The First Kilogram Synthesis of Beclabuvir, an HCV NS5B Polymerase

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The First Kilogram Synthesis of Beclabuvir, an HCV NS5B Polymerase Inhibitor Jeffrey Bien, Akin H. Davulcu, Albert J DelMonte, Kenneth J. Fraunhoffer, Zhinong Gao, Chao Hang, Yi Hsiao, Wenhao Hu, Kishta Katipally, Adam Littke, Aghogho Pedro, Yuping Qiu, Maria Sandoval, Richard L. Schild, Michelle Soltani, Anthony Tedesco, Dale Vanyo, Purushotham Vemishetti, and Robert E. Waltermire Org. Process Res. Dev., Just Accepted Manuscript • DOI: 10.1021/acs.oprd.8b00214 • Publication Date (Web): 16 Aug 2018 Downloaded from http://pubs.acs.org on August 16, 2018

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Organic Process Research & Development

The First Kilogram Synthesis of Beclabuvir, an HCV NS5B Polymerase Inhibitor Jeffrey Bien, Akin Davulcu, Albert DelMonte*, Kenneth J. Fraunhoffer, Zhinong Gao, Chao Hang, Yi Hsiao, Wenhao Hu, Kishta Katipally, Adam Littke, Aghogho Pedro, Yuping Qiu, Maria Sandoval, Richard Schild, Michelle Soltani, Anthony Tedesco, Dale Vanyo, Purushotham Vemishetti, Robert E. Waltermire Chemical and Synthetic Development, Bristol-Myers Squibb Company, One Squibb Drive, P.O. Box 191, New Brunswick, New Jersey, 08903, United States [email protected]

1 ACS Paragon Plus Environment

Organic Process Research & Development 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

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Abstract: The process development and the kilogram-scale synthesis of Beclabuvir BMS-791325 (1) is described. The convergent synthesis features the use of asymmetric catalysis to generate a chiral cyclopropane fragment and coupling with an indole fragment via an alkylation. Subsequent palladium catalyzed intramolecular direct arylation efficiently builds the central 7-membered ring. The target was prepared in 12 linear steps with 5 isolations in an overall yield of 8 %.

Keywords: asymmetric, cyclopropanation, direct arylation, Palladium, Rhodium, catalysis

Introduction The hepatitis C virus (HCV) is transmitted through infected blood and currently affects an estimated 2–3 percent of the world’s population.1 Infection with hepatitis C is frequently asymptomatic but left untreated often leads to cirrhosis of the liver, liver failure, hepatocellular cancer and even death. In the not too distant past, most infections were treated with pegylated interferon-α and ribavirin. Treatment typically lasted 24–48 weeks and was physically demanding as patients often experience flu-like side effects. Fortunately, for those afflicted with HCV, there have been a number of modern treatments that offer reduced side effects and shortened treatment duration.2,3

In addition, many patients are treated with

combinations of these drugs and experience even greater cure rates. The HCV NS5B RNA polymerase is an attractive target for drug inhibition as it is required for HCV replication. The potential for a very selective inhibitor exists as there is no analogous enzyme in human cells. It was for this purpose that Beclabuvir (BMS-791325) (1) was selected as a potential drug candidate for the inhibition of HCV NS5B RNA polymerase.4

This paper details the

development of a convergent chemical route and production of multi-kilogram quantities of 1, which were required to meet initial toxicological and clinical supply material demands.

Results and Discussions In order to support the clinical development, a short and efficient route was desired which addressed the challenges of the Discovery route (Scheme 1). Although this approach adequately met initial material requirements, a more convergent approach that would be 3 ACS Paragon Plus Environment


amenable to rapid scale up was desired. One aspiration for a scalable synthesis was to eliminate the late stage achiral cyclopropanation and preparatory HPLC chiral separation as this would greatly impact yield and throughput. There was also concern regarding use of the reagent methyl 2-(dimethoxyphosphoryl)acrylate (9) utilized in the synthesis of 8, as we felt it would require extensive development to understand and control potential polymerization challenges.


