Practical Synthesis of MDM2 Antagonist RG7388. Part 2: Development

Article pubs.acs.org/OPRD

Practical Synthesis of MDM2 Antagonist RG7388. Part 2: Development of the Cu(I) Catalyzed [3 + 2] Asymmetric Cycloaddition Process for the Manufacture of Idasanutlin Gösta Rimmler,† Andre Alker,‡ Marcello Bosco,† Ralph Diodone,† Dan Fishlock,† Stefan Hildbrand,† Bernd Kuhn,‡ Christian Moessner,† Carsten Peters,† Pankaj D. Rege,*,† and Markus Schantz† †

Small Molecules Technical Development, F. Hoffmann-La Roche Ltd., 4070 Basel, Switzerland Therapeutic Modalities, Roche Pharma Research and Early Development, Roche Innovation Center Basel, F. Hoffmann-La Roche Ltd., 4070 Basel, Switzerland

‡

S Supporting Information *

ABSTRACT: A concise catalytic asymmetric synthesis of idasanutlin (1) was developed in which the key pyrrolidine core, containing four contiguous stereocenters, was constructed via a Ag/MeOBIPHEP promoted [3 + 2] cycloaddition reaction. Further development of the [3 + 2] cycloaddition reaction resulted in an improvement in diastereoselectivity and enantioselectivity by changing the catalyst system to Cu(I)/BINAP. While producing equivalent high quality API, the copper(I) catalyzed process not only increased the overall yield but also demonstrated benefit with respect to cycle times, waste streams, and processability. The optimized copper(I) catalyzed process has been used to prepare more than 1500 kg of idasanutlin (1). KEYWORDS: [3 + 2] cycloaddition, asymmetric, idasanutlin, copper-catalyzed, silver-catalyzed

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literature2 and provides a logical synthetic disconnection for 1. The medicinal chemistry synthesis1a employed this synthetic disconnection in the form of a racemic [3 + 2] cycloaddition reaction to construct the pyrrolidine core for structure−activity relationship (SAR) studies and initial preclinical supplies. The medicinal chemistry route1a was not suitable for the preparation of larger amounts due to the reasons outlined in Part 1 of this work (10.1021/acs.oprd.6b00320).3 Therefore, upon selection of 1 as a clinical candidate, process research activities commenced to meet the demands of clinical trial supply and eventually develop a commercial manufacturing process. These efforts initially resulted in an asymmetric cycloaddition reaction using Ag(OAc) with (R)-MeOBIPHEP4 as a chiral ligand.5,6 Although the Ag-catalyzed process gave 1 in good quality on a 100 kg scale, a more efficient process was required to support the increasing API demand. Continuing development activities which were targeted to address limitations of the Ag-catalyzed process yielded a Cu(I)/(R)BINAP7 variant of the [3 + 2] cycloaddition reaction. Herein we report the development of a manufacturing process for 1 which employs a Cu(I)-catalyzed asymmetric [3 + 2] cycloaddition reaction. The optimized Cu(I)-catalyzed synthesis represents the industrial process, which to date has been used to manufacture more than 1500 kg of 1.

INTRODUCTION Idasanutlin (RG7388, 1, Figure 1) is a mouse double minute 2 (MDM2) homologue protein antagonist discovered by Roche.

Figure 1. Left: Structure and absolute stereochemistry of idasanutlin (1). Right: X-ray crystal structure of 1.

MDM2 (also known as E3 ubiquitin-protein ligase MDM2) is an important negative regulator of the p53 tumor suppressor protein. Idasanutlin (1) is designed to bind to MDM2 to potentially prevent the p53−MDM2 interaction and thereby result in activation of p53.1 Idasanutlin (1) shows efficacy in xenograft models and clinical trials of this compound are ongoing. Rapid and durable responses have been seen in patients suffering from acute myeloid leukemia (AML) in response to treatment with 1 in combination with cytarabine, and stable disease with evidence of p53 activation has been observed in early solid tumor studies. As depicted in Figure 1, 1 contains a congested pyrrolidine carboxamide core with four contiguous stereocenters with an all anti absolute configuration (2R,3S,4R,5S). Azomethine ylidebased [3 + 2] cycloaddition reaction with appropriately substituted electron-deficient olefins is well-established in the © XXXX American Chemical Society

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RESULTS AND DISCUSSION Synthesis of the Starting Materials for the [3 + 2] Cycloaddition Reaction. The imine 4 was prepared by the condensation reaction between 4-(2-amino-acetylamino)-3methoxy-benzoic acid ethyl ester hydrochloride (2) and 3,3Received: September 26, 2016 Published: October 31, 2016 A

