Process for the manufacture of cyclic undecapeptides

Abstract

The present invention relates to processes and intermediates useful for the manufacture of cyclic undecapeptides, such as Alisporivir.

Claims

1. An HBF.sub.4 salt of ##STR00017##

2. An HBF4 salt of ##STR00018##

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1 is a proton NMR spectra for compound 3.

(2) FIG. 2 is a proton NMR spectra for compound 4.

DETAILED DESCRIPTION OF THE INVENTION

(3) The general process according to the present invention for producing cyclic polypeptides, more specifically, cyclic undecapeptides, such as Alisporivir, is shown in the scheme below; however, this general scheme can also be used to make cyclic polypeptides, more specifically, cyclic undecapeptides, derived from cyclosporine A, B, D, or G.

(4) ##STR00009## ##STR00010##

(5) Specifically, alisporivir can be made by converting cyclosporin A (compound (1) wherein R.sup.2 is ethyl) into a compound of formula 4 as shown above by acylation of cyclosporin A, to form acetyl-Cyclosporin A (2); ring opening; crystallization to obtain a compound 3, Edman degradation of compound 3; crystallization to obtain a compound 4 and then cyclizing compound 4 to form alisporivir (as shown below).

(6) ##STR00011## ##STR00012##

(7) The invention specially relates to the processes described in each section. The invention likewise relates, independently, to every single step described in a process sequence within the corresponding section. Therefore, each and every single step of any process, consisting of a sequence of steps, described herein is itself a preferred embodiment of the present invention. Thus, the invention also relates to those embodiments of the process, according to which a compound obtainable as an intermediate in any step of the process is used as a starting material.

(8) The invention also relates to intermediates which have been specifically developed for the preparation of the compounds according to the invention, to their use and to processes for their preparation.

(9) It is noted that in the present application, explanations made in one section may also be applicable for other sections, unless otherwise stated.

(10) Cyclosporin A, cyclosporin B, cyclosporin D or cyclosporin G or salts thereof, may be prepared, for example by fermentation.

(11) In one embodiment the present invention relates to a method for preparing compound of formula 3, comprising the steps of acylation of cyclosporin A, cyclosporin B, cyclosporin D or cyclosporin G to form acetyl-Cyclosporin A, B, D, or G; ring opening; and crystallization.

(12) In one embodiment the present invention relates to a method for preparing compound of formula 4 or a salt thereof, comprising Edman degradation, a reaction well known in the art, of a compound of formula 3 and crystallization thereof to obtain compound of formula 4.

(13) Another embodiment of the present invention relates to a method for preparing a compound of formula 3 or formula 4 wherein the purity of the Cyclosporin A starting material is >80% by weight

(14) Another embodiment of the present invention relates to a method for preparing a compound of formula 3 or formula 4 wherein the purity of the Cyclosporin A starting material is >85% by weight.

(15) Another embodiment of the present invention relates to a method for preparing a compound of formula 3 or formula 4 wherein the purity of the Cyclosporin A starting material is 60 to 80%, weight % assay.

(16) In the processes shown above, novel and inventive compounds are involved. Consequently, further subjects of the present invention are the compounds shown below.

(17) Compounds of formula 3 or salts thereof,

(18) ##STR00013##
wherein R is methyl, ethyl, propyl or phenyl, R′ is methyl or ethyl, and R.sub.2 is methyl, ethyl, or propyl.

(19) Compounds of formula 4 or salts thereof,

(20) ##STR00014##
wherein R is methyl, ethyl, propyl or phenyl, R′ is methyl or ethyl, and R.sup.2 is methyl, ethyl, or propyl.

(21) Compounds of formula 3 or salts thereof,

(22) ##STR00015## wherein R is methyl, ethyl, propyl or phenyl and R′ is methyl or ethyl.

(23) Compounds of formula 4 or salts thereof,

(24) ##STR00016## wherein R is methyl, ethyl, propyl or phenyl and R′ is methyl or ethyl.

(25) The following Examples represent preferred embodiments of the reaction steps, intermediates and/or the process of the present invention and serve to illustrate the invention without limiting the scope thereof.

