- Efficient bioreductive production of (S)-N-Boc-3-hydroxypiperidine using ketoreductase ChKRED03
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Ibrutinib is an anticancer drug targeting B-cell malignancies. The key chiral intermediate for ibrutinib synthesis is the alcohol (S)-N-Boc-3-hydroxypiperidine ((S)-NBHP), which can be produced via ketoreductase (KRED)-catalyzed bioreduction. After screening a small inventory of 27 KREDs mined from the genome of Chryseobacterium sp. CA49, ChKRED03 was selected as the best performer, leading to the complete conversion of 100 g substrate/L within 10 h to yield (S)-NBHP with high enantiomeric excess (> 99% ee). The enzyme was NADPH dependent, and the highest enzymatic activity was observed at 30 °C in potassium phosphate buffer (pH 7.0). At a substrate/catalyst ratio of 66.7 (w/w), ChKRED03 catalyzed the complete conversion of 200 g/L substrate within 3 h to yield (S)-NBHP with >99% ee, demonstrating great potential for industrial application.
- Xu, Guang-Peng,Wang, Hai-Bo,Wu, Zhong-Liu
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Read Online
- Characterization of a carbonyl reductase from Rhodococcus erythropolis WZ010 and its variant Y54F for asymmetric synthesis of (S)-N-Boc-3-hydroxypiperidine
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The recombinant carbonyl reductase from Rhodococcus erythropolis WZ010 (ReCR) demonstrated strict (S)-stereoselectivity and catalyzed the irreversible reduction of N-Boc-3-piperidone (NBPO) to (S)-N-Boc-3-hydroxypiperidine [(S)-NBHP], a key chiral intermediate in the synthesis of ibrutinib. The NAD(H)-specific enzyme was active within broad ranges of pH and temperature and had remarkable activity in the presence of higher concentration of organic solvents. The amino acid residue at position 54 was critical for the activity and the substitution of Tyr54 to Phe significantly enhanced the catalytic efficiency of ReCR. The kcat/Km values of ReCR Y54F for NBPO, (R/S)-2-octanol, and 2-propanol were 49.17 s?1 mM?1, 56.56 s?1 mM?1, and 20.69 s?1 mM?1, respectively. In addition, the (S)-NBHP yield was as high as 95.92% when whole cells of E. coli overexpressing ReCR variant Y54F catalyzed the asymmetric reduction of 1.5 M NBPO for 12 h in the aqueous/(R/S)-2-octanol biphasic system, demonstrating the great potential of ReCR variant Y54F for practical applications.
- Ying, Xiangxian,Zhang, Jie,Wang, Can,Huang, Meijuan,Ji, Yuting,Cheng, Feng,Yu, Meilan,Wang, Zhao,Ying, Meirong
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Read Online
- Efficient synthesis of Ibrutinib chiral intermediate in high space-time yield by recombinant E. coli co-expressing alcohol dehydrogenase and glucose dehydrogenase
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The production of (S)-N-boc-3-hydroxy piperidine (NBHP) via asymmetric bioreduction of 1-boc-3-piperidinone with reductase is impeded by the need for expensive coenzymes NAD(P)H. In order to regenerate the coenzyme in situ, the gene of alcohol dehydrogenase from Thermoanaerobacter brockii and glucose dehydrogenase from Bacillus subtilis were ligated into the multiple cloning sites of pRSFDuet-1 plasmid to construct the recombinant Escherichia BL21 (DE3) that co-expressing alcohol dehydrogenase and glucose dehydrogenase. Different culture conditions including the medium composition, inducer and pH etc were systematically investigated to improve the enzyme production. The enzyme activity was increased more than 11-fold under optimal culture condition, from 12.7 to 139.8 U L?1. In the further work, the asymmetric reduction of 1-boc-3-piperidinone by whole cells of recombinant E. coli was systematic optimized to increase the substrate concentration and reaction efficiency. At last, S-NBHP (>99% ee) was prepared at 500 mM substrate concentration without external addition of cofactors. The conversion of S-NBHP reached 96.2% within merely 3 h, corresponding a high space-time yield around 774 g L?1 d?1. All these results demonstrated the potential of recombinant E. coli BL21 (DE3) coupled expressing alcohol dehydrogenase and glucose dehydrogenase for efficient synthesis of S-NBHP.
