Lipase-Catalyzed Regioselective Ester Hydrolysis as a Key Step in an

Apr 18, 2019 - Springtime in Grasse, France, is a fragrant affair. The town and its surroundings enjoy a warm, maritime... SCIENCE CONCENTRATES ...
4 downloads 0 Views 455KB Size
Article Cite This: Org. Process Res. Dev. XXXX, XXX, XXX−XXX

pubs.acs.org/OPRD

Lipase-Catalyzed Regioselective Ester Hydrolysis as a Key Step in an Alternative Synthesis of a Buprenorphine Pro-Drug John S. Carey* and Emily McCann Indivior, Henry Boot Way, Priory Park, Hull HU4 7DY, U.K.

Downloaded via UNIV AUTONOMA DE COAHUILA on April 19, 2019 at 01:02:58 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.

S Supporting Information *

ABSTRACT: This paper describes the development of an alternative route toward a hemiadipic acid pro-drug of buprenorphine. Buprenorphine was acylated with adipic acid monoethyl ester. A regioselective ester hydrolysis using C. antarctica lipase B cleaved the sterically less-hindered alkyl ester in the presence of the more labile phenolic ester. In this manner the pro-drug could be isolated in good yield and high purity. KEYWORDS: Buprenorphine, pro-drug, biocatalysis, C. antarctica lipase B, Novozyme 435



DISCUSSION In the preceding manuscript we reported our process development activities toward a pro-drug of buprenorphine 2.1 The route outlined in Scheme 1 had been used successfully

adipic acid and DCU byproduct at the end of the reaction was a significant challenge. The weight and volume of excess adipic acid to be removed was significantly greater than the weight and volume of intermediate to be isolated, so this caused significant equipment configuration restraints. (2) Effectively washing the adipic acid filtercake to maximize the recovery of the product was not trivial. Washing the adipic acid filtercake with a large volume of tetrahydrofuran (THF) risked redissolving significant amounts of the adipic acid, which in turn would lead to losses to the mother liquors upon precipitation of the crude 2 as the HCl salt. The use of alternative solvents was not effective, so the compromise position was to wash the adipic acid filtercake with a minimum amount of THF and accept a 5−10% loss in yield. (3) Although adipic acid is cheap and readily available, the use of 13 equiv of adipic acid is wasteful from a Green chemistry perspective. There were also significant concerns regarding the physical nature of the dichloromethane (DCM) reslurry used to reduce the levels of impurity 3 from the level of ∼4% present in the crude 3 to less than 1% present in the intermediate grade 2. The success of the reslurry of crude 2 in DCM appeared to be highly dependent upon the physical properties of the crude 2, in that only a partial THF solvate seemed to be a viable substrate. DCM was unique as the only solvent the process worked in. Given our lack of understanding around the process, the concern was, if for any reason the DCM reslurry stopped working there was no fallback option. In addition, DCM is an ICH class 2 solvent, and our desire was to eliminate class 2 solvents from the latter stages of our processes. During the discovery phase,2 many of the analogues were prepared by the opening of the cyclic anhydride with the phenol group. This approach is not possible for pro-drug 2 due to the known instability of the required monomeric cyclic adipic anhydride.3

Scheme 1. Current, Scale-up Route to Pro-Drug 2

to prepare seven batches of ∼7 kg of drug substance on a 250 L scale. However, there were some weaknesses in the route that meant investigations into alternative routes of synthesis were warranted. The weaknesses related to the control of the 2:1 byproduct 3 produced during the esterification reaction, Figure 1. By using 13 equiv of adipic acid the production of impurity 3 was limited to ∼5% at the end of the reaction. This in turn had several knock-on effects; (1) Removal of the excess