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Scheme 1. Discovery Synthesis of 1.a

` a

Reagents and conditions: (a) THF, Et3SiH, cyclohexanone (77%); (b) Br2, HOAc (76%); (c) NaOH, THF,

MeOH (99%); (d) CDI, THF; (e) N,N-dimethylsulfamide, DBU, THF (74%); (f) Pd(PPh3)4, LiCl, Na2CO3, EtOH, toluene (88%); The arylboronic acid aldehyde is synthesized in 4 steps;5 (g) Cs2CO3, DMF (71)%; (h) trimethylsulfoxonium iodide, NaH, DMSO (83%); (i) Chiracel-OJ chromatography, 44% recovery (50% theoretical); (j) NaOH, THF, MeOH, H2O, (86%); (k) TBTU, DIPEA, DMF; (l) Chromatography, MeOH, CH2Cl2, (70%).

New Route Strategy Although a number of potential synthetic approaches were considered, we concentrated our efforts on identification of a convergent route.

As shown in the

retrosynthetic analysis (Figure 1), a disconnection strategy leading back to an achiral indole fragment (16) and a chiral cyclopropane fragment (17) was envisioned.



Figure 1. Revised Retrosynthetic Analysis of 1.

When considering the variety of options within this strategy, we leveraged existing knowledge. Work by our Discovery colleagues suggested a final coupling of intermediate 13 with the bridged piperazine 14 was preferable, as they found this approach afforded 1 in higher purity than alternative end games. An added benefit is that the relatively expensive bridged piperazine 14 is introduced in the final step.

Published work by Kosikowski6

(Scheme 2) also suggested that if the indole and cyclopropyl fragments were already coupled through the carbon-nitrogen bond (as shown in intermediate 15), a direct intramolecular palladium catalyzed direct arylation was possible to form the 7-membered ring and produce compound 13.

This was an important precedent as the direct arylation strategy would

significantly streamline the route by avoiding unnecessary steps to install a bromide on the indole and a boronic ester on the aryl group.7

The alcohol moiety of the cyclopropyl

fragment had to be activated to function as an alkylating agent, and preparation of cyclopropyl tosylate 17 not only served in this role, it also imparted a high level of crystallinity, which was viewed to be advantageous at the time. Another approach to improve the convergent nature of the synthesis was to couple the indole and cyclopropyl fragments with the N,Ndimethylsulfamide moiety already installed on the indole 16, obviating the need for ester hydrolysis and sulfamide coupling in the end-game of the synthesis.

Scheme 2. Precedence of Direct Indole Arylation to Form a 7-Membered Ring 6 ACS Paragon Plus Environment

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There are several practical advantages to this synthetic approach: (1) the convergent nature leads to a more efficient synthesis; (2) an earlier introduction of the chiral cyclopropane results in higher throughput even if a resolution is required; (3) the opportunity to modify the synthesis of the individual fragments to iteratively evolve the synthesis into a commercially viable route without having to modify the overall strategy.

Synthesis of the Indole Fragment 16 Scheme 3 reveals our synthesis of the indole fragment 16. We adopted a reductive alkylation and hydrolysis approach to prepare indole 18. A telescoped multi-step process was utilized to install the N,N-dimethylsulfamide to afford 16. The literature method for the reductive alkylation8 only afforded a 36% yield of 3 when the reaction was conducted with cyclohexanone.

The original conditions were

optimized for order of addition, acid, temperature, stoichiometry, solvents, and crystallization conditions. The modified process involved the controlled addition of TFA to a cold (0 ºC) toluene mixture of 2, Et3SiH (3 equivalents), and cyclohexanone (2 equivalents) while maintaining the temperature ≤20 ºC. The reaction was slowly warmed to 20-25 ºC and monitored for reaction conversion. Upon reaction completion, the addition of petroleum ether produced a suspension. Filtration, washing and drying of the cake directly afforded the crystalline material. This process was demonstrated on pilot scale and afforded 89 kg of 3 in 64.7-69.9% yield and >99.6 HPLC area percent purity over 2 batches. These procedures were repeated in a subsequent campaign and afforded 374 kg of 3 in 74.5-75.3% yield9 and >99.3 HPLC area percent purity over 4 batches.10

Scheme 3. Synthesis of Indole 16.



