. MA calcd for C22H19N4O2F: C, 67.68%; H, 4.40%; N, 14.35%. Found: C, 67.52%; H, 4.38%; N, 14.42%.EXAMPLE 3
Preparation of 4-[N-cinnamoylaminomethyl]benzoic acid
To a suspension of 0.33 g (2.01 mmol) of N,N′-carbonyldiimidazole in tetrahydrofunan (10 ml) is added drop-wise a solution of 0.30 g (2.01 mmol) of cinnamic acid at 0° C. Then, the mixture is stirred at room temperature for 3 hours and added drop-wise to a separately prepared 2.0 ml (2.00 mmol) of 1N aqueous sodium hydroxide solution including 0.30 g (2.00 mmol) of 4-aminomethylbenzoic acid, followed by stirring at room temperature for 8 hours. The reaction mixture is evaporated under vacuum. To the residue is added a saturated solution of sodium chloride (2 ml), then the mixture is neutralized with concentrated hydrochloric acid to pH 7. The deposited white solid is collected by filtration, washed with ice-water, and then dried to give the title compound (0.51 g, 91%). HRMS calcd for C17H15NO3: 281.3242. Found: 281.3240. MA calcd for C17H15NO3: C, 72.58%; H, 5.38%; N, 4.98. Found: C, 72.42%; H, 5.37%; N, 4.98%.
EXAMPLE 4
Preparation of N-(2-amino-4-fluorophenyl)-4-[N-cinnamoylaminomethyl]benzamide
To a suspension of 0.29 g (1.78 mmol) of N,N′-carbonyldiimidazole in tetrahydrofunan (15 ml) is added 0.50 g (1.78 mmol) of 4-[N-cinnamoylaminomethyl]benzoic acid, followed by stirring at 45° C. for 1 hour. After cooling, the reaction mixture is added to a separately prepared tetrahydrofunan (10 ml) solution including 0.28 g (2.22 mmol) of 4-fluoro-1,2-phenylenediamine and 0.20 g (1.78 mmol) of trifluoroacetic acid at room temperature. After reaction at room temperature for 16 hours, the deposited white solid is collected by filtration, washed with tetrahydrofunan, and then dried to give the title compound (0.45 g, 64%). 1H NMR (300 MHz, DMSO-d6): δppm: 4.42 (2H, d), 4.92 (2H, br.s), 6.62 (1H, t), 6.78 (2H, m), 7.01 (1H, t), 7.32 (5H, m), 7.54 (5H, m), 8.76 (1H, br.t), 9.58 (1H, br.s). IR (KBr) cm−1: 3306, 1618, 1517, 1308, 745. HRMS calcd for C23H20N3O2F: 389.4292. Found: 389.4294. MA calcd for C23H20N3O2F: C, 70.94%; H, 5.18%; N, 10.79%. Found: C, 70.72%; H, 5.18%; N, 10.88%.
Chidamide (Epidaza), a class I HDAC inhibitor, was discovered and developed by ChipScreen and approved by the CFDA in December 2014 for the treatment of recurrent of refractory peripheral T-cell lymphoma. Chidamide, also known as CS055 and HBI- 8000, is an orally bioavailable benzamide type inhibitor of HDAC isoenzymes class I 1–3, as well as class IIb 10, with potential antineoplastic activity. It selectively binds to and inhibits HDAC, leading to an increase in acetylation levels of histone protein H3.74
This agent also inhibits the expression of signaling kinases in the PI3K/ Akt and MAPK/Ras pathways and may result in cell cycle arrest and the induction of tumor cell apoptosis.75
Currently, phases I and II clinical trials are underway for the treatment of non-small cell lung cancer and for the treatment of breast cancer, respectively.76 The scalable synthetic approach to chidamide very closely follows the discovery route,77–79 and is described in Scheme 10. The sequence began with the condensation of commercial nicotinaldehyde (52) and malonic acid (53) in a mixture of pyridine and piperidine. Next, activation of acid 54 with N,N0-carbonyldiimidazole (CDI) and subsequent reaction with 4-aminomethyl benzoic acid (55) under basic conditions afforded amide 56 in 82% yield.
Finally, activation of 56 with CDI prior to treatment with 4-fluorobenzene- 1,2-diamine (57) and subsequent treatment with TFA and THF yielded chidamide (VIII) in 38% overall yield from 52. However, no publication reported that mono-N-Boc-protected bis-aniline was used to approach Chidamide.
74. Ning, Z. Q.; Li, Z. B.; Newman, M. J.; Shan, S.; Wang, X. H.; Pan, D. S.; Zhang, J.;
Dong, M.; Du, X.; Lu, X. P. Cancer Chemother. Pharmacol. 2012, 69, 901.
75. Liu, L.; Chen, B.; Qin, S.; Li, S.; He, X.; Qiu, S.; Zhao, W.; Zhao, H. Biochem.
Biophys. Res. Commun. 2010, 392, 190.
76. Gong, K.; Xie, J.; Yi, H.; Li, W. Bio. Chem. J. 2012, 443, 735.
77. Lu, X. P.; Li, Z. B.; Xie, A. H.; Shi, L. M.; Li, B. Y.; Ning, Z. Q.; Shan, S.; Deng, T.;
Hu, W. M. US Patent 2004224991A1, 2004.
78. Lu, X. P.; Li, Z. B.; Xie, A. H.; Shi, L. M.; Li, B. Y.; Ning, Z. Q.; Shan, S.; Deng, T.;
Hu, W. M. CN Patent 1513839A, 2003.
79. Yin, Z. H.; Wu, Z. W.; Lan, Y. K.; Liao, C. Z.; Shan, S.; Li, Z. L.; Ning, Z. Q.; Lu, X.
P.; Li, Z. B. Chin. J. New Drugs 2004, 13, 536.
Example 2. Preparation of
N-(2-amino-5-fluorophenyl)-4-[N-(Pyridn-3-ylacryloyl)aminomethyl]benzamide
To a suspension of 0.29 g (1.78 mmol) of N, N’-carbonyldiimidazole in tetrahydrofunan (15 ml) is added 0.50 g (1.78 mmol) of 4-[N-(Pyridn-3-ylacryloyl)aminomethyl]benzoic acid, followed by stirring at 45°C for 1 hour. After cooling, the reaction mixture is added to a separately prepared tetrahydrofunan (10 ml) solution including 0.28 g (2.22 mmol) of 4-fluoro-1,2-phenylenediamine and 0.20 g (1.78 mmol) of trifluoroacetic acid at room temperature. After reaction at room temperature for 24 hours, the deposited white solid is collected by filtration, washed with tetrahydrofunan, and then dried to give the title compound (0.40 g, 57%). 1H NMR (300 MHz, DMSO-d6): δppm: 4.49 (2H, d), 4.84 (2H, br.s), 6.60 (IH, t), 6.80 (2H, m), 6.96 (IH, t), 7.18 (IH, d), 7.42 (2H, d), 7.52 (IH, d), 7.95 (2H, d), 8.02 (IH, d), 8.56 (IH, d), 8.72 (IH, br. t), 8.78 (IH, s), 9.60 (IH, br.s). IR (KBr) cm“1: 3310, 1655, 1631, 1524, 1305, 750. HRMS calcd for C22Hι9N4O2F: 390.4170. Found: 390.4172. MA calcd for C22Hι9N4O2F: C, 67.68%; H, 4.40%; N, 14.35. Found: C, 67.52%; H, 4.38%; N, 14.42%.
Photo taken on May 22, 2015 shows a box of Chidamide in Shenzhen, south China’s Guangdong Province. Chidamide is the world’s first oral HDAC inhibitor …
A New Cancer Drug, Made in China
After 14 years, Shenzhen biotech’s medicine is one of the few locally developed from start to finish
Xian-Ping Lu left his research job at a drug maker in the U.S. to co-found a biotech company in his native China.PHOTO: SHENZHEN CHIPSCREEN BIOSCIENCES
HONG KONG— Xian-Ping Lu left his job as director of research at drug maker Galderma R&D in Princeton, N.J., to co-found a biotech company to develop new medicines in his native China.
It took more than 14 years but the bet could be paying off. In February, Shenzhen Chipscreen Biosciences’ first therapy, a medication for a rare type of lymph-node cancer, hit the market in China.
The willingness of veterans like Dr. Lu and others to leave multinational drug companies for Chinese startups reflects a growing optimism in the industry here. The goal, encouraged by the government, is to move the Chinese drug industry beyond generic medicines and drugs based on ones developed in the West.
Chipscreen’s drug, called chidamide, or Epidaza, was developed from start to finish in China. The medicine is the first of its kind approved for sale in China, and just the fourth in a new class globally. Dr. Lu estimates the research cost of chidamide was about $70 million, or about one-tenth what it would have cost to develop in the U.S.
“They are a good example of the potential for innovation in China,” said Angus Cole, director at Monitor Deloitte and pharmaceuticals and biotechnology lead in China.
China’s spending on pharmaceuticals is expected to top $107 billion in 2015, up from $26 billion in 2007, according to Deloitte China. It will become the world’s second-largest drug market, after the U.S., by 2020, according to an analysis published last year in the Journal of Pharmaceutical Policy and Practice.