- Chen, Yitong,Ma, Baodi,Cao, Songshuang,Wu, Xiaomei,Xu, Yi
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Read Online
- Engineering an Alcohol Dehydrogenase from Kluyveromyces polyspora for Efficient Synthesis of Ibrutinib Intermediate
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(S)-N-Boc-3-hydroxypiperidine [(S)-NBHP] is a key intermediate for the synthesis of mantle cell lymphoma drug, ibrutinib. Here, KpADH, an alcohol dehydrogenase from Kluyveromyces polyspora, exhibits evolutionary potential in the asymmetric reduction of N-Boc-3-piperidone (NBPO) to (S)-NBHP. By screening key residues in substrate binding pocket of KpADH, an excellent variant Y127W was obtained with 6-fold improved activity of 119.3 U mg?1, 1.8-fold enhanced half-life of 147 h and strict S-stereoselectivity (>99% ee). When catalyzed by Y127W, a complete conversion of 600 g L?1 NBPO was achieved at a substrate to catalyst ratio (S/C) of 30 in 10 h. Based on crystal-structure of Y127W, molecular docking and dynamic simulations reveal hydrogen bonds formed between W127 and Boc group of NBPO, as well as improved structural stability mainly contribute to the increased catalytic activity and stereoselectivity of Y127W. This study offers guidance for engineering ADHs for biosynthesis of chiral heterocyclic alcohols, and provides insights into mechanisms in catalytic activity and stereoselectivity toward carbonyl-containing heterocyclic substrates. (Figure presented.).
- Wu, Yanfei,Zhou, Jieyu,Ni, Jie,Zhu, Cheng,Sun, Zewen,Xu, Guochao,Ni, Ye
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- Preparation method of N-tert-butyloxycarbonyl-3-piperidone and derivative thereof
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The invention discloses a preparation method of N-tert-butyloxycarbonyl-3-piperidone, and the method comprises the following step: oxidizing N-tert-butyloxycarbonyl-3-hydroxypiperidine by virtue of aNaDCC-TEMPO system so as to generate N-tert-butyloxycarbonyl-3-piperidone. The invention also discloses a method for preparing (S)-N-tert-butyloxycarbonyl-3-hydroxypiperidine through an oxidation reaction and a reduction reaction by virtue of the NaDCC-TEMPO system. The method has the advantages of simple operation steps, high conversion rate and high product purity.
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Paragraph 0063; 0073-0078
(2020/07/21)
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- Preparation method of (S)-1-tert-butyloxycarbonyl-3-hydroxypiperidine
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The invention discloses a preparation method of (S)-1-tert-butyloxycarbonyl-3-hydroxypiperidine, and relates to the technical field of biological pharmacy. The preparation method comprises the following steps: 1, preparing 3-hydroxypyridine (2) as a raw material; 2, preparing racemic 3-hydroxypiperidine (3) from 3-hydroxypyridine (2); 3, preparing a derivative (2S, 3S)-N-(4-chlorphenyl)-2, 3-dihydroxy succinamic acid (4) from racemic 3-hydroxypiperidine (3) and D-tartaric acid; 4, carrying out resolution reaction on the derivative (2S, 3S)-N-(4-chlorphenyl)-2, 3-dihydroxy succinamic acid (4) prepared from D-tartaric acid to obtain a (S)-hydroxypiperidine salt (5); and 5, subjecting an obtained mixture to extraction, concentration and crystallization, to prepare a (S)-1-tert-butyloxycarbonyl-3-hydroxypiperidine salt (5). The preparation process has the advantages that the synthesis steps are reduced, the yield is increased, an adopted chiral resolving agent can be recycled, and the preparation process is suitable for industrial production.
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- PROCESSES AND INTERMEDIATES FOR PREPARING A BTK INHIBITOR
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Disclosed is a process for the preparation of certain intermediates, e.g. the following compound: (I) which intermediate and processes are useful in the preparation of a BTK inhibitor, such as ibrutinib.
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Page/Page column 15; 16
(2018/04/21)
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- Methodology Development in Directed Evolution: Exploring Options when Applying Triple-Code Saturation Mutagenesis
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Directed evolution of stereo- or regioselective enzymes as catalysts in asymmetric transformations is of particular interest in organic synthesis. Upon evolving these biocatalysts, screening is the bottleneck. To beat the numbers problem most effectively, methods and strategies for building “small but smart” mutant libraries have been developed. Herein, we compared two different strategies regarding the application of triple-code saturation mutagenesis (TCSM) at multiresidue sites of the Thermoanaerobacter brockii alcohol dehydrogenase by using distinct reduced amino-acid alphabets. By using the synthetically difficult-to-reduce prochiral ketone tetrahydrofuran-3-one as a substrate, highly R- and S-selective variants were obtained (92–99 % ee) with minimal screening. The origin of stereoselectivity was provided by molecular dynamics analyses, which is discussed in terms of the Bürgi–Dunitz trajectory.