(26) Preparation of Compound 3 HBF.sub.4 Salt with Merwein Salt

(27) Acetyl-Cyclosporin A (100 g as is) was reacted with trimethyloxonium tetrafluoroborate (32 g) at 20-25° C. in dichloromethane (180 mL). After 20 h, acetonitrile (200 mL) and water (650 mL) were added to perform the hydrolysis. After 3 h, at 20-25° C., the phases were separated and the reaction mixture was dried by azeotropic distillation with 2-Methyl-Tetrahydrofuran (solvent exchange dichloromethane/2-Methyl-Tetrahydrofuran). The desired product was then crystallized from 2-Methyl-Tetrahydrofuran (900 mL) and 2-Methoxy-2-methylpropane (400 mL) to provide compound 3 HBF.sub.4 as a white crystalline powder (63.9 g, after drying, purity >92%). 0.69, (3H,d,J=6.6 Hz); 0.71, (3H,d,J=6.5 Hz); 0.81, (6H,m); [0.82 . . . 0.89], (24H,m); 0.90, (3H,d,J=6.6 Hz); 0.93, (3H,d,J=6.6 Hz); 1.16, (6H,m); [1.23 . . . 1.50], (4H,m); 1.52, (1H,m); [1.32 . . . 1.73], (8H,m); 1.59, (3H,d,J=6.0 Hz); 1.65, (2H,m); 1.65, 2.13, (2H,m); 1.93, 1.94, (3H,s); 2.03, (1H,m); 2.19, (1H,m); 2.45, (3H,s); 2.72, (3H,s); 2.84, (3H,s); 2.86, (3H,s); 2.99, (3H,s); 3.02, (3H,s); 3.06, (3H,s); 3.62, 3.68, (3H,s); 3.78, (1H,m); 3.87, 4.53, (1H,d,J=17.2 Hz,18.6 Hz); 4.10, 4.26, (1H,d,J=18.6 Hz,16.8 Hz); 4.23, (1H,m); 4.60, (1H,m); 4.62, (1H,m); 4.66, (1H,m); 5.02, (1H,m); 5.13, (1H,dd, J=11.3 Hz,4.7 Hz); 5.26, (1H,m); 5.29, (1H,m); 5.32, (1H,m); 5.36, (1H,m); 5.39, (2H,m); 7.72, (1H,d,J=7.3 Hz); 8.14, (1H,d,J=7.3 Hz); 8.21, 8.35, (1H,d,J=7.3 Hz,8.1 Hz); 8.85, (2H,s,br); 8.96, (1H,d,J=8.4 Hz).

(28) Preparation of Compound 3 HBF.sub.4 Salt with Use of Trimethylorthoformate and Borontrifluoride Etherate

(29) A solution of Acetyl-Cyclosporin A (10 g) in dichloromethane (20 mL) was added at −15° C. to a slurry of dimethoxycarbenium tetrafluoroborate generate at −20° C. by a slow addition of borontrifluoride (2 ml) to a solution of trimethylorthoformate (2 ml) in dichloromethane (20 mL). After the addition, the slurry was allowed to warm up to room temperature and was kept stirring for 20 h. Afterward, Acetonitrile (10 ml) and water (10 ml) were added. After 2 h stirring at 0° C., phases were split. Then, after having washed the organic phase with water, solvent switched to 2-Methyl-Tetrahydrofuran and saturation with 2-Methoxy-2-methylpropane, compound 3 was obtained as a white solid which was dried under vacuum (5.1 g, >90% purity) (see FIG. 1)

(30) Preparation of Compound HBF.sub.4 Salt:

(31) The previously prepared salt of compound 3 (34.62 g) was charged to a reactor along with sodium carbonate (4.8 g), Toluene (50 mL) and water (50 mL). The resulting mixture was stirred at 20-25° C. for 30 minutes, and the phases were separated. Phenylisothiocyanate (3.81 g) was added drop wise in 1 h at 20-25° C. and the resulting reaction mixture was stirred until completion. Then methanol (20 mL), and 48% fluoroboric acid in water (2.5 g) was added and the mixture was stirred for an additional 1 h. Then water (25 mL) was added, and the phases were split. The aqueous layers were extracted once more with toluene (50 mL) and then extracted with 2-Methyl-Tetrahydrofuran (100 mL). The organic extract was dried azeotropically and the desired product was crystallized from 2-Methyl-Tetrahydrofuran (100 mL) and 2-Methoxy-2-methylpropane (50 mL) to provide compound 4 HBF.sub.4 as a white crystalline powder (ca. 30 g, after drying, >93% purity). (see FIG. 2) 0.69, (3H,d,J=6.2 Hz); 0.73, (3H,d,J=7.0 Hz); 0.81, (3H,t,J=7.3 Hz,7.3 Hz); 0.82, (3H,m); 0.85, (9H,m); 0.88, (6H,m); 0.91, (3H,d,J=7.0 Hz); 0.93, (3H,d,J=6.6 Hz); 0.99, (3H,d,J=7.0 Hz); 1.17, (6H,d,J=6.6 Hz); [1.30 . . . 1.55], (9H,m); 1.60, (3H,d,J=5.5 Hz); [1.56 . . . 1.72], (4H,m); 1.93, 1.95 (3H,s); 2.09, (1H,m); 2.14, (1H,m); 2.20, (1H,m); 2.74, (3H,s); 2.82, 3.06, (3H,s); 2.84, (3H,s); 2.87, (3H,s); 2.94, (3H,s); 3.02, (3H,s); 3.63, 3.68, (3H,s); 3.88, 4.52, (1H,d,J=17.2 Hz,18.6 Hz); 4.10, 4.24, (1H,d,J=18.7 Hz,m); 4.24, (2H,m); 4.39, 4.62, (1H,m); 4.66, (1H,m); 5.02, (1H,m); 5.08, (1H,m); 5.26, (2H,m); 5.32, (1H,m); 5.37, (1H,m); 5.39, (2H,m); 7.84, 8.51 (1H,d,J=7.3 Hz,8.1 Hz); 7.98, (3H,s,br); 8.07, 8.18 (1H,d,J=7.7 Hz,7.3 Hz); 8.13, 8.27, (1H,d,J=7.3 Hz,8.1 Hz).

(32) Preparation of Alisporivir:

(33) The previously described intermediate 4 (ca. 109 g) was added in portions at 15° C. to a mixture of toluene and sodium carbonate/water and stirred for 3 hours at 15° C. The aqueous phase was separated. The dipeptide (ca. 28 g) and N-hydroxybenzotriazole-mono-hydrate (8.1 g) was added at 0° C. Water (2.5 mL) was added to the mixture and N-methylmorpholine (17.9 g) was dosed at −10° C., followed by a solution of dicyclohexylcarbodiimide (21.9 g) in toluene dosed at −10° C. and stirred at that temperature for 3 hours. The reaction mixture was heated to 0° C. in a period of 10 hours. At completion, the reaction mixture was filtered and the filtrate was extracted with 5% sodium carbonate solution, 5% salt solution, 2M hydrochloric acid and 10% salt solution. The organic phase was filtered again and concentrated at 50° C. in vacuum.

(34) Sodium Borohydride (0.59 g) was dissolved in diglyme (14.3 g) at 20 to 40° C. The cloudy solution was chilled to 12-16° C. Glycine (0.59 g) was added at that temperature. The previously prepared product (6.2 g) in solution into ca. 6 g toluene was dosed to the white suspension at 12-16° C. Then the reduction was accelerated by adding methanol in toluene at 12-16° C. This addition was controlled and performed in 3 portions. The reaction was stirred for an additional 3 hours, and added to the quench solution (aqueous acetic acid) at an internal temperature of 10-20° C. The rate of addition was modified in line with the hydrogen evolution (no accumulation). The phases were separated, the upper organic product phase was extracted with water and subsequently concentrated.

(35) The previously prepared product (5 g) was dissolved in toluene (ca. 5 g) and dosed in a period of 40-60 minutes to a solution of sulphuric acid (0.8 g) in methanol (50 mL) at 50° C. After ca. 2 hours, the solution was chilled to 15-20° C. and 25% benzyltrimethylammoniumhydroxide (ca. 12 g) in methanol was dosed (exothermic reaction). After 2 hours, water was dosed and the mixture was stirred for approx. 20 hours at 22° C. At completion, the solution was chilled to 10-15° C., diluted with water and neutralised with dilute sulphuric acid. Following separation of the top toluene phase, methanol was removed from the lower product phase by distillation. The oily separated product was extracted with ethyl acetate and washed in sequence with water and salt solution, adjusting the pH to 7.0-7.5 with dilute sulphuric acid. Then the organic phase was dried azeotropically and then precipitated onto 0-5° C. cold heptane. The product suspension was filtered, washed with heptane and dried in vacuum at 60° C.

(36) The “undecapeptide amino acid” precursor (5 to 13% to the overall end mass) dissolved in dichloromethane and the DCC dissolved into dichloromethane were added continuously in parallel in ca. 10 h to a mixture of Cl-HOBT, and NMM in dichloromethane at 40° C. At the end of the addition, the mixture was stirred for an additional 2 h, filtered to remove the DCU salt and concentrated to give Alisporivir as a crude product.