China has on-the-ground infrastructure labs, a critical mass of leading scientists and interested investors, according to Franck Le Deu, head of consultancy McKinsey & Co.’s pharmaceuticals and medical-products practice in China. “There’re all the elements for the recipe for potential in China,” he said.
But there are obstacles to an industry where companies want big payoffs for a decade or more of work and tremendous costs it takes to develop a drug.
While the protection of intellectual property has improved, China’s cumbersome rules for drug approval and a government effort to cut health-care costs, particularly spending on drugs, could hurt the Chinese drug companies’ efforts, said Mr. Cole of Deloitte.
“Will you start to see success? Of course you will,” said Mr. Cole. However, “I’ve yet to see convincing or compelling evidence that it’s imminent.”
To date, many of the Chinese companies that are flourishing in the life sciences are contract research organizations that help carry out clinical trials, as well as providers of related services.
Some companies, like Shanghai-based Hua Medicine, are buying the rights to develop new compounds in China from multinational drug companies, what some experts consider more akin to an intermediate step to innovation.
Late last year, Hua Medicine completed an early-stage human clinical trial of a diabetes drug in China and in March filed an application to the Food and Drug Administration to develop it in the U.S. as well. The company has raised $45 million in venture funding to date.
Li Chen, who left an 18-year career at Roche Holding AG as head of research and development in China to help start Hua Medicine, said the company’s goal is to “create a game-changer of drug discovery.”
At Chipscreen Biosciences, Dr. Lu and his co-founders set up the company in 2001 in Shenzhen, a city that was quickly growing into a technology and research hub, just over the border from Hong Kong. They created a lab of 10 scientists to use a new analytic technique known as “chemical genomics” to examine the relationships between molecular structures of the existing and failed drugs, how they act on different targets in the body and what genes were being activated or repressed. Now they have more than 60 scientists.
By better predicting how chemicals would act on the body before entering human testing, they hoped they would be more likely get a drug to market.
“How can a small company compete with a multinational?” said Dr. Lu. “The only thing we can compete with is the scientific brain.”
The biggest challenges for the company have been financing and the Chinese regulatory system, said Dr. Lu. The company has raised a total of 300 million yuan ($48 million) over five rounds of venture funding, said Dr. Lu. Chipscreen also receives grant money from the Chinese government.
The company filed its application for approval of chidamide to the Chinese Food and Drug Administration, or CFDA, in early 2013. It had to wait nearly two years for approval, receiving the OK only in December.
Chidamide now is on the market in China for 26,500 yuan ($4,275) a month, a price far lower than patients in the U.S. pay for some of the newest cancer medicines but much more than the typical Chinese patient pays for drugs. Dr. Lu said the price reflects a balance between affordability for patients and return for shareholders. Some investors wanted to price the drug higher.
PAPER
Discovery of an orally active subtype-selective HDAC inhibitor, chidamide, as an epigenetic modulator for cancer treatment
Tumorigenesis is maintained through a complex interplay of multiple cellular biological processes and is regulated to some extent by epigenetic control of gene expression. Targeting one signaling pathway or biological function in cancer treatment often results in compensatory modulation of others, such as off-target drivers of cell survival. As a result, overall survival of cancer patients is still far from satisfactory. Epigenetic-modulating agents can concurrently target multiple aberrant or compensatory signaling pathways found in cancer cells. However, existing epigenetic-modulating agents in cancer treatment have not yet fully translated into survival benefits beyond hematological tumors. In this article, we present a historical rationale for use of chidamide (CS055/Epidaza), an orally active and subtype-selective histone deacetylase (HDAC) inhibitor of the benzamide chemical class. This compound was discovered and successfully developed as mono-therapy for relapsed and refractory peripheral T cell lymphoma (PTCL) in China. We discuss the evidence supporting chidamide as a durable epigenetic modulator that allows cellular reprogramming with little cytotoxicity in cancer treatments.
CLIPS
Chinese scientists develop world’s 1st oral HDAC inhibitor
Lu Xianping works in a lab at Shenzhen Chipscreen Biosciences Ltd. in Shenzhen, south China’s Guangdong Province, May 20, 2015. Lu Xianping, together with other four returned overseas scientists, spent 14 years to develop Chidamide, the world’s first oral HDAC inhibitor, which was given regulatory approval in January. (Xinhua/Mao Siqian)
GNT Biotech and Medicals Corporation Licenses Novel Cancer Molecule from Shenzhen Chipscreen Biosciences Ltd.
PR Newswire
SHENZHEN, China, Oct. 10, 2013
SHENZHEN, China, Oct. 10, 2013 /PRNewswire/ — GNT Biotech and Medicals Corporation announces the grant of an exclusive license from Shenzhen Chipscreen Biosciences Ltd.for the development and commercialization of Chidamide in Taiwan. Chidamide, an oral, selective histone deacetylase (HDAC) inhibitor, is currently being evaluated in Phase II trials by Chipscreen Biosciences in Peripheral T-Cell Lymphoma (PTCL), Cutaneous T-Cell Lymphoma (CTCL) and Non-Small Cell Lung Cancer patients (NSCLC). GNTbm will develop and commercialize Chidamide primarily in PTCL, NSCLC and will also retain the rights to develop and commercialize Chidamide in other oncology indications in Taiwan.
About Chidamide
Chidamide is a selective HDAC inhibitor against subtype 1, 2, 3 and 10, and being studied in multiple clinical trials as a single agent or in combination with chemotherapeutic agents for the treatment of various hematological and solid cancers. Its anticancer effects are thought to be mediated through epigenetic modulation via multiple mechanisms of action, including the inhibition of cell proliferation and induction of apoptosis in blood derived cells, inhibition of epithelial to mesenchymal transition (EMT, a process that is highly relevant to tumor cell metastasis and drug resistance), induction of tumor specific antigen and antigen-specific T cell cytotoxicity, enhancement of NK cell anti-tumor activity, induction of cancer stem cell differentiation, and resensitization of tumor cells that have become resistant to anticancer agents such as platinums, taxanes and topoisomerase II inhibitors. Chidamide has demonstrated clinical efficacy in pivotal phase II trials on Cutaneous T-Cell Lymphoma (CTCL) and Peripheral T-Cell Lymphoma (PTCL) conducted in China, and is currently undergoing phase II trial in NSCLC together with first line PC therapeutic treatment. Due to its superior pharmacokinetic properties and selectivity, Chidamide may offer better clinical profile over the other HDAC inhibitors currently under development or being marketed.
About GNTbm
GNTbm is a subsidiary of GNT Inc, a Taiwanese company focused on the manufacture of nano-scale metallic particles for food and medical purposes. Founded in 1992 by a team of electronic professionals, GNT has successfully developed the innovative technology of physical metal miniaturization based on the patent of MBE (Molecular Beam Epitaxy). Further information about GNT Inc is available at www.gnt.com.tw.
GNTbm was established in August 2013, and housed in the Nankang Biotech Incubation Center, (NBIC), in Nankang, Taipei. Lead by Dr. Chia-Nan Chenalong with an experienced team of scientists, GNTbm will explore development and commercialization of novel drug delivery systems, Innovative biomedical and diagnostic tools based on gold nanoparticles.
About Shenzhen Chipscreen Biosciences Ltd.
Chipscreen is a leading integrated biotech company in China specialized in discovery and development of novel small molecule pharmaceuticals. The company has utilized its proprietary chemical genomics-based discovery platform to successfully develop a portfolio of clinical and preclinical stage programs in a number of therapeutic areas. Chipscreen’s business strategy is to generate differentiated drug candidates across multiple therapeutic areas. Drug candidates are either developed by Chipscreen or co-developed and commercialized in a partnership at the research, preclinical and clinical stages. The company was established as Sino-foreign joint venture in 2001. Further details about Chipscreen Bioscience is available atwww.chipscreen.com.
Qiao, Z (2013-04-26). “Chidamide, a novel histone deacetylase inhibitor, synergistically enhances gemcitabine cytotoxicity in pancreatic cancer cells.”. Biochem Biophys Res Commun.434 (1): 95–101. doi:10.1016/j.bbrc.2013.03.059. PMID23541946.
References:
1. Ning, Z. Q.; et. al. Chidamide (CS055/HBI-8000): a new histone deacetylase inhibitor of the benzamide class with antitumor activity and the ability to enhance immune cell-mediated tumor cell cytotoxicity. Cancer Chemother Pharmacol2012, 69(4), 901-909. (activity)
2. Gong, K.; et. al. CS055 (Chidamide/HBI-8000), a novel histone deacetylase inhibitor, induces G1 arrest, ROS-dependent apoptosis and differentiation in human leukaemia cells. Biochem J 2012, 443(3), 735-746. (activity)
3. Hu, W.; et. al. N-(2-amino-5-fluorophenyl)-4-[N-(Pyridin-3-ylacryloyl) aminomethyl ]benzamide or other derivatives for treating cancer and psoriasis. US7244751B2
4. Lu, X.; et. al. Crystal form of chidamide, preparation method and use thereof. WO2014082354A1
5. Yin, Z.-H.; et. al. Synthesis of chidamide,a new histone deacetylase (HDAC) inhibitor. Chin J New Drugs 2004, 13(6), 536-538. (starts with basic raw materials)
Zhongguo Xinyao Zazhi (2004), 13(6), 536-538.