The hydrolysis conditions for the conversion of 3 to 18 were optimized for concentration of base, temperature, solvents, and crystallization conditions. The optimized conditions required the treatment of 3 in 2 N NaOH solution (6 L/kg)11 and isopropanol (IPA) (6 L/kg) at 75 ºC. Increased concentrations of NaOH lead to transesterification with IPA. Upon reaction completion, the mixture was cooled to 50 ºC and neutralized with 2.5 N HCl to pH ≤1 resulting in a white suspension. Cooling to 25 ºC and filtration afforded the desired product 16. This process was demonstrated on scale and afforded 39.9 kg in 95.2-96.5% yield and 99.8-99.9 HPLC area percent purity over 2 batches. These results were also duplicated in a subsequent campaign and afforded 342 kg in 96.2-97.2% yield and 99.6-99.7 HPLC area percent purity over 4 batches. The coupling of 18 with sulfonamide proved more challenging than initially anticipated. Although the desired product 16 was obtained when indole 18 was subjected to CDI mediated coupling conditions analogous to those used successfully for the 2-bromo analog 5, unexpectedly high levels of impurity 22 (Figure 2) were also produced by a competitive nucleophilic attack by the indole nitrogen on the activated intermediate 19.12 Although impurity 22 showed some promise of being able to convert to the desired indole 16 under relatively harsh conditions, (prolonged heating in the presence of excess N,N-


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dimethylsulfonamide), we felt a more attractive approach would be to circumvent the formation of impurity 22 entirely via an in-situ protection of the indole nitrogen.

Figure 2. Indole Coupling Impurity 22.

During the development of this chemistry, we found the optimal sequence involved generation of the acyl imidazole 19 with CDI, followed by Boc protection. By introducing catalytic 4-dimethylaminopyridine (DMAP) (0.2 equivalents) during formation of 19, it was found there was no initiation period and that the off-gassing was addition controlled which is an important control feature from a safety perspective. Boc protection of 19 effectively led to the formation of intermediate 20 which upon treatment with N,N-dimethylsulfamide and 1,8diazabicyclo[5.4.0]undec-7-ene (DBU) afforded intermediate 21. DBU was found to perform better than triethylamine (TEA), diisopropylethylamine (DIPEA), Cs2CO3, pyridine and 1,4diazabicyclo[2.2.2]octane (DABCO). Although acidic conditions were effective at removing the Boc protecting group, the associated CO2 off-gassing was a concern as it was not easily controlled. However, basic conditions at elevated temperatures (65 °C) were effective at removing the Boc group from 21 and allowed for the addition controlled evolution of CO2 during the subsequent neutralization with HCl. A number of solvents were screened for this reaction sequence with dichloromethane (DCM), acetonitrile (ACN), tetrahydrofuran (THF), dimethylformamide (DMF), and dimethylacetamide (DMAc) all demonstrated to be viable. Ultimately, DCM was chosen as it allowed for a facile distillative swap to isopropanol for the basic hydrolysis of the Boc group. Utilizing this approach, 38 kg of indole 16 was produced in 98.9-99.3 HPLC area percent purity and 65.6-76.6% yield in two batches. These results



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were also duplicated in a subsequent campaign in which 254 kg of 16 were prepared in 99.499.7 HPLC area percent purity and an improved 76.3-77.4% yield in 4 batches. Overall, the synthesis of the indole fragment 16 was conducted in 6 chemical transformations, but only 3 isolations in 53 % overall yield.

Synthesis of the Chiral Cyclopropane Multiple potential approaches to the cyclopropane fragment were considered. During our investigations, the acid alcohol cyclopropane 23 (Scheme 4) emerged as a key intermediate in the synthesis of the tosylate 17.