DOI: 10.1021/acs.oprd.6b00319 Org. Process Res. Dev. XXXX, XXX, XXX−XXX

Organic Process Research & Development

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Scheme 1. Preparation of 4

impurity was not depleted in the downstream chemistry. Further optimization of reaction conditions showed that a catalytic amount of 30% solution of sodium methoxide in methanol facilitated the reaction at ambient temperature within only 3 h and the methoxy-substituted impurity was not formed. Thus, upon slow addition of the 30% methanolic solution of sodium methoxide (5 mol %) to a 1:1 mixture of 5 and 6 in ethanol containing 0.5% w/w water, the desired product 7 precipitated from the reaction mixture and was isolated by filtration. The trace amounts of water in the mixture were found to significantly improve the filterability of 7. The only observed side product, with levels up to 6%, was the corresponding E-silbene 8 which does not precipitate from the reaction mixture and was efficiently removed to the mother liquor during the filtration of 7. This optimized manufacturing process for 7 resulted in a yield of 91% with purity of 99.9%. This process was executed on up to 160 kg batch size of 6 to produce 11 batches of 7 (1818 kg in total). Metal-Catalyzed Asymmetric Cycloaddition Process. In the original medicinal chemistry route, the pyrrolidine core in 1 was constructed in a racemic form using stoichiometric AgF promoted [3 + 2] cycloaddition reaction.1a The enantiopure 1 was obtained employing chiral chromatography. Although that route was not suitable to support upcoming clinical demands, it provided the basis for the initial process development work. Analogous to the medicinal chemistry route, the pyrrolidine ring in compound 9c was constructed

dimethyl butyraldehyde (3) (Scheme 1). The reaction was performed in the presence of triethylamine in methyl-tertbutylether (MTBE) at 20−25 °C within 6 h. After separation of precipitated NEt3·HCl by filtration, the imine 4 was obtained by crystallization, following a solvent exchange to n-heptane, in a yield of 94% with excellent quality (99.9% purity). Six batches have been prepared starting with 110 kg of 2 to produce 703 kg of 4. Stilbene 7 was prepared by Knoevenagel condensation between aldehyde 5 and 2-(4-chloro-2-fluorophenyl)acetonitrile (6) (Scheme 2). Originally a stoichiometric amount Scheme 2. Preparation of 7

of sodium methoxide in methanol at 50 °C for 3 h was used to effect the condensation. However, the use of 5 mol % of NaOH (50% w/w aqueous solution) instead of sodium methoxide was also effective. Unfortunately, the methoxy substitution of fluorine (ortho to the chlorine) in the aromatic ring of product 7 was observed when the reaction mixture was stirred for an extended period of time. Moreover this methoxy-substituted

Scheme 3. Silver-Catalyzed Asymmetric Synthesis of 1 (Absolute Stereochemistry Assigned Based on X-ray Crystal Structure Analysis, Vide Infra)a

Reagents and conditions: (a) AgOAc (1.0 mol %), 12 (1.1 mol %), 2-MeTHF. (b) Anhydrous LiOH (milled), 60 °C, then n-heptane, 97% yield. (c) i. LiOH, 2-PrOH, H2O, 70 °C; ii. filter-off lithium salt of rac-1; iii. AcOH, H2O, 70 °C, 45% yield, er >99:1. a

B



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Scheme 4. Cu(I)-Catalyzed Asymmetric [2 + 3] Cycloaddition Reactiona

a

Reagents and conditions: (a) CuOAc (0.50 mol %), 13 (0.53 mol %), 2-MeTHF, Et3N. (b) i. THF, ethanol, NaOH(aq). ii. cryst from 2-PrOH, H2O, 78% yield, er 93:7.