/////////Chidamide, Epidaza, CS055, HBI-8000, orally active subtype-selective HDAC inhibitor, epigenetic modulator, cancer treatment
SELECTBIO is delighted to announce its 4th International Drug Discovery India 2016 Conference and Exhibition. This conference will be held in Le Méridien Bangalore Hotel, Bengaluru on September 29-30, 2016. The theme of the conference is “Deriving Best Out of Chemistry, Biotechnology and Natural Products“.
The event aims to expand knowledge by providing insights into the latest developments and innovations in the field of drug discovery, medicinal chemistry, natural products and chemical biology. Every year this event bring together about 200 drug discovery scientists together at a platform to discuss and share drug discovery related research and facilitates collaborations amongst scientists from across the globe.
This meeting will be co-located with our 2nd International Conference Antibodies and Antibody Drug Conjugates. Registered delegates will have unrestricted access to all co-located meetings ensuring a comprehensive learning and sharing experience as well as being financially beneficial for attendees.
Running alongside the Drug Discovery India 2016 conference will be an Exhibition covering the latest technological advances within these fields. We look forward to welcoming you at the Drug Discovery India 2016 Conference and Exhibition and hope that the two days will be both informative and enjoyable.
Who Should Attend
Drug Discovery Scientists, Medicinal Chemists, Biotechnologists & Researchers from Pharmaceutical Industry R&D and Academic institutions working in the area of New Drug Discovery Research, Discovery and Development of New Chemical entities, Biomolecular Screening Technologies, Drug Target Identification, Structure-based and Target-based, Drug Design, Protein-Protein Interactions, Drug Repurposing, Orphan Drugs, Chemical Biology, Stem Cell, Epigenetics as well as Natural Products.
A phase III clinical trial for rheumatoid arthritis was halted in Feb 2013.[3] In September 2014, a second phase III trial focussing on treating systemic lupus erythematosus, was terminated early as the study failed to meet its primary endpoint.[4]
Continuous production is a flow production method used to manufacture, produce, or process materials without interruption. Continuous production is called a continuous process or a continuous flow process because the materials, either dry bulk or fluids that are being processed are continuously in motion, undergoing chemical reactions or subject to mechanical or heat treatment. Continuous processing is contrasted with batch production.
Continuous usually means operating 24 hours per day, seven days per week with infrequent maintenance shutdowns, such as semi-annual or annual. Some chemical plants can operate for more than one or two years without a shutdown. Blast furnaces can run four to ten years without stopping.[1]
Production workers in continuous production commonly work in rotating shifts.
Processes are operated continuously for practical as well as economic reasons. Most of these industries are very capital intensive and the management is therefore very concerned about lost operating time.
Shutting down and starting up many continuous processes typically results in off quality product that must be reprocessed or disposed of. Many tanks, vessels and pipes cannot be left full of materials because of unwanted chemical reactions, settling of suspended materials or crystallization or hardening of materials. Also, cycling temperatures and pressures from starting up and shutting down certain processes (line kilns, boilers, blast furnaces, pressure vessels, etc.) may cause metal fatigue or other wear from pressure or thermal cycling.
In the more complex operations there are sequential shut down and start up procedures that must be carefully followed in order to protect personnel and equipment. Typically a start up or shut down will take several hours.
Continuous processes use process control to automate and control operational variables such as flow rates, tank levels, pressures, temperatures and machine speeds.[2]
Semi-continuous processes
Many processes such as assembly lines and light manufacturing that can be easily shut down and restarted are today considered semi-continuous. These can be operated for one or two shifts if necessary.
History
The oldest continuous flow processes is the blast furnace for producing pig iron. The blast furnace is intermittently charged with ore, fuel and flux and intermittently tapped for molten pig iron and slag; however, the chemical reaction of reducing the iron and silicon and later oxidizing the silicon is continuous.
Semi-continuous processes, such as machine manufacturing of cigarettes, were called “continuous” when they appeared.
Many truly continuous processes of today were originally batch operations.
The Fourdrinier paper machine, patented in 1799, was one of the earliest of the industrial revolution era continuous manufacturing processes. It produced a continuous web of paper that was formed, pressed, dried and reeled up in a roll. Previously paper had been made in individual sheets.
Another early continuous processes was Oliver Evans‘es flour mill (ca. 1785), which was fully automated.
Early chemical production and oil refining was done in batches until process control was sufficiently developed to allow remote control and automation for continuous processing. Processes began to operate continuously during the 19th century. By the early 20th century continuous processes were common.
Shut-downs
In addition to performing maintenance, shut downs are also when process modifications are performed. These include installing new equipment in the main process flow or tying-in or making provisions to tie-in sub-processes or equipment that can be installed while the process is operating.
Shut-downs of complicated processes may take weeks or months of planning. Typically a series of meetings takes place for co-ordination and planning. These typically involve the various departments such as maintenance, power, engineering, safety and operating units.
All work is done according to a carefully sequenced schedule that incorporates the various trades involved, such as pipe-fitters, millwrights, mechanics, laborers, etc., and the necessary equipment (cranes, mobile equipment, air compressors, welding machines, scaffolding, etc.) and all supplies (spare parts, steel, pipe, wiring, nuts and bolts) and provisions for power in case power will also be off as part of the outage. Often one or more outside contractors perform some of the work, especially if new equipment is installed.
Safety
Safety meetings are typically held before and during shutdowns. Other safety measures include providing adequate ventilation to hot areas or areas where oxygen may become depleted or toxic gases may be present and checking vessels and other enclosed areas for adequate levels of oxygen and insure absence of toxic or explosive gases. Any machines that are going to be worked on must be electrically disconnected, usually through the motor starter, so that it cannot operate. It is common practice to put a padlock on the motor starter, which can only be unlocked by the person or persons who is or are endangered by performing the work. Other disconnect means include removing couplings between the motor and the equipment or by using mechanical means to keep the equipment from moving. Valves on pipes connected to vessels that workers will enter are chained and locked closed, unless some other means is taken to insure that nothing will come through the pipes.
Continuous processor (equipment)
Continuous Production can be supplemented using a Continuous Processor. Continuous Processors are designed to mix viscous products on a continuous basis by utilizing a combination of mixing and conveying action. The Paddles within the mixing chamber (barrel) are mounted on two co-rotating shafts that are responsible for mixing the material. The barrels and paddles are contoured in such a way that the paddles create a self-wiping action between themselves minimizing buildup of product except for the normal operating clearances of the moving parts. Barrels may also be heated or cooled to optimize the mixing cycle. Unlike an extruder, the Continuous Processor void volume mixing area is consistent the entire length of the barrel ensuring better mixing and little to no pressure build up. The Continuous Processor works by metering powders, granules, liquids, etc. into the mixing chamber of the machine. Several variables allow the Continuous Processor to be versatile for a wide variety of mixing operations:[3]
Barrel Temperature
Agitator speed
Fed rate, accuracy of feed
Retention time (function of feed rate and volume of product within mixing chamber)
Continuous Processors are used in the following processes:
Compounding
Mixing
Kneading
Shearing
Crystallizing
Encapsulating
The Continuous Processor has an unlimited material mixing capabilities but, it has proven its ability to mix:
Plastics
Adhesives
Pigments
Composites
Candy
Gum
Paste
Toners
Peanut Butter
Waste Products
EXAMPLE…………….
In the development of a new route to bendamustine hydrochloride, the API in Treanda, the key benzimidazole intermediate 5 was generated via catalytic heterogeneous hydrogenation of an aromatic nitro compound using a batch reactor. Because of safety concerns and a site limitation on hydrogenation at scale, a continuous flow hydrogenation for the reaction was investigated at lab scale using the commercially available H-Cube. The process was then scaled successfully, generating kilogram quantities on the H-Cube Midi. This flow process eliminated the safety concerns about the use of hydrogen gas and pyrophoric catalysts and also showed 1200-fold increase in space–time yield versus the batch processing.