Received: January 16, 2019

Figure 1. Structure of 2:1 adduct 3. © XXXX American Chemical Society

A

DOI: 10.1021/acs.oprd.9b00026 Org. Process Res. Dev. XXXX, XXX, XXX−XXX

Organic Process Research & Development

Article

adduct 3, less than 0.1% of acid 2, and less than 0.1% unreacted buprenorphine 1. A limited screen of hydrolytic enzymes for the conversion of ester 5 to acid 2 was undertaken. From which, C. antarctica lipase B was identified as a suitable enzyme that was able to perform the selective hydrolysis of the ethyl ester, without significant hydrolysis of the phenolic ester. On the basis of prior experience,6 we elected to use Novozym 435 as a supported version of the C. antarctica lipase B enzyme for the development work. The same prior experience suggested that performing the reaction at pH 7.0 in aqueous t-BuOH could be effective. By maintaining the pH of reaction medium at pH 7.0 by the constant addition of aqueous NH3, the loading of Novozym 435 could be reduced to less than 10% by weight relative to the substrate 5. A simple Design of Experiments looking at reaction temperature (20 vs 40 °C) and t-BuOH/H2O ratio (50/50 vs 90/10 v/v) provided some workable condition. At 40 °C, ∼4% of buprenorphine 1 was produced via background hydrolysis. With a 50/50 v/v t-BuOH/H2O ratio significant loss of enzyme from the support was observed. The preferred reaction conditions were: ester 5 was suspended in 90/10 v/v t-BuOH/H2O at 20 °C. The pH was adjusted from ∼pH 3 to pH 7.0 by the addition of aqueous NH3. The Novozym 435 was added, and the reaction continued at pH 7.0 and 20 °C for 24 h.7 At the end point of the reaction, the level of unreacted ester 5 was less than 0.5%, the level buprenorphine 1 was less than 1.0%, and the level of 2:1 adduct 3 remained constant at less than 0.1%. Following completion of the reaction, the Novozym 435 was removed by filtration, and the resin was washed with aqueous t-BuOH. The filtrate and wash were combined and acidified to pH 2.0 by the addition of hydrochloric acid, thus causing the product 2 to crystallize from solution as the HCl salt. Intermediate grade 2 was isolated following filtration in 84% yield. The high-performance liquid chromatography (HPLC) purity of the intermediate grade product was high; unreacted ester 5 was present at 0.3%, buprenorphine 1 was at 0.5%, and the 2:1 adduct 3 was less than 0.1%. The overall yield from buprenorphine 1 to the intermediate grade 2 for this alternative route was 75%, which compares favorably with the yield for the current supply route (72.5%).1 The intermediate grade 2 was recrystallized using dimethyl sulfoxide (DMSO)/ethyl acetate (EtOAc).1 The purified acid 2 was isolated in 92% yield and with high purity. The levels of both unreacted ester 5 and buprenorphine 1 had reduced slightly, with both being at 0.2%.8 In summary, an alternative route from buprenorphine 1 to pro-drug 2 was developed that avoided (1) the use of a vast excess of adipic acid and (2) the previously reported, but poorly understood, slurry process that controlled the levels of the 2:1 impurity 3. With high-quality adipic acid monoethyl ester 4 that contained low levels of residual adipic acid, the 2:1 impurity 3 could be avoided. A highly regioselective ester hydrolysis using an enzyme was developed that cleaved the sterically less-hindered alkyl ester in the presence of a more labile phenolic ester. In this manner pro-drug 2 could be isolated in good yield and high purity. The process was demonstrated on multigram scale by processes that would have been amenable to further scale-up if required.

To enable the acylation to occur without using a large excess of the acid, it was decided to investigate the use of adipic acid monoethyl ester 4 to generate the corresponding ester 5 (Scheme 2). It was then hoped that an enzyme could be Scheme 2. Alternative Route for the Synthesis of Pro-Drug 2