using an asymmetric version of the Ag-catalyzed [3 + 2] cycloaddition reaction. AgOAc with bidentate phosphine ligands was found to provide the best selectivity among all of the systems investigated, and eventually AgOAc/(R)-MeOBIPHEP catalyst system was chosen.6 In the presence of 1.0 mol % of AgOAc and 1.1 mol % (R)MeOBIPHEP 12,4 the reaction of 4 and 7 in MeTHF at 0 °C afforded a mixture of pyrrolidine diastereoisomers 9a, 9b, and 9c (total 75% area), the ring-opened Michael addition product 10 (11% area) and a mixture of imine-hydrolyzed Michael addition products 11 (15% area) after 16−24 h (Scheme 3). Upon treatment of this reaction mixture with micronized anhydrous lithium hydroxide the pyrrolidine diastereoisomers 9a and 9b, as well as the Michael addition product 10, were converted to the desired ester 9c (er 84:16) which was isolated by crystallization from n-heptane/MeTHF. Hydrolysis of 9c with lithium hydroxide in isopropanol followed by the removal of the insoluble racemate (as lithium salt) by filtration provided enantioenriched filtrate. Subsequent acidification of the filtrate with acetic acid and isolation by crystallization from isopropanol/water produced 1 with an er of >99:1 in 44% overall yield from 7. The Ag-catalyzed process outlined above was successfully employed to produce more than 100 kg of API to support ongoing clinical trials. Despite the successful supply campaigns, this process was not considered appropriate for commercial manufacturing for the following reasons: (a) the efficient and complete epimerization can only be achieved with finely milled lithium hydroxide, making an extra milling step necessary, (b) silver oxide and other very fine particulates precipitate during the hydrolysis process leading to serious filter or centrifuge clogging during racemate filtration, (c) intermediate reactor cleaning is necessary due to formation of silver deposites on the reactor walls, and (d) the modest enantioselectivity of the

cycloaddition reaction leads to an erosion of the yield due to enantioenrichment via racemate removal. The limitations of the Ag-catalyzed approach warranted continuation of development efforts in search of a more robust scalable process for commercial manufacturing. Along with Agbased catalysts, Cu-based catalysts are widely employed in asymmetric [3 + 2] cycloaddition.2 In an effort to address the disadvantages of the Ag-catalyzed process and primarily to improve the diastereo- and enantioselectivity of the cycloaddition reaction and issues related to precipitation of Ag salts, the use of Cu catalysts was investigated. As reported by Shu et al.,3 the initial focus of this investigation was Cu(II)-based catalyst systems, and a process using Cu(OAc)2/(R)-BINAP (1.3% catalyst loading) delivered 1 with an er of approximately 94:6 on a 30 g scale. Further enantioenrichment was achieved by the filtration of the racemate from EtOAc/THF, followed by the crystallization of idasanutlin (1). During further development, the Cu(OAc)2 catalytic system was found to give inconsistent results when starting materials (4 and 7) of representative quality were employed. Experiments on 1 kg scale confirmed the lack of robustness of the cycloaddition step. These reactions were carried out at room temperature with 1.0 mol % of Cu(OAc)2 and (R)-BINAP in 2MeTHF. The consumption of stilbene 7 required 16 h, and the contents of diastereoisomers 9a−c in the reaction mixture varied substantially between single experiments (70−80%) along with side products 11 in the range of 5−15%. The ringopened Michael addition product 10 was present in 1−2%. The ratio between 9a−c (10:0.7:0.85) remained fairly constant. The variability of side product formation, extended reaction times of the cycloaddition reaction, and occasional stalling of the reaction merited more thorough investigation of the Cucatalyzed process. During their studies of Cu(II)-catalyzed systems, Shu et al. had noted that the Cu(I)-based catalyst system also provided C



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Scheme 5. Z- and E-Stilbene, 7 and 8, and Resulting Cycloaddition Product Mixtures