Improved Continuous Flow Processing: Benzimidazole Ring Formation via Catalytic Hydrogenation of an Aromatic Nitro Compound
Org. Process Res. Dev., 2014, 18 (11), pp 1427–1433
Correia et al. have published a three-step flow synthesis of rac-Effavirenz. This short synthetic route begins with cryogenic trifluoroacetylation of 1,4-dichlorobenzene. After quench and removal of morpholine using silica gel, this intermediate could either be isolated, or the product stream could be used directly in the next alkynylation step. Nucleophilic addition of lithium cyclopropylacetylide to the trifluoroacetate gave the propargyl alcohol intermediate in 90% yield in under 2 min residence time. This reaction was temperature-sensitive, and low temperatures were required to minimize decomposition. Again silica gel proved effective in the quench of the reaction. However, residual alkyne and other byproducts were difficult to remove. Thus, isolation of this intermediate was performed to minimize the impact of impurities on the final copper catalyzed cyanate installation/cyclization step to afford Effavirenz. Optimization of this step in batch mode for both copper source and ligand identified Cu(NO3)2 and CyDMEDA in a 1:4 molar ratio (20 mol % and 80 mol %, respectively) produced the product in 60% yield. Adaptation of this procedure to flow conditions resulted in poor conversion due to slow in situ reduction of the Cu(II) to Cu(I). Thus, a packed bed reactor of NaOCN and Cu(0) was used. Under these conditions, the ligand and catalyst loading could be reduced without compromising yield. Due to solubility limitations of Cu(NO3)2, Cu(OTf)2 was used with CyDMEDA in 1:2 molar ratio (5 mol % and 10 mol % loading, respectively). Under these optimized conditions, rac-Effavirenz was obtained in 62% isolated yield in reaction time of 1 h. This three-step process provides 45% overall yield of rac-Effavirenz and represents the shortest synthesis of this HIV drug reported to date
1 T. J. Connolly; A. W.-Y Chan; Z. Ding; M. R. Ghosh; X. Shi; J. Ren, E. Hansen; R. Farr; M. MacEwan; A. Alimardanov; et al, PCT Int. Appl. WO 2009012201 A2 20090122, 2009.
2 (a) Z. Dai, X. Long, B. Luo, A. Kulesza, J. Reichwagen, Y. Guo, (Lonza Ltd), PCT Int. Appl. WO2012097510, 2012; (b) D. D. Christ; J. A. Markwalder; J. M. Fortunak; S. S. Ko; A. E. Mutlib; R. L. Parsons; M. Patel; S. P. Seitz, PCT Int. Appl. WO 9814436 A1 19980409, 1998 (c) C. A. Correia; D. T. McQuade; P. H. Seeberger, Adv. Synth. Catal. 2013, 355, 3517−3521.
Wang et al. developed a flow process that uses metal catalyzed hydrogenation of NAB (2-nitro-2′-hydroxy-5′-methylazobenzene) to BTA (2-(2′-hydroxy-5′-methylphenyl)benzotriazole), a commonly used ultraviolet absorber. The major challenge in this process was to optimize the reduction of the diazo functionality over the nitro group and control formation of over reduction side products. The initial screen of metals adsorbed onto a γ-Al2O3 support indicated Pd to be superior to the other metals and also confirmed that catalyst preparation plays an important role in selectivity. To better understand the characteristics of the supported metal catalyst systems, the best performing were analyzed by TEM, XRD, H2-TPR, and N2 adsorption–desorption. Finally, solvents and bases were screened ultimately arriving at the optimized conditions using toluene, 2 equiv n-butylamine over 1% Pd/Al2O3, which provided 90% yield BTA in process with 98% conversion. The process can run over 200 h without a decrease in performance
( ACS Sustainable Chem. Eng. 2015, 3,1890−1896)
.
The synthesis of 2-(2′-hydroxy-5′-methylphenyl)benzotriazole from 2-nitro-2′-hydroxy-5′-methylazobenzene over Pd/γ-Al2O3 in a fixed-bed reactor was investigated. Pd/γ-Al2O3 catalysts were prepared by two methods and characterized by XRD, TEM, H2-TPR, and N2 adsorption–desorption. Employed in the above reaction, the palladium catalyst impregnated in hydrochloric acid exhibited much better catalytic performance than that impregnated in ammonia–water, which was possibly attributed to the better dispersion of palladium crystals on γ-Al2O3. This result demonstrated that the preparation process of the catalyst was very important. Furthermore, the reaction parameters were optimized. Under the optimized conditions (toluene, NAB/triethylamine molar ratio 1:2, 60 °C, 2.5 MPa hydrogen pressure, 0.23 h–1 liquid hourly space velocity), about 90% yield of 2-(2′-hydroxy-5′-methylphenyl)benzotriazole was obtained. Finally, the time on stream performance of the catalyst was evaluated, and the reaction could proceed effectively over 200 h without deactivation of the catalyst.
Construction of 2-(2′-Hydroxy-5′-methylphenyl)benzotriazole over Pd/γ-Al2O3 by a Continuous Process
Continuous Flow-Processing of Organometallic Reagents Using an Advanced Peristaltic Pumping System and the Telescoped Flow Synthesis of (E/Z)-Tamoxifen
A new enabling technology for the pumping of organometallic reagents such as n-butyllithium, Grignard reagents, and DIBAL-H is reported, which utilises a newly developed, chemically resistant, peristaltic pumping system. Several representative examples of its use in common transformations using these reagents, including metal–halogen exchange, addition, addition–elimination, conjugate addition, and partial reduction, are reported along with examples of telescoping of the anionic reaction products. This platform allows for truly continuous pumping of these highly reactive substances (and examples are demonstrated over periods of several hours) to generate multigram quantities of products. This work culminates in an approach to the telescoped synthesis of (E/Z)-tamoxifen using continuous-flow organometallic reagent-mediated transformations.
Multi-step Continuous Flow Pyrazole Synthesis via a Metal-free Amine-redox Process
A versatile multi-step continuous flow synthesis for the preparation of substituted pyrazoles is presented.
The automated synthesis utilises a metal-free ascorbic acid mediated reduction of diazonium salts prepared from aniline starting materials followed by hydrolysis of the intermediate hydazide and cyclo-condensation with various 1,3-dicarbonyl equivalents to afford good yields of isolated functionalised pyrazole products.
The synthesis of the COX-2 selective NSAID was demonstrated using this approach.
Synthesis of a Precursor to Sacubitril Using Enabling Technologies
Continuous flow methodologyhas been used to enhance several steps in the synthesis of a precursor to Sacubitril.
In particular, a key carboethoxyallylation benefited from a reducedprocessing time and improved reproducibility, the latter attributable toavoiding the use of a slurry as in the batch procedure. Moreover, in batchexothermic formation of the organozinc species resulted in the formation ofside products, whereas this could be avoided in flow because heat dissipationfrom a narrow packed column of zinc was more efficient
RAFT RAFT (Reversible Addition Fragmentation chain Transfer), a type of controlled radical polymerization, was invented by CSIRO in 1998 but developed in partnership with DuPont over a long term collaboration. Conventional polymerisation is fast but gives a wide distribution of polymer chain lengths. (known as a high polydispersity index ). RAFT is more versatile than other living polymerization techniques, such as atom transfer radical polymerization (ATRP) or nitroxide-mediated polymerization (NMP), it not only leads to polymers with a low polydispersity index and a predetermined molecular weight, but it permits the creation of complex architectures, such as linear block copolymers, comblike, star, brush polymers and dendrimers. Monomers capable of polymerizing by RAFT include styrenes, acrylates, acrylamides, and many vinyl monomers. CSIRO is the owner of the RAFT patents and is actively commercialising the technology. There are 12 licences in force and CSIRO is pursuing interest in a number of fields including human health, agriculture, animal health and personal care. RAFT is the dominant polymerization technique for the creation of polymer-protein or polymer-drug conjugates, permitting (for example) the combination of a polymer exhibiting high solubility with a drug molecule with poor solubility.. Though RAFT can be carried out in batch, it also lends itself to continuous flow processing, as this processing method offers an easy and reproducible scale-up route of the oxygen sensitive RAFT process. The possibility to effectively exclude oxygen using continuous flow reactors in combination with inline degassing methods offers advantages over batch processing at scales beyond the laboratory environment. Challenges associated with the high viscosity of the polymer product solution can be controlled using pressuriseable continuous flow reactor systems. http://www.csiro.au/products/RAFT.html
Examples………..
Cyclohexaneperoxycarboxylic acid (6, has been developed as a safe, inexpensive oxidant, with demonstrated utility in a Baeyer−Villiger rearrangement.34 Solutions of cyclohexanecarboxylic acid in hexane and 50% aqueous H2O2 were continuously added to 45% H2SO4 at 50−70 °C and slightly reduced pressure. The byproduct H2O was removed azeotropically, and the residence time in the reactor was 3 h. Processing was adjusted to maintain a concentration of 6 at 17−19%, below the detonable level, and the product was kept as a stable solution in hexane. These operations enhanced the safety margin in preparing 6.
Scheme . Generation of cyclohexaneperoxycarboxylic acid
Examples………..
The conversion of a batch process to continuous (flow) operation has been investigated. The manufacture of 4,d-erythronolactone at kilogram scale was used as an example. Fully continuousprocessing was found to be impracticable with the available plant because of the difficulty in carrying out a multiphase isolation step continuously, so hybrid batch–continuous options were explored. It was found that very little additional laboratory or process safety work other than that required for the batch process was required to develop the hybrid process. A hybrid process was chosen because of the difficulty caused by the precipitation of solid byproduct during the isolation stage. While the project was a technical success, the performance benefits of the hybrid process over the batch were not seen as commercially significant for this system.
Multikilogram Synthesis of 4-d-Erythronolactone via Batch andContinuousProcessing
Org. Process Res. Dev., 2012, 16 (5), pp 1003–1012
Benett, Stuart (1986). A History of Control Engineering 1800-1930. Institution of Engineering and Technology. ISBN978-0-86341-047-5.