Intermediate 23 was not only highly

crystalline, but also formed a highly crystalline intermediate with (R)-α-methylbenzylamine ((R)-AMBA) to afford 23••(R)-AMBA. These crystalline properties provided flexibility to the synthetic approach to either leverage chiral salt formation as a way to conduct a resolution of racemic 23 or as a tool to upgrade 23 of moderate enantiomeric excess (ee). The 23••(R)AMBA could then be converted to tosylate 17 via an esterification and subsequent tosylation reaction (vide infra).

Scheme 4. Synthesis of Cyclopropane Fragment 17.

We envisioned that cyclopropane 23 could be produced by functional group transformation of the vinyl cyclopropane 24 as shown retrosynthetically in Figure 3. The seminal work by Huw Davies13 was clear precedent for the synthesis of the trisubstituted cyclopropane

24

in

good

enantioselectivity

from

C13)pheynyl]sulfonyl]-(2S)-pyrrolidinecarboxylate]dirhodium reaction between vinyl diazoacetate 25 and styrene 26.


the

tetrakis[1-[[4-alkyl(C11-

(Rh2(S-DOSP)4)

catalyzed

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Figure 3. Retrosynthetic Analysis to Cyclopropane 23

The

synthesis

of

vinyl

diazoester

25

involved

reacting

27

acetamidobenzenesulfonyl azide (4-ABSA) and DBU in ACN (Scheme 5).

with

4-

The crude

reaction mixture was diluted with methyl tert-butyl ether (MTBE) and an aqueous workup effectively removed the DBU by-products and ACN. This also induced the crystallization of the sulfonamide by-product, which was removed via filtration. The mixture was solvent swapped to n-heptane and afforded a 6–9 wt% solution of diazoester 25 in n-heptane in 75–80 % yield. The removal of ACN was found to be a critical operation as residual levels were detrimental to catalyst performance in the cyclopropanation. Controlling ACN levels to ≤ 0.5 % (relative to the n-heptane solvent) ensured high quality product in the subsequent step. Scheme 5. Synthesis of Vinyl Diazoacetate 25.

Before scaling up it was important to understand any stability or safety issues associated with this specific vinyl diazoester 25. Fortuitously, different from most diazo compounds, decomposition of 25 did not lead to nitrogen off-gassing. The inherent reactivity of 25 instead resulted in electrocylization to form pyrazole 28 (Figure 4). In the solid state,



this electrocyclization event had an onset temperature of 42 °C, and a heat of reaction of 872 J/g. In solution, it was shown that this reaction was solvent and concentration dependent.14 Calorimetry studies revealed that solutions of ≤10 wt% of 25 in n-heptane were suitable for processing, wherein the rate of the decomposition to 28 was approximately 3-5 % per day. One additional advantage of this electrocyclization decomposition mode was that the resulting pyrazole 28 was not soluble in n-heptane, and could easily be filtered off before the solution was introduced to the cyclopropanation reaction. Potency of the n-heptane solution of 25 would slowly decrease over time, however the purity of the reagent solution was always maintained.15

Figure 4. Electrocyclization of 25 to Afford Pyrazole 28

The styrene substrate 26 was generated in a two-step sequence (Scheme 6). In the first step the commercially available 2-methoxybenzaldehye (29) was brominated in glacial AcOH at 30–35 °C. Addition of water led to the direct precipitation of the crude product (30). The resulting wet cake was reslurried with a dilute sodium bicarbonate solution to remove any residual acid, affording the desired pure 2-bromo-5-methoxybenzaldehyde 30 in 83–90% isolated yield.16 Selection of the appropriate solvent, AcOH, was critical to minimize overbromination. The aldehyde 30 was then subjected to a Wittig olefination in THF. n-Heptane was added followed by an aqueous workup.