encouraging results, so CuOAc was tested as an alternative to Cu(OAc)2 in the [3 + 2] cycloaddition reaction. Using only 0.50 mol % CuOAc in combination with 0.53 mol % (R)BINAP led to smooth conversion of stilbene 3 within 6 h, and the overall amount of diastereomers 9a−c was raised to 90− 93% with a fairly constant ratio between the single diastereoisomers 9a (74−76%), 9b (5−6%), and 9c (7−8%) (Scheme 4). The formation of side product diastereoisomers 11 could be reduced to 2−4% and the ring opened Michael addition product 10 was formed in 3−4%. In contrast to Cu(OAc)2, the CuOAc system did not require any premixing or aging of the metal−ligand complex. The selectivity of the cycloaddition reaction was found to be very robust, and within the temperature range of 0−40 °C no change in selectivity was observed. To better understand the Cu-catalyzed reaction, Cu-chiral ligand catalyst systems were analyzed using 31P NMR.8 These experiments showed that the complexation of Cu(I) with (R)BINAP occurs immediately at room temperature. On the other hand, Cu(II) system showed significantly less complexation of the chiral phosphine ligand with the metal, even after 80 h. Since the CuOAc/(R)-BINAP system provided robust conditions and desired selectivity and (R)-BINAP is a moderately priced commercially available ligand, a repetition of the chiral ligand screening was not initiated, and the CuOAc/(R)-BINAP system was therefore chosen for further development. Selectivity in [3 + 2] Cycloaddition Reaction. Copperbased catalysts are reported to deliver exo selective [3 + 2] cycloaddition products of metalated azomethine ylides that lead to 4,5-trans substituted pyrrolidines.2 Indeed, the relative stereochemistry between electron-withdrawing group (CN) at C4 and the neopentyl group at C5 in the main product 9a from the cycloaddition reaction is trans. Therefore, the [3 + 2] cycloaddition reaction under our conditions can be presumed to proceed analogously as reported by Garner et al.2d The endo product 9b was observed in 5−6%, and another product observed in minor amounts 7−8% is the diastereomer 9c. The latter can be an outcome of a nonconcerted stepwise mechanism (Michael addition followed by Mannich ring closure) via a zwitterionic intermediate as postulated by Garner et al.2d Further evaluation of the stereoisomeric outcome of this cycloaddition addressed the configuration of the dipolarophile. The Z- and E-stilbenes, 7 and 8, would lead to two distinct series of diastereoisomers differing in the relative stereochemistry of the two aromatic rings in 3,4-position of the cycloaddition product (Scheme 5). The Z-stilbene 7 generates pyrrolidine stereoisomers 9 with the desired anti orientation of the aromatic rings, whereas the corresponding E-stilbene 89 leads to product 14 with undesired 3,4-cis orientation. It could be demonstrated that the Z-stilbene 7 is configurationally stable under the reaction conditions, and as a consequence no isomers with 3,4-syn configuration were detected in the reaction mixture. Therefore, the formation of only four diastereomers must be considered. Furthermore, for unequivocal structural assignment diastereoisomers 9a−c and ring-opened product 1010 (Scheme 4) were isolated by preparative chromatography from the cycloaddition mixture and characterized by NMR. The isolated diastereoisomers were further subjected to preparative chiral HPLC separation, and then each enantiomer was fully characterized by NMR and chiral HPLC. In addition, the

absolute configuration for at least one of the two enantiomers of each diastereoisomer 9a, 9b, and 9c were determined by Xray crystallography (see Supporting Information).11 It was found that the main product 9a was formed with high enantioselectivity of 99:1 (with RSRR as the major isomer), while negligible enantioselectivity (40:60) was observed for the endo-product 9b. The isomer 9c bearing the same configuration as idasanutlin was formed with moderate enantioselectivity of 86:14 (with RSRS as the major isomer). The enantiomeric purity of the ring-opened product 10 could not be determined at the ester stage, but 10 was also converted to idasanutlin (1) during hydrolysis/isomerization (vide infra) with er >99:1, indicating that the stereocenter at C3 was formed with a high enantioselectivity in the cycloaddition. The undesired constitutional isomer (ethyl ester of 15) could only be generated in trace amounts (99.9:0.1) in 79% yield. Absolute stereochemistry of 1 has been confirmed by single crystal X-ray analysis (see Supporting Information for details). E



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THF solution of 1 (14% w/w). Crystallization of 1 starts after addition of approximately 1/3 of the aqueous mixture. The isolated acetonitrile solvate (Form IX) of 1 is dried at 80 °C under vacuum to obtain 1 in 93% yield. The acetonitrile in the solvate is easily removed by drying, resulting in the solvate free Form III. As listed in Table 1, Form III is a slightly hygroscopic polymorph of 1 and is reliably produced during clinical supply. The commercial manufacturing process for 1 is shown in Scheme 6. Genotoxic Impurity Control. Accompanying process development activities, starting materials and their observed impurities, reagents, solvents, process intermediates, observed impurities, degradation products, and reasonably expected reaction byproducts involved in the synthesis of 1 were assessed for genotoxicity risk. The assessment included in silico evaluation (using two complementary methods, DEREK 10.0.2 for Windows and Leadscope Model, according to ICH M7 guidance: Assessment and Control of DNA Reactive (Mutagenic) Impurities in Pharmaceuticals to Limit Potential Carcinogenic Risk), Ames testing, and analytical testing. Two compounds, 16 and its ethyl ester 17, produced structural alerts for mutagenicity during in silico evaluation and were subsequently subjected to Ames testing. As outlined in Scheme 7, there are at least two potential pathways for formation of 16 and 17. Compound 16 is not only a byproduct during the saponification reaction but also a degradation product of 1 formed by hydrolysis of the amide bond, and ethyl ester 17 is a potential degradation product of imine 4 and a byproduct in the hydrolysis/isomerization reaction. The saponification of 17 under the reaction conditions was found to be significantly slower as compared to the saponification of intermediate products 9a−c. Therefore, both 17 and 16 are typically detected in the reaction mixture after the hydrolysis in only up to 2% and 1%, respectively. These amounts are effectively purged during the crystallization of