Ziegler, Gregory R.; Aguilar, Carlos A. (2003). “Residence Time Distribution in a Co-rotating, Twin-screw Continuous Mixer by the Step Change Method”. Journal of Food Engineering(Elsevier) 59 (2-3): 1–7.
Sources and further reading
R H Perry, C H Chilton, C W Green (Ed), Perry’s Chemical Engineers’ Handbook (7th Ed), McGraw-Hill (1997), ISBN 978-0-07-049841-9
Major industries typically each have one or more trade magazines that constantly feature articles about plant operations, new equipment and processes and operating and maintenance tips. Trade magazines are one of the best ways to keep informed of state of the art developments.
The present invention provides a process for the preparation and isolation of crystalline miglustat without the use of a column chromatography or ion exchange purification. The crystalline miglustat has a high purity and a melting point of 128 °C and an endothermic peak is 133 °C.
Process for preparing and isolating crystalline form of miglustat with a high purity is claimed. Represents a first PCT filing from the inventors on miglustat. Actelion, under license from Oxford GlycoSciences (OGS; then Celltech, now UCB), which licensed the compound from GD Searle & Co, has developed and launched miglustat.
Product patent WO9426714, will expire in the US in 2018.
Kharatkar is affiliated with Sterling Biotech, Bhatt is affiliated with Intas and Kevat is affiliated with Orchid Chemicals & Pharmaceuticals.
Navinta, LLC in Ewing, N.J. is a technology driven Pharmaceutical Company that focuses on novel routes of synthesis of new and existing drug molecules, complex pharmaceutical ingredients, novel formulations of liquid dosage form, novel oral dosage form, novel injectable dosage form and implantable drug delivery devices. Navinta has currently at least fifteen (15) patents granted or pending with the United States Patent and Trademark Office.
Miglustat is a potent inhibitor of glycosyltransferase. It is primarily used in the treatment of Gaucher’s disease. Miglustat is chemically known as N-butyl-1,5-dideoxy-1,5-imino-D-glucitol of formula (I) and is sometimes referred as N-butyl-1-deoxynojirimycin. Miglustat is a white to off-white crystalline solid with a melting point of 125-126° C. Its empirical formula is C10H21NO4 and has a molecular weight of 219.28 g/mol.
Miglustat belongs to the class of azasugars or iminosugars. Ever since the discovery of iminosugars in the 1960s, iminosugars have been subject of extensive studies in both the organic chemistry and biochemistry fields. Iminosugars are polyhydroxylated alkaloids, which may be described as monosaccharide analogues with nitrogen replacing oxygen in the ring. A well-known member of this extensive family of compounds is 1-deoxynojirimycin of formula (II).
1-Deoxynojirimycin was initially synthesized in a laboratory. Subsequently, 1-deoxynojirimycin was isolated from natural sources, such as from leaves of mulberry trees and certain species of bacteria. 1-Deoxynojirimycin was shown to be an enzyme inhibitor.
Further research on 1-deoxynojirimycin analogs revealed that N-alkylated derivatives of 1-deoxynojirimycin exhibited greater biological activity than 1-deoxynojirimycin. Among them, N-butyl-1-deoxynojirimycin or miglustat of formula (I), was identified as a very potent inhibitor of glycosyltransferase. Miglustat was later approved by the FDA for human use.
Preparation of azasugars has been a very active area of research for a long time. A seminal synthesis of the compounds of formulas (I) and (II) by double reductive aminations of 5-keto-D-glucose was developed by Baxter and Reitz (J. Org. Chem. 1994, 59, 3175). This method was later refined by Matos and Lopes (Synthesis 1999, 571), in which tetra-O-benzyl-glucose was used as a starting material. Synthesis of miglustat can be traced back to 1977, when chemists from Bayer reported a synthesis of miglustat from 1-deoxynojirimycin and patented in U.S. Pat. No. 4,639,436. Other variations of this general scheme have also appeared in patents and non-patent literature, for example, U.S. Pat. No. 8,802,155 and U.S. Application Publication No. 2014/0243369.
A major drawback of those protocols is that all of them require the use of ion-exchange resins for purification of miglustat. In those protocols, an aqueous solution of miglustat obtained after running an ion-exchange column was concentrated to isolate miglustat. Due to the presence of four hydroxyl groups and a tertiary amine moiety in its chemical structure, miglustat is extremely hydrophilic. Thus, isolation of miglustat from an aqueous solution is quite challenging. In particular, it was very difficult to remove diastereomers and inorganic impurities formed during the reactions from miglustat by those protocols. Sometimes a second chromatographic purification was required to separate these impurities from miglustat. As a result, the yields of miglustat were generally low. The requirement to use a column purification (e.g. ion exchange column, flash column chromatography) further limits the scale of miglustat that could be prepared.
Scheme 1 is a synthetic scheme of miglustat in accordance with one embodiment of the invention:
As depicted in scheme 1, the method of preparing miglustat may include the steps of: (1) providing or synthesizing a compound of formula (V); (2) conducting a reductive amination to provide a compound of formula (VI); (3) performing a hydrogenation reaction; and (4) isolating a free base miglustat.
The starting material, 2,3,4,6-tetra-O-benzyl-1-deoxynojirimycin hydrochloride of formula (V) may be prepared by following the methods described in Organic Process Research and Development, 2008, 12, 414-423.
Example 1
Synthesis of 2, 3, 4, 6-tetra-O-benzyl-N-butyl-1-deoxynojirimycin hydrochloride of Formula (VI)
To a solution of 2, 3, 4, 6-tetra-O-benzyl-1-deoxynojirimycin hydrochloride (V) (prepared as in Organic Process Research & Development, 2008, 12, 414-423) (45 g, 0.08 mol) in 1575 mL of methanol, n-butyraldehyde (21.6 g, 0.24 mol) and sodium cyanoborohydride (25.2 g, 0.4 mol) were added and stirred. The reaction was maintained under stirring at a temperature from about 25.degree. C. to about 30.degree. C. After the completion of the reaction, the reaction was quenched by adding 765 ml of 10% HCl in methanol, while keeping the temperature between 25.degree. C. to 30.degree. C. The reaction mass was cooled to 0.degree. C. to 5.degree. C. and the resulting precipitate solids were filtered. The filtrate was treated with aqueous HCl and the solid formed was filtered, suspended in 1 N HCl, stirred for 1 hour and filtered. The collected solid was washed with diisopropylether and dried under vacuum to furnish 46.2 g of compound (IV) (46.2 g, 0.075 mol, 94% yield) of high chemical purity based on HPLC analysis (>99.0%).
Example 2
Synthesis of Miglustat Hydrochloride of Formula (III)
A solution of 2, 3, 4, 6-tetra-O-benzyl-N-butyl-1-deoxynojirimycin hydrochloride (VI) (100 g, 0.16 mol) in methanol (1000 mL), 10% HCl solution in methanol (100 mL), and 10% Pd/C (50% wet) (10 g) were mixed and stirred under hydrogen atmosphere at a temperature of about 25.degree. C. to about 30.degree. C. until completion of the reaction. The reaction mass was filtered and the solvent was removed from the filtrate by rotary evaporation. Ethyl acetate (1000 mL) was added to the residue from the rotary evaporation to precipitate the solid. The solid was filtered and dried to isolate Miglustat hydrochloride (III) (42 g, 0.16 mol, 100% yield) of >99.5% purity as measured by HPLC analysis. The DSC thermogram of this product is provided as FIG. 3, and the FTIR spectrum of this product is provided as FIG. 4.
Example 3
Synthesis of Miglustat of Formula (I)
Miglustat hydrochloride (III) (42 g, 0.16 mol) obtained from Example 2 was dissolved in 420 mL of methanol and DBU (1,8-diazabicycloundec-7-ene) (34.1 mL) was added. The reaction mass was warmed slightly and stirred for about 2 hours. The reaction was concentrated by removal of methanol. Dichloromethane (900 mL) was added to the residue. The resulting solid was filtered and dried to obtain crystalline miglustat (I) (27 g, 0.12 mol, 75% yield) of >99.5% purity as measured by HPLC analysis. The melting point of the crystalline miglustat (I) is 128.degree. C. The DSC thermogram and FTIR spectrum of the product are provided as FIG. 1 and FIG. 2, respectively. The crystalline miglustat (I) contained <0.05% of the 5R isomer (IV) as measured by HPLC.
Mitsubishi reported that tenatoprazole was still in Phase I clinical trials in 2007[4]:27 and again in 2012.[3]:17
Tenatoprazole has an imidazopyridine ring in place of the benzimidazole moiety found in other proton pump inhibitors, and has a half-life about seven times longer than other PPIs.[5]
Tenatoprazole is a novel imidazopyridine derivative and has an imidazopyridine ring in place of the benzimidazole moiety found in other proton pump inhibitors. It is activated more slowly than other proton pump inhibitor, but its inhibition is resistant to reversal.Tenatoprazole has an extended plasma half-life in comparison with those of all other proton pump inhibitors; this makes it more potent in the treatment of nocturnal acid breakthrough than esomeprazole, one of the most popular proton pump inhibitors.