Optimization of the reaction and workup

conditions led to a facile crystallization of the triphenylphosphine oxide reaction by-product. However, the cyclopropanation reaction proved highly sensitive even to residual levels of triphenylphosphine oxide, so the mixture was passed through a basic alumina plug. Surprising, we still observed variable catalyst performance. Careful evaluation of the mixture revealed the culprit as very low levels of triphenylphosphine (200 - 400 ppm). To address the trace levels of this highly impactful impurity, an oxidative wash with sodium percarbonate and hydrogen peroxide was implemented during the aqueous workup. This converted any residual triphenylphosphine to triphenylphosphine oxide, which was readily removed via the original removal strategy (crystallization and passage through a basic alumina plug). After 12 ACS Paragon Plus Environment

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distillative solvent reduction, pure styrene 26 was obtained as a solution in n-heptane.17 Two batches were prepared on kilogram scale. The first was produced as a 31.2 wt% n-heptane solution in 64.7% yield and the second batch was prepared as a 21.5 wt% n-heptane solution in 54.8% yield from aldehyde 30.

Scheme 6. Synthesis of Styrene 26.

With access to n-heptane streams of the styrene 26 as well as the vinyl diazoester 25, the cyclopropanation chemistry was investigated.

The reactor was charged with 1.1

equivalents of 26 relative to diazoester 25 (the limiting agent). The Rh2(S-DOSP)4 catalyst was charged, and the diazoester 25 was added over approximately 1 hour at room temperature. Lab experiments demonstrated that 0.2 mol% catalyst loading could afford a 90% solution yield and 85–86% ee. On scale-up, enantioselectivity ranged between 80.6 and 83.2% ee. Upon reaction completion n-heptane was removed via distillation and methylene chloride was charged.

Scheme 7. Rhodium Catalyzed Asymmetric Cyclopropanation of Styrene 26.



The crude reaction stream, which consisted of a ~ 10:1 mixture of cyclopropane 24 to residual styrene 26, was then subjected to several telescoped functional group transformations to afford crystalline 23••(R)-AMBA (Scheme 8). This approach was necessary both due to the lack of crystalline intermediates and the desire to avoid diminishing the ee during isolations.

Scheme 8. Telescoped Functional Group Transformations of Vinyl Cyclopropane 24

The DCM solution of cyclopropane 24 was first cooled to -60 ˚C and exposed to a stream of ozonized air while maintaining the internal temperature ≤ -40 ˚C. Upon complete consumption of 24 a methanolic solution of sodium borohydride was then introduced to


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reduce the ozonide to the corresponding alcohol 31.18 The mixture was quenched with an aqueous HCl solution and an extractive workup afforded a DCM solution of 31. Along with desired product 31, the expected benzyl alcohol by-product 32 and alcohol 33, resulting from ozonolysis and reduction of the excess 0.1 equivalents of the styrene 26, were produced (Figure 5). Solvent exchange to MeOH/THF and subsequent introduction of an aqueous LiOH solution effected ester hydrolysis to afford a solution of 23••Li. Extraction of the reaction mixture with MTBE served to conveniently remove the two benzylic alcohols 32 and 33, along with other organic impurities, affording a relatively pure aqueous solution of lithium carboxylate 23••Li.

Figure 5. Alcohol Impurities Generated after Ozonolysis and Reduction

The aqueous lithium carboxylate stream was then acidified, and the corresponding acid was extracted into dichloromethane. While the free acid is crystalline (mp > 140 °C), we found that isolation from a variety of solvents failed to improve the optical purity. The dichloromethane solution was solvent exchanged to IPA:water, warmed to 60 °C, and treated with (R)-AMBA resulting in the crystallization of 23••(R)-AMBA. This process performed well in the lab, furnishing approximately 65% isolated yield of optically enriched 23••(R)AMBA. At pilot scale, this telescoped sequence afforded 61 kg with a 53% isolated yield in high chemical and optical purity (>99.0 HPLC area percent purity, >98 % ee). To our knowledge, this was the first use of the Davies catalyst system on multikilogram scale.

It was ideal from a fit-for-purpose perspective as it identified a key

intermediate to control quality and facilitate isolation (23••(R)-AMBA), was developed very quickly, and successfully enabled the delivery of early clinical supplies. The overall route to



23••(R)-AMBA was 7 linear steps, but only 2 isolations, affording high quality product in an overall 26% yield (Scheme 9).

Scheme 9. First Generation Synthesis of the Chiral Cyclopropane 23.