Although the enantiomeric excess in compounds 9a, 9b, and 9c obtained via the Ag-catalyzed process is similar to the Cu(I)catalyzed process, significantly less racemate is formed in the Cu(I)-catalyzed process due to the better diastereoselectivity in the [3 + 2] cycloaddition reaction. This results in an improvement in the overall yield from 44% to 61%. Furthermore, the aqueous waste streams generated in the Cuprocess are biodegradable, thus reducing the environmental impact. Final Recrystallization of Idasanutlin. Several polymorphs, pseudopolymorphs, hydrates, and an amorphous form of 1 are known, and the most relevant forms are summarized in Table 1. The access to the different polymorphic Table 1. Summary of the Most Important Polymorphs and Pseudo-Polymorphs of 1 and Thermal Behavior form Form I Form II Form III Form V Form IX amorphous

thermal events endothermic solid−solid conversion around 270 °C followed by melting as Form III around 280 °C (melting) around 282 °C (melting) between 25 and 140 °C loss of weight (dehydration) and conversion to Form I desolvation and conversion into Form III followed by melting of Form III glass transition around 146 °C

solid form polymorph polymorph polymorph hydrate acetonitrile solvate amorphous

forms is mainly determined by the solvate present in the crystallization process. The use of acetonitrile in the crystallization process of 1 (vide supra) results in the formation of a corresponding low soluble acetonitrile solvate. The acetonitrile in the solvate can easily be removed by drying, resulting in the solvate free Form III. The final recrystallization process is the following: a water− acetonitrile mixture (1:2.5% w/w) is added to a polish-filtered

Scheme 6. Commercial Manufacturing Process for Idasanutlin (1)

F



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Scheme 7. Pathways for Formation of Genotoxic Impurities 16 and 17

Table 2. Theoretical and Measured Purge Factors for 16 and 17 GTI

step no.

reactivity

solubility

volatility

theoretical purge factor

measured purge factor

16

2a/2b 3 4

1 1 1

10 10 10

1 1 1

17

2a/2b 3 4

3 1 1

10 10 10

1 1 1

10 10 10 1000 30 10 10 3000

>150 >15 >330 >7 × 105 >60 >50 >80 >2.4 × 105

process

process

starting materials, the exceptionally selective and consistent cycloaddition/isomerization sequence using a cheap chiral Cu(I) catalyst system, the overall robustness and the high throughput in production, and the low environmental impact. The process ensures control of impurity formation and purging (including isomers and genotoxic impurities) and the crystallization of a desired polymorphic form. This process has been utilized to produce more than 1500 kg of idasanutlin (1) to date to support ongoing clinical studies.

enantioenriched 1 to below the reporting limit of the analytical method.18 In the racemate removal step, the main purging effect was observed during the crystallization of 1. The filter cake containing racemic 1 did not show significant levels for both impurities. Furthermore, the final recrystallization step provides good purging for both genotoxic compounds. Analysis of enantiopure 1 consistently proved both the compounds to be below 10 ppm. Despite the low levels routinely obtained during manufacturing, the purging powers of both crystallizations were investigated to correlate the experimental purge factors and the theoretical purge factors calculated following the procedure by Teasdale et al.19 The experimental purge factors were determined using data from spiking experiments. For the crystallization of enantioenriched 1, both compounds were spiked to the mixture after the hydrolysis just before the neutralization. Purging factors of >150 for 16 and >60 for 17 were observed. In the racemate removal step as well as the final crystallization, both compounds were spiked to a level of 1000 ppm to 1. As anticipated the ethyl ester 17 is significantly better depleted than 16 in the racemate removal step showing a purge factor of 50 compared to 15 for 16. The final recrystallization step proved to have a very high purging power for 17 due to the presence of water in the crystallization mixture. The overall purge factors (experimental) were determined to be >7 × 105 and >2.4 × 105 for compounds 16 and 17, respectively. As shown in Table 2, these values are significantly higher than the theoretical purge factors of 1000 and 3000 for compound 16 and 17, respectively. Therefore, in the case provided here, the theoretical purge factor underestimates the real purging effect of the crystallization process and provides a worst case estimate for the impurity control.