identified to perform a regioselective ester hydrolysis of ester 5 to produce pro-drug 2 without hydrolysis of the phenolic ester linkage to regenerate buprenorphine 1. Although a phenolic ester is more reactive than an alkyl ester under the standard conditions used for ester saponification, it was anticipated that the very different steric environments around each ester linkage would enable an enzyme to perform the selective hydrolysis of the sterically less hindered ethyl ester in the presence of the sterically more hindered, yet hydrolytically more labile, phenolic ester. The use of adipic acid mono t-butyl ester and subsequent deprotection under acidic conditions would not be practical due to the known instability of buprenorphine under acidic conditions.4 Ester 5 was readily formed by reaction of buprenorphine base 1 with 1.1 equiv of adipic acid monoethyl ester 4 in the presence of N-ethyl-N′-(3-(dimethylamino)propyl)carbodiimide hydrochloride (EDC·HCl) and catalytic 4dimethylaminopyridine (DMAP). Following aqueous (aq) workup and solvent removal the free base of ester 5 could be isolated as a viscous oil. A point to note, the preliminary enzyme screening was performed on the free base. To improve the ease of handling the free base was converted into the corresponding HCl salt following treatment of with anhydrous HCl in ethanol. The corresponding HCl of ester 5 was a highly crystalline compound that could be isolated by filtration. A process was developed, whereby, following the aqueous workup, a solvent exchange from DCM5 to EtOH was performed thus enabling the precipitation of the HCl salt without isolation of the free base. With this process, ester 5 could be produced in 89% yield. A key to the success of this approach was the quality of the adipic acid monoethyl ester. Any unprotected adipic acid present would react to generate small amounts of the desired pro-drug 2 and the 2:1 adduct 3. The presence of small amounts of the ultimate target compound was not an issue. However, the presence of the 2:1 adduct 3 was problematic, as it would track through the subsequent stages of chemistry unreacted and end up as an undesired impurity in the drug substance. With adipic acid monoethyl ester 4 that contained less than 0.1% of adipic acid, ester 5 could be produced that contained less than 0.1% of 2:1 B

DOI: 10.1021/acs.oprd.9b00026 Org. Process Res. Dev. XXXX, XXX, XXX−XXX

Organic Process Research & Development



Article

over 18 h, then filtered. The filtercake was washed with EtOAc (2 × 100 mL) and then dried under vacuum at 60 °C for 24 h to afford purified compound 2 as a white solid (18.30 g, 92%). δH ((CD3)2SO, 400 MHz) 0.38−0.44 (1H, m), 0.55−0.72 (4H, m), 1.02 (9H, s), 1.09−1.16 (1H, m), 1.27 (3H, s), 1.36 (1H, td, J = 12.8, 6.8 Hz), 1.47 (1H, dd, J = 13.2, 8.8 Hz), 1.54−1.71 (5H, m), 1.81 (1H, t, J = 12.4 Hz), 1.91 (1H, d, J = 12 Hz), 2.14 (1H, t, J = 9.4 Hz), 2.21−2.26 (3H, m), 2.58 (2H, t, J = 6.8 Hz), 2.76−2.81 (1H, m), 2.88 (1H, dd, J = 20.0, 6.8 Hz), 3.00−3.45 (8H, m), 3.96 (1H, d, J = 6.8 Hz), 4.66 (1H, s), 5.46 (1H, br s), 6.76 (1H, d, J = 8.4 Hz), 6.96 (1H, d, J = 8.0 Hz), 9.91 (1H, br s), 12.07 (1H, br s); δC ((CD3)2SO, 100 MHz) 3.0, 5.8, 6.1, 17.9, 20.5, 24.3, 24.5, 25.0, 26.8, 29.4, 31.5, 31.8, 33.3, 33.8, 35.8, 42.0, 44.5, 45.0, 52.5, 57.5, 57.7, 79.3, 79.9, 95.4, 120.3, 124.0, 131.7, 131.9, 132.2, 150.2, 171.4, 174.7; MS (ESI+) m/z 596.5 [M + H]+.