It was recognized there would be a number of challenges for implementation of this 1st generation cyclopropanation chemistry in the commercial environment. The issues identified to be addressed or mitigated with the next generation approach19 included: (1) Cost of the chiral catalyst; (2) Stability of the diazo compound and safety concerns; (3) The sensitivity of the catalyst to impurities in the substrates or reagents which could result in lower than desired ee’s or conversions; (4) Difficulty directly isolating material in high ee and yield from a reaction stream which was only 82-86% ee; (5) Specialized equipment and facilities to accommodate the diazo chemistry as well as the ozonolysis chemistry.

Conversion of the Chiral Cyclopropane to an Alkylating agent As mentioned previously, 23••(R)-AMBA was targeted because it could be readily converted to the desired cyclopropyl tosylate 17 through simple functional group conversions (Scheme 4). In our initial investigations, we were freebasing the salt and removing the (R)AMBA, however we quickly found that this extra step was not required. Esterification could be done directly via the in-situ generation of anhydrous HCl with trimethylsilylchloride


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(TMSCl) in EtOH. One of the challenges with the esterification is related to the fact that 23 has alcohol functionality and can self-esterify. Although this dimer was not impactful to quality as the ester is ultimately hydrolyzed, high levels would correspond to decreased yields, so two control points (concentration and temperature) were identified. The concentration of 9 L/kg was a good balance between minimizing dimer formation observed under high concentration reaction conditions as well as managing volumes for efficiency, throughput, and cost. Upon reaction completion, the bulk of the ethanol was removed via distillation, again to control volumes. By operating under vacuum, it was possible to maintain the temperature at ≤40 °C, thus minimizing further dimerization. After an aqueous workup which served to remove the one equivalent of (R)-AMBA, the final toluene solution was azeotropically distilled.

Development of the tosylation reaction quickly identified DABCO and tosyl

chloride (TsCl) as the ideal reagents. The DABCO not only served as a base, but it also activated the TsCl and promoted fast reactions (1–2 h).20 The toluene stream was treated with DABCO and TsCl to generate the desired 17 in high in-process yield (> 90%). Levels of the dimer were typically at 0.6-1.1 HPLC area percent. After an aqueous extraction, the mixture was azeotropically distilled under vacuum and the product was crystallized by charging heptane resulting in good purging of the dimer (99.6 HPLC area percent purity and < 100 ppm Pd.

Scheme 13. Modified Pd Catalyzed Intramolecular Direct Arylation



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BMS-791325 (1) Development and Synthesis Our investigation into the API (active pharmaceutical ingredient) step began with an evaluation of the Discovery conditions which employed 2-(1H-benzotriazole-1-yl)-1,1,3,3tetramethylaminium tetrafluoroborate (TBTU). We quickly found the TBTU chemistry was not amenable for scale-up as TBTU activation was accompanied by the formation of a number of impurities and the reaction mixtures were typically very highly colored (dark red-brown). Similar to our Discovery colleagues, we were also forced to employ preparative HPLC separation in order to obtain material of the requisite purity. While evaluating alternative coupling

reagents,

we

rapidly

narrowed

our

focus

to

1-ethyl-3-(3-

dimethylaminopropyl)carbodiimide (EDAC), 1-hydroxybenzotriazole (HOBt) and N,Ndiisopropylethylamine (DIPEA) conditions as there was extensive experience with many successful on-scale implementations of this chemistry.22

We first sought to determine

whether the EDAC/HOBt protocol was viable in terms of reactivity (note the sterically congested nature of the amide bond that is formed in this process) and impurity profile. We were pleased to find that the EDAC/HOBt chemistry (Scheme 14) was very well-suited for the preparation of BMS-791325 (1) via coupling of compound 13••Κ•EtOH and the bridged piperazine 14.

Scheme 14. EDAC and HOBt Coupling of 13••K and 14 to Produce BMS-791325 (1)


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Solvents that were assessed for the reaction were ACN, THF, DMF and 2methyltetrahydrofuran (2-MeTHF). ACN and THF were found to behave equally well, with both giving excellent HPLC purity profiles for the reaction at completion.