Tenatoprazole belongs to the class of covalent proton pump inhibitors (PPIs), which is converted to the active sulfenamide or sulfenic acid by acid in the secretory canaliculus of the stimulated parietal cell of the stomach.This active species binds to luminally accessible cysteines of the gastric H+,K+-ATPase, resulting in disulfide formation and acid secretion inhibition.Tenatoprazole binds at the catalytic subunit of the gastric acid pump with a stoichiometry of 2.6 nmol mg−1 of the enzyme in vitro. In vivo, maximum binding of tenatoprazole was 2.9 nmol mg−1of the enzyme at 2 h after intravenous (IV) administration.
Tenatoprazole, or (+)-5-methoxy-2-{[(4-methoxy-3,5-dimethyl-2-pyridyl) methyl] sulfinyl} imidazo-[4,5-b] pyridine, is described in Patent No. EP 254,588. It belongs to the group of drugs considered as proton pump inhibitors, which inhibit the secretion of gastric acid and are useful in the treatment of gastric and duodenal ulcers. It can also be used to treat gastro-oesophageal reflux, digestive bleeding and dyspepsia, because of its relatively long elimination half-life, as described in the application for French patent No. FR 02. 13113.
The first known derivative of this series of proton pump inhibitors was omeprazole, described in Patent No. EP 001,529, which is endowed with properties which inhibit the secretion of gastric acid and is widely employed as an anti-ulcerative in human therapeutics.
In addition to omeprazole, other proton pump inhibitors are well known, and particular mention can be made of rabeprazole, pantoprazole and lansoprazole, which all exhibit structural analogy and lansoprazole, which all exhibit structural analogy and belong to the group of pyridinyl methyl sulfinyl benzimidazoles. These compounds are sulfoxides presenting with asymmetry at the level of the sulphur atom, and therefore generally take the form of a racemic mixture of two enantiomers.
Like omeprazole and other sulfoxide with an analogue structure, tenatoprazole has an asymmetric structure and may therefore be present in the form of a racemic mixture or of its enantiomers. Thus it may exist in the form of its two enantiomers with R and S configurations, or (+) or (−), respectively.
Recent studies have shown that, unlike all the other proton pump inhibitors such as, for example, omeprazole or lansoprazole, and unexpectedly, tenatoprazole is endowed with a markedly prolonged duration of action, resulting from a plasma half-life which is about seven times longer. Thus the clinical data collected have shown that tenatoprazole enables a degree of symptom relief and healing of gastric lesions which is superior to that achieved by other drugs belonging to the same therapeutic category of proton pump inhibitors, which thus allows its effective use in the treatment of atypical and oesophageal symptoms of gastro-oesophageal reflux, digestive bleeding and dyspepsia, as indicated above.
Studies performed by the application have made it possible to show that the two enantiomers contribute differently to the properties of tenatoprazole, and that the two enantiomers, (+) and (−) exhibit significantly different pharmacokinetic properties. Thus it is possible to prepare medicinal products with specific activity by isolating the enantiomers, and these enantiomers themselves exhibit a different pharmacokinetic profile from that of the known racemic mixture. It then becomes possible to use each of these enantiomers more effectively in precise indications for the treatment of perfectly identified pathologies.
Anti-ulcer drug tenatoprazole (tenatoprazole) is a new proton pump inhibitor, by the Japanese company Tokyo Tanabe, Japan’s Mitsubishi Corporation and Japan’s Hokuriku pharmaceutical companies jointly developed, has passed Phase II clinical trials. It acts on gastric parietal cells, reducing treatment of gastric ulcer, duodenal ulcer, reflux wall cell H + / K + -ATP activity, inhibition of gastric acid secretion, and H. pylori antibacterial activity, mainly for esophagitis and Zhuo – Ellison syndrome and gastric acid secretion disorders related diseases. Compared with the same types of drugs, Tenatoprazole suppress H + / K + -ATP enzyme activity is stronger, more stable, its efficacy than similar products currently widely used in clinical omeprazole strong 7 times. It has not been in the domestic market, nor ratified the production, with broad market prospects and development potential.
Proton pump inhibitors (proton pump inhibitors) for the treatment of acid-related diseases, the past ten years a wide range of clinical applications, better effect of the drug. It can quickly pass through the stomach wall membrane, gathered in a strongly acidic secretory tubules, and H + / K + -ATP enzyme (proton pump) thiol groups covalently bonded to form a disulfide bond, proton pump inactivation, inhibition of the enzyme H + / K + transport, so as to achieve the effect of acid suppression. Proton pump inhibitors and conventional clinical application of gastric acid suppression drugs H2 receptor antagonists compared with different sites of action and have different characteristics, namely acid-suppressing effect at night is good, rapid onset of acid inhibition strong and long time, easy to take these drugs can quickly and efficiently inhibit gastric acid secretion and clearance of Helicobacter pylori, it is widely used gastric ulcer, duodenal ulcer, reflux esophagitis and Zhuo – Ellison syndrome and other diseases treatment. Currently, proton pump inhibitors has been listed on the main omeprazole, lansoprazole, pantoprazole, rabeprazole and esomeprazole.Physical and
chemical properties ofwhite or white crystalline powder, melting point 174 ~ 175 ℃. Soluble in chloroform, insoluble in alcohol and water.
This product and other proton pump inhibitors as compared to chemically stable. China had 34 omeprazole preparations from Portugal, Brazil, India, China and other 13 countries, the stability of the measurements were made. The results showed that only six products (18%) during the trial showing good physical and chemical stability of. 27 products (79%) less (including Chinese product), the active ingredient a significant chemical decomposition, color and physical properties such as dissolution, are also a corresponding change. The results of a stability test designed to compare the various proton pump inhibitors show investigated eight days at 60 ℃, relative humidity of 75%, after omeprazole decomposition only 3% of the active ingredient, the tenatoprazole 77% of the data, said Alpha pantoprazole stability far superior to omeprazole, is already developed similar products in the most promising products.
Synthesis
Matsuishi, N.; Takeda, H.; Iizumi, K.; Murakami, K.; Hisamitsu, A. US Patent 4,808,596, 1989
Synthesis of Tenatoprazole 1 commences with the coupling of 2-mercapto-5-methoxyimidazo[4,5-b]pyridine 2 with 2-chloromethyl-4-methoxy-3,5-dimethyl pyridine hydrochloride 3 in the presence of potassium hydroxide affords 4 with 73% yield in ethanol and chloroform. The oxidation of the penultimate sulfide intermediate4 with m-CPBA in chloroform (100 vol) afforded 1
synthesis of 1 begins with the solvent-free condensation of 2-mercapto-5-methoxyimidazo[4,5-b]pyridine 2 with 2-chloromethyl-4-methoxy-3,5-dimethyl pyridine hydrochloride 3 to deliver the sulfide intermediate4 with 98% yield.
The final step of the synthesis is the oxidation of the sulfide intermediate with m-CPBA to form tenatoprazole 1. The sulfide intermediate 4 on treatment with 0.9 equiv of m-chloroperbenzoic acid (m-CPBA) at −10 to −15 °C afforded the crude tenatoprazole which was isolated as its sodium salt. The sodium salt of tenatoprazole 5 was purified by recrystallsation using dimethyl formamide and ethyl acetate (2:1 ratio) to yield the pure crystalline tenatoprazole sodium 5. Treatment of tenatoprazole sodium 5 with dil. HCl in the presence of acetone and water afforded the pure tenatoprazole 1
Tenatoprazole is a new type of gastric H + / K + -ATP enzyme inhibitors (proton pump inhibitor PPI), the chemical name 5-methoxy-2- (4-methoxy-3, 5-dimethyl-2-methylsulfinyl) imidazole and W, 5-b] pyridine, useful in the treatment of gastric ulcer, duodenal ulcer, reflux esophagitis and Zhuo – Ai syndrome and gastric acid secretion disorders related diseases.The drug was developed by Japan’s Tokyo Tanabe, Japan’s Mitsubishi Corporation and Japan’s Hokuriku pharmaceutical companies.Compared with other varieties of the same type, which inhibit H + / K + -ATP enzyme activity is stronger, ulcers of various tests are effective, and significantly improve the stability compared with other proton pump inhibitors.
US patent US4808596 “hidazo [4,5_b] pyridine compounds and pharmaceutical compositions containing same)) synthesis process disclosed Tenatoprazole the below formula:
By The route of 2-chloro-3,5-dimethyl-4-methoxypyridine hydrochloride with 2-mercapto-5-methoxy-imidazole, 5-b] pyridine under basic conditions condensation of Intermediate 2- [2- (3,5_-dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide] imidazo W, 5-b] pyridine, and then oxidizing the Thai duly omeprazole.This route for the synthesis of pull azole classic line, many pull azoles such as omeprazole can be synthesized by a similar route, this route mild condition, simple operation.But the route condensation, oxidation treatment after use of large amounts of toxic solvent chloroform, is not conducive to industrial scale; lower oxidation yields, the resulting Tenatoprazole containing unreacted starting materials 2- [2_ (3,5 – dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide] imidazo W, 5-b] pyridine, further comprising a sulfone by-product, N- oxide, N- oxide sulfone, These by-products may interfere with purification of tenatoprazole.