EXPERIMENTAL SECTION Buprenorphine Hemiadipate Monoethyl Ester Hydrochloride 5. Buprenorphine base 1 (30.00 g, 64.16 mmol, 1.0 equiv), DMAP (0.393 g, 3.21 mmol, 0.05 equiv), and EDC· HCl (13.53 g, 70.58 mmol, 1.10 equiv) were dissolved in dichloromethane (180 mL) at ambient room temperature. Adipic acid monoethyl ester 4 (12.29 g, 70.58 mmol, 1.10 equiv) was dissolved in dichloromethane (60 mL) and added to the buprenorphine solution. The reaction was stirred at ambient room temperature overnight. The reaction mixture was washed consecutively with 5% w/v aq citric acid (120 mL), 5% w/v aq sodium bicarbonate (120 mL), then water (120 mL). The solution was then concentrated by distillation at atmospheric pressure to leave a residual volume of 60 mL. Ethanol (240 mL) was added, and the solution was concentrated by distillation at atmospheric pressure to leave a residual volume of 120 mL. The resulting solution was cooled to 0 °C, and acetyl chloride (5.70 mL, 80.17 mmol, 1.25 equiv) was added dropwise. The resulting suspension was stirred at 0 °C for 2 h and then filtered. The filtercake was washed with chilled ethanol (60 mL) and then dried in a vacuum oven to afford compound 5 as a white solid (37.53 g, 89%). mp 241 °C (differential scanning calorimetry (DSC)). δH (CDCl3, 400 MHz) 0.43−0.50 (1H, m), 0.67−0.75 (1H, m), 0.78−0.89 (3H, m), 1.11 (9H, s), 1.24−1.37 (8H, m),1.58 (1H, dd, J = 13.6, 8.8 Hz), 1.69−1.79 (4H, m), 1.86−2.00 (3H, m), 2.27 (1H, t, J = 9.4 Hz), 2.32−2.36 (2H, m), 2.56− 2.64 (3H, m), 2.73−2.91 (3H, m), 3.17 (1H, d, J = 19.2 Hz), 3.36−3.39 (1H, m), 3.46 (3H, s), 3.49−3.55 (1H, m), 3.57− 3.65 (1H, m), 3.93 (1H, d, J = 6.4 Hz), 4.13 (2H, q, J = 7.2 Hz), 4.53 (1H, s), 5.77 (1H, s), 6.72 (1H, d, J = 8.0 Hz), 6.91 (1H, d, J = 8.0 Hz), 11.16 (1H, br s); δC (CDCl3, 100 MHz) 3.0, 5.9, 6.2, 14.3, 17.3, 20.5, 24.2, 24.4, 25.4, 26.3, 30.1, 31.7, 32.5, 33.5, 33.8, 36.1, 40.6, 42.5, 44.6, 45.6, 52.7, 57.5, 58.7, 60.4, 79.5, 79.6, 96.6, 119.7, 124.0, 129.4, 131.4, 132.6, 150.3, 170.9, 173.1; mass spectrometry (MS) electrospray ionization (ESI+) m/z 624.5 [M + H]+. Buprenorphine Hemiadipate Hydrochloride 2. Ethyl ester 5 (25.00 g, 37.86 mmol) was suspended in 88% w/w aq tBuOH (250 mL), and the pH was adjusted to pH 7 by the addition of 1.5 M aq NH3 (28.1 mL). Novozym 435 (1.88 g) was added, and the mixture was stirred at 20 °C for 24 h. The pH was maintained at a constant pH 7.0 by the addition of 1.5 M aq NH3 (21.6 mL). Reaction completion was checked by HPLC analysis, target residual ester 5 less than 0.5%. The enzyme resin was removed by filtration, and the resin was washed with 88% aq t-BuOH (50 mL). The filtrate and wash were combined and cooled to 0 °C. 1 M aq HCl (70 mL) was added to achieve a final pH of pH 2.0. The resulting slurry was aged at 0 °C for 1 h and then filtered. The filtercake was washed with water (50 mL) followed by ethanol (50 mL) and then dried under vacuum to afford intermediate grade compound 2 as a white solid (20.17g, 84%). Intermediate grade buprenorphine hemiadipate hydrochloride 2 (19.91 g, 31.49 mmol) was suspended in DMSO (80 mL) and heated to 85−90 °C to dissolve the solids. The resulting solution was cooled to 60−65 °C, and seed crystals (0.02 g, 0.1% by weight) were added. EtOAc (20 mL) was added over 20 min while maintaining the temperature at 60− 65 °C. Further EtOAc (480 mL) was added over 4 h while maintaining the temperature at 60−65 °C. The resulting slurry was allowed to cool to ambient room temperature and aged



ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.oprd.9b00026. 1



H and 13C NMR spectra for compound 5 (PDF)

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. ORCID

John S. Carey: 0000-0002-3654-0063 Author Contributions

The manuscript was written through contributions of all authors. Notes

The authors declare no competing financial interest.



ACKNOWLEDGMENTS The authors would like to thank Dr. S. Martin (Aesica Pharmaceuticals) for invaluable advice and for sourcing highpurity adipic acid monoethyl ester 4.



ABBREVIATIONS DCC, N,N′-dicyclohexylcarbodiimide; DCM, dichloromethane; DCU, N,N′-dicyclohexylurea; DMAP, 4-dimethylaminopyridine; DMSO, dimethyl sulfoxide; EDC·HCl, N-ethylN′-(3-(dimethylamino)propyl)carbodiimide hydrochloride; equiv, equivalents; h, hour(s); NMR, nuclear magnetic resonance spectroscopy; THF, tetrahydrofuran; vs, versus; v/ v, volume/volume ratio; w/w, weight/weight ratio



REFERENCES

(1) Berrell, S. L.; Byard, S. J.; Carey, J. S.; Codina, A.; Davis, J. A.; Filer, K.; Garnett, A.; Marshall, R.; Martin, S. J.; Mykytiuk, J.; Northen, J.; Pandya, U.; Reid, G.; Smyth, C.; Watson, L. Process Development toward a Pro-Drug of Buprenorphine. Org. Process Res. Dev. 2019, DOI: 10.1021/acs.oprd.9b00025 (2) Chapleo, C. B.; Lewis, J. W. Buprenorphine derivatives and uses thereof. WO2007/110636, 2007. (3) Hill, J. W. Studies on Polymerization and Ring Formation. VI. Adipic Anhydride. J. Am. Chem. Soc. 1930, 52, 4110−4114. (4) Grivas, K.; Breeden, S. W.; Ganter, C.; Husbands, S. M.; Lewis, J. W. Acid Catalysed Rearrangement of the Thevinols: The Mechanism of Furanocodide Formation. Tetrahedron Lett. 1999, 40, 1795−1798.

C

DOI: 10.1021/acs.oprd.9b00026 Org. Process Res. Dev. XXXX, XXX, XXX−XXX

Organic Process Research & Development

Article

(5) One of the stated aims of the alternative route development work was to avoid the use of DCM. Although DCM was used as the reaction solvent for Stage 1 it was not perceived to be the only solvent the reaction would work in. Had more development time been available then the complete elimination of DCM would have been possible. (6) Atkins, R. J.; Banks, A.; Bellingham, R. K.; Breen, G. F.; Carey, J. S.; Etridge, S. K.; Hayes, J. F.; Hussain, N.; Morgan, D. O.; Oxley, P.; Passey, S. C.; Walsgrove, T. C.; Wells, A. S. The Development of a Manufacturing Route for the GPIIb/IIIa Receptor Antagonist SB214857-A. Part 2: Conversion of the Key Intermediate SB-235349 to SB-214857-A. Org. Process Res. Dev. 2003, 7, 663−675. (7) After 6−8 h, ∼3−4% of ester 5 remained unreacted. (8) As this work was at a relatively early stage of development, the determination of residual enzyme present in the drug substance was not performed.

D

DOI: 10.1021/acs.oprd.9b00026 Org. Process Res. Dev. XXXX, XXX, XXX−XXX