DMF was

satisfactory as a reaction solvent, but had the potential for prohibitively high volumes during the workup. 2-MeTHF did not perform well; specifically, the reaction proceeded at a slower rate when compared with acetonitrile and THF, presumably due to diminished solubility of the reaction components in 2-MeTHF. As discussed (vide supra), our initial development work and scale-up to produce intermediate 13 would inevitably contain high levels of water and EtOH (~10% by weight). In the API step, EtOH was shown to generate the ethyl ester impurity, and excess water caused reaction stalling. As a result, the API step required modification to contend with these contaminants. Azeotropic removal of EtOH and water was found to be an effective solution, and in this regard, acetonitrile proved to be far superior to THF in its ability to purge EtOH and water via azeotropic distillation. Thus, acetonitrile was selected as the reaction solvent. The primary impurities of concern in the amidation chemistry are the N-acylureas (compounds 43 and 44, Figure 9) that form via isomerization of the primary O-acylurea.23 These compounds are dead ends on the reaction pathway. They form in approximately a 2:1 ratio favoring isomer 44.

Implementation of HOBt, at loadings of > 0.5 equivalents,

minimized the formation of the N-acylurea impurities 43 and 44. From the perspective of



overall reaction purity/impurity profile and reaction rate (our targeted time to reaction completion was ≤ 10 hours), 1.15 equivalents of HOBt was found to be an optimal loading.

Figure 9. Observed N-Acylurea Impurities 43 and 44

The other components in this reaction (EDAC, DIPEA, and the bridged piperazine 34) were not subjected to extensive range-finding studies at the time of initial scale-up. EDAC and 14 loadings were simply adjusted to match the HOBt loading, and DIPEA was adjusted to approximate the stoichiometric HCl loading in the reaction system. In the reaction sequence, reagents were added in the following order: 14, HOBt, EDAC and DIPEA and delays of up to 1 hour in the addition of each reagent were proactively explored and found to be well tolerated. The workup is designed to minimize losses to the aqueous layers via maintaining aqueous pH within the isoelectric zone (~pH 4.5 to 7.0) for the API as the aqueous solubility of BMS-791325 increases dramatically outside of this range. For the reaction quench, a half saturated aqueous ammonium chloride solution was found to effectively extract residual EDAC, the corresponding EDAC-derived urea (EDCU), DIPEA and residual 14 into the aqueous layer. However, quenching the reaction directly into this solution led to significant loss of API due to the fact that the resulting aqueous pH is approximately 10. To solve this problem, five equivalents of acetic acid were added to the quench medium. As a result, the post-quench aqueous pH was typically about 5, a value that was within the desired isoelectric range. Implementation of the acetic acid charge had an additional benefit in that the aqueous pH does not fall significantly below the lower limit of the isoelectric range, due to the pKa of this acid. The phase split at pH~5 still removed most of the excess bridged piperadine 14, DIPEA-HCl, EDCU and residual EDAC.


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The derived organic layer still contained HOBt. In most amidation reactions, HOBt is removed by extraction with an aqueous base.24 In our case, this was not a viable approach due to the aqueous solubility of BMS-791325 at pH values greater than 7. As a result, we chose to extract the organic layer with pH 7 phosphate buffer (K2HPO4/KH2PO4), an aqueous medium that provided good extraction efficiency for HOBt while maintaining the aqueous pH in the desired range. It is important to note that the choice of co-solvent for the workup is critical to the effectiveness of the phosphate buffer extraction.