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EXPERIMENTAL DETAILS General Remarks. Reagents and solvents were used as received from commercial suppliers. All reactions were carried out in standard pilot plant equipment consisting of a setup of three connected 2500 L reactors with variable speed agitator, 50 to 150 °C jacket temperature range, and a 20 mbar vacuum to 1050 mbar pressure rating. Solids were separated on a 100 cm basket centrifuge. All reactions were carried out under a nitrogen atmosphere. Nitrogen was also routinely used to break vacuums for safety reasons. The four starting materials 2, 3, 5, and 6 and the intermediates 4, 7, and 9c were tested in the central safety laboratory of F. Hoffmann-La Roche AG, and the chemical transformations were investigated via reaction calorimetric measurements. Common calorimetric methods such as differential scanning calorimetry (DSC), accelerating rate calorimetry (ARC), and reaction calorimetry were used to determine safety of reaction mixtures, handling of reactants, reagents, and solvents. For each step reported in this publication, the processes were deemed safe based on data from safety analysis. All of the GC and HPLC data were generated using calibrated equipment. Unless otherwise specified, all reported purity and assay values are measured using HPLC methods. MS and NMR spectra were measured by the central analytical service of F. Hoffmann-La Roche AG. The 1H NMR and 13C NMR spectra were measured on Bruker 600 MHz NMR spectrometers at 600, 400, and 150 MHz, respectively. The

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CONCLUSION A short and high-yielding manufacturing process for idasanutlin (1) was developed. The salient features of the process include the convergent route using readily accessible noncomplex G



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(220 kg) at 4 °C, a previously prepared thin suspension of (R)MeOBIPHEP 12 (1.01 kg; 1.73 mol) and AgOAc (0.26 kg; 1.56 mol) in 2-MeTHF (8.6 kg) was added and the feeding tank rinsed with 2-MeTHF (4 kg) into the vessel. Afterward the reaction mixture was stirred at 2−4 °C for 16 h. After successful IPC, the reaction mixture was warmed to 23 °C and diluted by the addition of 2-MeTHF (36 kg). Then a suspension of previously wet milled LiOH20 in 2-MeTHF was added, and the reaction mixture was heated to 65 °C and stirred at this temperature for 19 h. After complete conversion, n-heptane (460 kg) was added within 50 min, and afterward the suspension was cooled to 15 °C at a rate of 0.15 °C/min and stirred at target temperature for 4 h. The product was collected by centrifugation and washed with a mixture of nheptane (156 kg) and 2-MeTHF (98 kg). It was dried to constant weight (55 °C, 7 mbar, 5 h) to furnish 103.6 kg of enantiomerically enriched 4-{[(2R,3S,4R,5S)-4-(4-chloro-2-fluorophenyl)-3-(3-chloro-2-fluorophenyl)-4-cyano-5-(2,2-dimethylpropyl)-pyrrolidine-2-carbonyl]-amino}-3-methoxybenzoic acid ethyl ester (9c) (94.9% w/w, er: 78.5:21.5, 97% yield) as an off-white solid. Peparation of Idasanutlin (1) from Ester (9c). The reactor was charged with 4-{[(2R,3S,4R,5S)-4-(4-chloro-2-fluorophenyl)-3-(3-chloro-2-fluorophenyl)-4-cyano-5-(2,2-dimethylpropyl)-pyrrolidine-2-carbonyl]-amino}-3-methoxybenzoic acid ethyl ester (9c) (128.0 kg; 198.6 mol) followed by isopropanol (447 kg) at 20−25 °C, and to the resulting suspension a previously prepared solution of lithium hydroxide (0.97 kg; 40.5 mol) in water (109 kg) was added followed by the addition of water (19 kg) for rinsing of the feed vessel. The thin suspension was heated to 63−67 °C and stirred at this temperature for 6 h. After completion of the reaction, the reaction mixture was transferred to a crystallization vessel, and the reactor and lines were rinsed with isopropanol (45 kg). The thin suspension was cooled within 3 h to 15−20 °C and stirred for an additional 8 h at this temperature. The solids were collected by centrifugation, and the filter cake was rinsed with isopropanol (507 kg). The combined filtrates were afterward polish filtered and warmed to 63−67 °C. At this temperature, glacial acetic acid (26.4 kg; 439.6 mol) was added followed by water (8.2 kg). After adjusting the pH to

Practical Synthesis of MDM2 Antagonist RG7388. Part 2: Development

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