Japanese Patent invention Wo 丨 J JP05222038 “5_methoxy-2- [[(4_methoxy-3, 5-dimethyl-2-pyridyl) methyl] thio] imidazo [4,5 ~ b] pyridine and intermediates)) male
Synthesis open Tenatoprazole the below formula:
4-chloro-2-chloromethyl-3,5-dimethylpyridine -N- oxide 2_ mercapto _5_ methoxy-imidazo – [4, 5-b] pyridine in alkaline under condensation of Intermediate 5-Methoxy-2- (4-chloro-3,5-dimethyl-2-methylthio Bi) imidazo W, 5-b] pyridine-oxide -N- ( yield 82%), then refluxed in a solution of sodium methoxide in methanol to give 5-methoxy-2- (4-oxo-3,5-dimethyl-2-methyl sulfide) imidazo W , 5-b] pyridine -N- oxide (income ¥ 71%), and then at room temperature in methylene chloride, phosphorus trichloride treated with deoxy (yield 95%), and finally oxidation in Tenatoprazole (income Rate not reported).The novel synthetic route, mild reaction conditions, simple operation, the yield of each step is higher, but the route is too long resulting in a total yield is not high, prolonged and rising production costs.
To a reaction flask was added 2-mercapto-5-methoxy-imidazole, 5-b] pyridine 18. lg, 12g of sodium hydroxide and water 144. 8g, stirred and dissolved at 25 ° C, was added dropwise within Ih 20g of the 2-chloromethyl-dimethyl-4-methoxy _3,5- pyridine hydrochloride and 60g of water were mixed solution dropwise at 25 ° C the reaction 2h, the reaction is completed, filtered, washed with water 144. 8g, 36. 2mL ethanol and washed to obtain a wet powder; wet powder was dried at 50 ° C in vacuo to constant weight to give 2- [2_ (3,5-dimethyl) -4-methoxy-pyridylmethyl sulfide -5 – methoxy] imidazo [4,5-b] pyridine 32. Og;
2) Preparation of tenatoprazole lithium salt: To a reaction flask was added 2- [2- (3,5_-dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide] imidazo W, 5-b] pyridine 30g, dichloromethane 300g, methanol 15g, and dissolved with stirring; cooled to -10 ° C, was added dropwise the 15g and 485g m-chloroperbenzoic acid in methylene chloride mixed solution, dropwise addition the reaction temperature was controlled at -10 ° C, the dropping time of the pool; the dropwise addition, the temperature control at -10 ° C, the reaction 30min; completion of the reaction, at 10 ° C by the dropwise addition of lithium hydroxide and 135g water 15g mixed solution, drip complete, insulation stirred Ih; filtered cake was washed with acetone 60mL, get wet powder; wet powder was dried at 35 ° C under vacuum to constant weight to give Tenatoprazole lithium salt ^ g;
3) Preparation Tenatoprazole: To a reaction flask 加入泰 pantoprazole lithium salt 25g, acetone 63mL, water IOOmL, cooling M0 ° C, dropping lmol within lh / L hydrochloric pH7 0, drops. Albert, stirring 30min; the filter cake washed with water 50mL, washed with acetone and 50mL, wet powder was dried at 35 ° C under vacuum to constant weight to give Tenatoprazole 19. Sg.
Example 2:
a) 2- [2- (3,5-dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide] imidazo W, 5_b] pyridine (4) Preparation: To the reaction flask was added 2-mercapto-5-methoxy-imidazo 44,5-b] pyridine 18. lg, 11. 2g of potassium hydroxide and water 217mL, stirred and dissolved at! 35 ° C, was added dropwise within 2h by the 33. 3g of 2-chloro-3,5-dimethyl-4-methoxypyridine hydrochloride and 133. 2mL water mixed solution, dropwise at 35 ° C the reaction 4h, the reaction is completed, filtration, water 217mL, 72. 4mL ethanol and washed to obtain a wet powder; wet powder was dried at 60 ° C in vacuo to constant weight to give 2- [2- (3,5-dimethyl) -4-methoxy-pyridylmethyl sulfide -5-methoxy-yl] imidazo W, 5-b] pyridine 33. Ig;
2) Preparation of tenatoprazole lithium salt: To a reaction flask was added 2- [2- (3,5_-dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide] imidazole and W, 5-b] pyridine 30g, dichloromethane 400mL, methanol 50mL, stirring to dissolve; cooled to _15 ° C, was added drop by the m-chloroperoxybenzoic acid 16g of mixed solution of dichloromethane and 400mL , the process reactor temperature control was added dropwise at -20 ° C, the dropping time 2. 5h; the dropwise addition, the temperature control _15 ° C, the reaction 35min; completion of the reaction, at 15 ° C by the dropwise addition of 20g of hydrogen Lithium oxide and 200mL water mixed solution, drip completed, insulation mixing 1. 5h; filtration, the filter cake washed with acetone 90mL, get wet powder; wet powder was dried at 40 ° C under vacuum to constant weight to give Tenatoprazole lithium salt 28. 6g;
3) Preparation Tenatoprazole: To a reaction flask 加入泰 pantoprazole lithium salt 25g, ethanol 75mL, water 150mL, cooled to 10 ° C, dropping 6mol / L hydrochloric pH8 0 within 2h,. drops Albert, stirring 40min; the filter cake washed with water 100mL, washed with acetone IOOmL, wet powder was dried at 40 ° C under vacuum to constant weight, yield powder was Tenatoprazole 19. 5g.
Example 3:
a) 2- [2- (3,5-dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide] imidazo W, 5_b] pyridine (4) Preparation: To the reaction flask was added 2-mercapto-5-methoxy-imidazo 44,5-b] pyridine 18. lg, 8.4g of lithium hydroxide and water 180ml, stirred and dissolved at 30 ° C, was added dropwise within 1. 5h by the Guang .6g 2-chloro-3,5-dimethyl-4-methoxy-pyridine hydrochloride and 90mL water mixed solution, drop end at 30 ° C reaction 3h, the reaction is complete, filtration, water 217mL , washed with 85mL ethanol to obtain a wet powder; wet powder was dried at 55 ° C in vacuo to constant weight to give 2- [2- (3,5-dimethyl) -4-methoxy-5-pyridylmethyl sulfide oxy] imidazo [4,5-b] pyridine 32. 4g;
2) Preparation of tenatoprazole lithium salt: To a reaction flask was added 2- [2- (3,5_-dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide] imidazole and W, 5-b] pyridine 30g, dichloromethane 600mL, methanol 60mL, stirring to dissolve; cooled to -20 ° C, was added drop by the m-chloroperoxybenzoic acid 18g of mixed solution of dichloromethane and 600mL , dropwise addition the reaction temperature is controlled at _20 ° C, the dropping time of the pool; the dropwise addition, the temperature control at _20 ° C, the reaction 40min; completion of the reaction, at 20 ° C by the dropwise addition of lithium hydroxide and 300mL 30g water mixed solution, drip complete insulation mixing tank; filter, the filter cake washed with acetone and 120mL, get wet powder; wet powder was dried at 40 ° C under vacuum to constant weight to give Tenatoprazole lithium salt 28. 7g;
3) Preparation Tenatoprazole: To a reaction flask 加入泰 pantoprazole lithium salt 25g, methanol 75mL, water 120mL, cooled to 5 ° C, dropping dilute hydrochloric acid within 1 5h tune pH7 5,.. drops Albert, stirring 35min; the filter cake washed with water 75mL, 75mL acetone washed, wet powder was dried at 40 ° C under vacuum to constant weight, yield powder was Tenatoprazole 19. 6g.
Example 4:
a) 2- [2- (3,5-dimethyl) -4-methoxy-picolyl thioether _5_ methoxy] imidazo [4,5_b] pyridine ⑷ Preparation of: To a solution The reaction flask was added 2-mercapto-5-methoxy imidazole -½, 5-b] pyridine 18. lg, IOg sodium hydroxide and water 150ml, stirred and dissolved at 30 ° C, the 1. 5h dropwise added from 21 . 5g of 2-chloro-3,5-dimethyl-4-methoxypyridine hydrochloride and 90mL water mixed solution, dropwise at 30 ° C the reaction 3h, completion of the reaction, was filtered, washed with water 217mL, The wet powder was washed with ethanol to give 85mL; wet powder was dried at 55 ° C in vacuo to constant weight to give 2- [2- (3,5_-dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide ] imidazo [4,5-b] pyridine 32. 3g;
2) Preparation of tenatoprazole lithium salt: To a reaction flask was added 2- [2- (3,5_-dimethyl) -4-methoxy-5-methoxy-pyridylmethyl sulfide] imidazole and W, 5-b] pyridine 30g, dichloromethane 500mL, methanol 60mL, stirring to dissolve; cooled to -20 ° C, was added drop by the m-chloroperoxybenzoic acid 18g of mixed solution of dichloromethane and 500mL , the process reactor temperature control was added dropwise at -20 ° C, the dropping time pool; the dropwise addition, the temperature control in -20 ° C, the reaction 40min; completion of the reaction, at 20 ° C by the dropwise addition of lithium hydroxide 30g and 300mL water mixed solution, drip complete insulation mixing tank; filter, the filter cake washed with acetone and 120mL, get wet powder; wet powder was dried at 40 ° C under vacuum to constant weight to give Tenatoprazole lithium salt 28. 6g;
3) Preparation Tenatoprazole: To a reaction flask 加入泰 pantoprazole lithium salt 25g, isopropanol 75mL, water 120mL, cooled to 5 ° C, dropping 3mol / L hydrochloric within 1 5h. . pH7 5, drops Albert, stirring 35min; the filter cake washed with water 75mL, 75mL acetone washed, wet powder was dried at 40 ° C under vacuum to constant weight, yield powder was Tenatoprazole 19. 7g.