It was found that

isopropylacetate (IPAc) provided 2x better partitioning of HOBt into the aqueous layer when compared to ethyl acetate, and 10x better partitioning when compared to 2-MeTHF. Two extractions with 12.5 volumes of phosphate buffer led to consistent removal of HOBt to ≤ 2 HPLC area percent relative to BMS-791325. Following the workup sequence, the IPAc:ACN mixture is subjected to constantvolume distillation to effect solvent exchange to IPAc. Laboratory studies showed that ACN is typically reduced to non-detect levels25 prior to meeting a KF specification of ≤500 ppm. Therefore, we only monitored KF as a solvent exchange endpoint thus reducing the analytical burden. The KF specification of ≤500 ppm maximized the effectiveness of the upcoming clarification to remove residual inorganic impurities and other insolubles. A low water level at this stage was also desired because a favorable ethanol:water azeotrope is not accessible. Due to limited solubility of BMS-791325 in IPAc, we elected to solvent exchange to ethanol prior to clarification via a polish filtration. Once the solution was filtered, a 1.25N solution of HCl in ethanol was slowly charged, the batch was seeded and MTBE was charged over 1 h. The batch was held at 25 °C for 12 h, the resulting slurry of the BMS-791325••HCl salt was filtered and the cake was washed with an MTBE:ethanol mixture. This process, in four batches, afforded a total of 6.39 kg of 1 in 76.4% average yield26 in > 99.97 HPLC area percent purity and > 99.9 ee in 4 batches. A subsequent delivery afforded 68.9 kg in 78.5% average yield27 in >99.6 HPLC area percent purity and >99.9 % ee in 3 batches. One of the challenges with the crystallization of BMS-791325••HCl is the fact that up to 16% of the product often remained in the mother liquors post-crystallization despite all efforts to optimize anti-solvent ratio and temperature of the crystallization.

This was

identified as something that would have to be addressed in subsequent development work.



Conclusions In summary, we describe an efficient, highly convergent synthesis (Scheme 15) of BMS-791325•HCl (1) which obviated the need for an end game chiral resolution. The chiral cyclopropane fragment (23•(R)-AMBA) was prepared in a highly telescoped fashion (7 linear steps and only 2 isolations) in 26 % yield via asymmetric catalysis using the Rh2(S-DOSP)4 catalyst. This is the first known kilogram-scale demonstration of the Davies methodology. The indole fragment was prepared in 6 steps and 3 isolations in 53% yield. After the cyclopropane fragment is activated and indole is N-alkylated, the palladium catalyzed cyclization provides quick and direct access to the 7-membered ring. Hydrolysis and coupling affords BMS-791325•HCl (1) in high quality. The telescoped endgame is conducted in 6 steps and 3 isolations in 32% yield. The overall process has been demonstrated on multikilogram scale and was considered fit-for-purpose successfully fulfilling initial toxicological and clinical requirements and laying the ground work for development of the commercial process. The target was prepared in 12 linear steps with 5 isolations in an overall yield of 8.2%. Scheme 15. Complete Linear Synthesis of BMS-791325••HCl (1)


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Experimental Section All reactions were performed under a nitrogen atmosphere unless otherwise noted. Reagents were used as received from vendors, unless otherwise noted. All reagents purchased from vendors were used as received unless otherwise indicated. NMR spectra were recorded on a Bruker DRX-400 or AV III 600 instrument and are referenced to residual undeuterated solvents. The following abbreviations are used to explain multiplicities: br = broad, s = singlet, d = doublet, t = triplet, q = quartet, m = multiplet. High-resolution mass spectrometry (HRMS) was performed on an Agilent 6230B TOF instrument. Preparation of methyl 3-cyclohexyl-1H-indole-6-carboxylate (3). To a reactor was charged methyl indole-6-carboxylate (2) (85.0 kg, 485.2 mol, 1.0 equiv.) and toluene (206.0 kg). The resulting slurry was stirred for 15 min at 20 oC. To this mixture was charged cyclohexanone (94.0 kg, 957.7 mol, 2.0 equiv.) while maintaining the temperature at 20 oC. To the solution was charged Et3SiH (172.0 L, 1479.2 mol, 3.0 equiv.) in 3 portions. The mixture was cooled to 0 oC. Trifluoroacetic acid (172.0 kg, 1508.5 mol, 3.1 equiv.) was charged over 90 minutes controlling the temperature below 30 °C (note: this addition is exothermic and requires active cooling). After the addition was complete, the



reaction was maintained at 20 °C and monitored for reaction completion (starting material

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