PAPER
An Improved Synthesis of Antiulcerative Drug: Tenatoprazole
Department of Research and Development, Srini Pharmaceuticals Ltd., Plot No. 10, Type-C, Road No. 8, Film Nagar, Jubilee Hills, Hyderabad-500033, Andhra Pradesh, India, Department of Chemistry, Osmania University, Tarnaka, Hyderabad-500007, Andhra Pradesh, India and Research and Development, Integrated Product Development Organization, Innovation Plaza, Dr. Reddy’s Laboratories Ltd., Bachupally, Qutubullapur, R. R. Dist. 500 072, Andhra Pradesh, India
* To whom correspondence should be addressed. Telephone: +91 9490783736. E-mail: drkvr_ou@yahoo.com;kvgr1951@rediffmail.com., †Srini Pharmaceuticals Ltd.
, ‡Osmania University.
, §Dr. Reddy’s Laboratory Ltd.
An efficient, cost-effective and multikilogram-scale process for the synthesis of tenatoprazole 1, an antiulcerative drug, is described. The key steps in this synthesis involve the coupling of 2-mercapto-5-methoxyimidazo[4,5-b]pyridine 2 with 2-chloromethyl-4-methoxy-3,5-dimethyl pyridine hydrochloride 3 to yield 4 and its subsequent oxidation with m-CPBA to produce sulfoxide 1. The process has been scaled up for the multikilogram-scale of compound 1 with an overall yield of 72%. The new process requires no purification process and affords the target compound 1 with 99.8% purity by HPLC.
the (+) enantiomer of tenatoplazole can be obtained by using chloroform, an industrially acceptable solvent, in accordance with the method proposed by Umemura et al. (J. Org. Chem. 1993, 58, 4592) as follows:
Example 1 (−)-5-methoxy-2-{(4-methoxy-3,5-dimethyl-2-pyridyl)methyl]sulfinyl}-1H-imidazo[4,5-b]pyridineThe conditions for preparative chromatography, shown as an example, are as follows:
Column: 265×110 mm ChiralPak®
Chiral Stationary Phase selector of the Amylose tris type [(S)-a methylbenzylcarbamate]
Flow rate: 570 ml/min
Detection: UV 240 nm
Temperature: Ambient temperature
These conditions are implemented on a liquid preparative chromatography apparatus.
Introduce approximately 2 g of the racemic mixture if tenatoprazole exhibiting purity higher than 99.5%. The (−) enantiomer is identified by measuring the angle of optical rotation, which must be laevogyre. This measurement can be performed directly on the column, the product being dissolved in the solvent (acetonitrile).
Example 2 (+)-5-methoxy-2-{(4-methoxy-3, 5-dimethyl-2-pyridyl)methyl]sulfinyl}-1H-imidazo[4,5-b]pyridine(R)-(+)-binaphthol 85 g (0.311 mol, 0.2 equivalence), ortho titanic acid isopropyl 42 g (0.148 mol, 0.1 equivalence), water 55 g (3.06 mol) and chloroform 7.5 L were stirred for 1 hour at room temperature. To the resultant, 5-methoxy-2-{(4-methoxy-3,5-dimethyl-2-pyridyl)methyl]thio}imidazo[4,5-b]pyridine (MPI), 0.5 kg, was added and stirred for 0.5 hours at room temperature. The thus-prepared mixture was cooled to 5° C. and then 70% aqueous solution of tert-butylhydroperoxide, 0.4 L (approx. 3.0 mol, 2.0 equivalence) was added and stirred for 72 hours at the same temperature as above. After the reaction endpoint was confirmed by HPLC, an aqueous solution of sodium hydroxide was added thereto to separate the aqueous layer, thus removing foreign matter. Then, the resultant was concentrated. Ethyl acetate was added to concentrated residues, which were then heated and suspended. The thus-prepared crude crystalline substances were dissolved in water and neutralized to pH 6.8 with a diluted sulfuric acid solution which was chilled with ice. Deposited crystals were filtered, dried and recrystallized by addition of ethanol to obtain (+)-5-methoxy-2-{(4-methoxy-3,5-dimethyl-2-pyridyl)methyl]sulfinyl}-1H-imidazo[4,5-b]pyridine {(+)-TU-199}
When measurements were carried out, for a solubility of (+), (−) forms and a racemic form (±) of tenatoprazole in relation to water, it was found that the (+) form dissolved almost 3 times greater than the racemic body and (−) form dissolved over 2 times greater than the racemic form, exhibiting favorable physical properties in preparing drugs (refer to Table 2 below).
TABLE 2
(+) form
(−) form
(±)racemic form
Solubility (water) μg/mL
93.0
74.4
34.6
CLIPS
Tenatoprazole is a pyridinylmethylsulfinyl imidazopyridine compound, which is a weak base. This compound has three pKas. One is the pyridine pKa of pyridinylmethyl moiety and the others are the imidazole pKa and the pyridine pKa of the imidazopyridine moiety. The pyridine pKa1 enables tenatoprazole accumulation in the acidic canaliculus of the parietal cell. Protonation of the imidazopyridine ring enhances electron deficiency at the C-2 position, allowing intramolecular rearrangement to the active form. The active form is the sulfenic acid and/or cyclic sulfonamide, and reacts with luminal cysteine thiols of the enzyme to inhibit the enzyme activity
Synthesis route from 2-mercapto-5-methoxy-imidazo [4,5-b] pyridine (2) and 2-chloro-3,5-dimethyl-4-methoxypyridine hydrochloride ( 3) by nucleophilic substitution synthesis of 2- (4-methoxy-3,5-dimethyl-2-methylthio) -5-methoxy-imidazo [4,5-b] pyridine (4) the oxidation of 4 1. Figure 1 is a synthesis route of tenatoprazole
《Organic Process Research & Development》 20081112 Somaiah Sripathi et al. An Improved Synthesis of Antiulcerative Drug:Tenatoprazole 第804-806页 1-6 第13卷,
2
*
《Synthetic Communication》 20080101 Liyan Dai et al. Improved Synthetic Approach to Tenatoprazole 第576-582页 1-6 第38卷,
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. Mass spectrometry was performed for this isolated impurity by ESI technique in positive mode. The positive DI-MS spectrum of isolated impurity exhibited molecular ion peak as [M + H]+ at m/z 592.29 and as sodium adduct [M + Na]+ at m/z 614.28. The MS/MS data displayed a dominant fragment at m/z 574.28 which is 17 amu less than the molecular ion peak indicating a removal of the hydroxyl group.
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Chidamide (Epidaza) is an 
To a suspension of 0.29 g (1.78 mmol) of N, N’-carbonyldiimidazole in tetrahydrofunan (15 ml) is added 0.50 g (1.78 mmol) of 4-[N-(Pyridn-3-ylacryloyl)aminomethyl]benzoic acid, followed by stirring at 45°C for 1 hour. After cooling, the reaction mixture is added to a separately prepared tetrahydrofunan (10 ml) solution including 0.28 g (2.22 mmol) of 4-fluoro-1,2-phenylenediamine and 0.20 g (1.78 mmol) of trifluoroacetic acid at room temperature. After reaction at room temperature for 24 hours, the deposited white solid is collected by filtration, washed with tetrahydrofunan, and then dried to give the title compound (0.40 g, 57%). 1H NMR (300 MHz, DMSO-d6): δppm: 4.49 (2H, d), 4.84 (2H, br.s), 6.60 (IH, t), 6.80 (2H, m), 6.96 (IH, t), 7.18 (IH, d), 7.42 (2H, d), 7.52 (IH, d), 7.95 (2H, d), 8.02 (IH, d), 8.56 (IH, d), 8.72 (IH, br. t), 8.78 (IH, s), 9.60 (IH, br.s). IR (KBr) cm“1: 3310, 1655, 1631, 1524, 1305, 750. HRMS calcd for C22Hι9N4O2F: 390.4170. Found: 390.4172. MA calcd for C22Hι9N4O2F: C, 67.68%; H, 4.40%; N, 14.35. Found: C, 67.52%; H, 4.38%; N, 14.42%.
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