- Qu, Ge,Lonsdale, Richard,Yao, Peiyuan,Li, Guangyue,Liu, Beibei,Reetz, Manfred T.,Sun, Zhoutong
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p. 239 - 246
(2018/02/09)
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- Preparation of structurally diverse chiral alcohols by engineering ketoreductase CgKR1
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Ketoreductases are tools for the synthesis of chiral alcohols in industry. However, the low activity of natural enzymes often restricts their use in industrial applications. On the basis of computational analysis and previous reports, two residues (F92 and F94) probably affecting the activity of ketoreductase CgKR1 were identified. By tuning these two residues, the CgKR1-F92C/F94W variant was obtained that exhibited higher activity toward all 28 structurally diverse substrates examined than the wild-type enzyme. Among them, 13 substrates have a specific activity over 50 U mg-1 (54-775 U mg-1). Using CgKR1-F92C/F94W as a catalyst, five substrates at high loading (>100 g-1 L-1) were reduced completely in gramscale preparative reactions. This approach provides accesses to pharmaceutically relevant chiral alcohols with high enantioselectivity (up to 99.0% ee) and high space-time yield (up to 583 g-1 L-1 day-1). Molecular dynamics simulations highlighted the crucial role of residues 92 and 94 in activity improvement. Our findings provide useful guidance for engineering other ketoreductases, especially those possessing a similar active pocket to that in CgKR1.
- Zheng, Gaowei,Liu, Yuan-Yang,Chen, Qi,Huang, Lei,Yu, Hui-Lei,Lou, Wen-Yong,Li, Chun-Xiu,Bai, Yun-Peng,Li, Ai-Tao,Xu, Jian-He
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p. 7174 - 7181
(2017/11/06)
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- Stereo-complementary bioreduction of saturated N-heterocyclic ketones
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The asymmetric bioreduction of several saturated N-heterocyclic ketones is demonstrated in a stereo-complementary fashion using the ketoreductases READH and ChKRED20 for the production of (S)- and (R)-alcohols, respectively. The reaction accepts substrates with a five-, six- or seven-membered ring, and exhibits excellent stereoselectivity when using 2-propanol as both the ultimate reducing agent and cosolvent, achieve >99% ee in the majority of cases for both enantiomers.
- Li, Chao,Liu, Yan,Pei, Xiao-Qiong,Wu, Zhong-Liu
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- Development of a practical biocatalytic process for (S)-N-Boc-3-hydroxypiperidine synthesis
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(S)-N-Boc-3-hydroxypiperidine ((S)-NBHP) is an important chiral intermediate for the synthesis of ibrutinib, an anticancer drug targeting B-cell malignancies. An NADPH-dependent reductase (YDR541C) from Saccharomyces cerevisiae was isolated and found to show excellent catalytic activity in the production of (S)-NBHP. The reductase YDR541C was cloned and overexpressed in Escherichia coli BL21 (DE3), purified to homogeneity, and its catalytic properties were studied. Furthermore, an ethyl caprylate-water (1:1, v/v) biphasic system was introduced to alleviate product inhibition. After optimization of the reaction, as much as 1200?mM N-Boc-piperidin-3-one (NBPO) (240?g/L) was asymmetrically reduced to (S)-NBHP within 6?h, resulting in a yield of 99%, an enantioselectivity of >99.5% ee, and a total turnover number (TTN) of 8000. These results indicate great potential for industrial application.
- Chen, Li-Feng,Fan, Hai-Yang,Zhang, Yi-Ping,Wu, Kai,Wang, Hua-Lei,Lin, Jin-Ping,Wei, Dong-Zhi
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supporting information
p. 1644 - 1650
(2017/04/04)
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- Selection of microbial biocatalysts for the reduction of cyclic and heterocyclic ketones
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The reduction of carbonyl compounds plays an important role in the synthesis of complex chiral molecules. In particular, enantiopure substituted cyclic and heterocyclic compounds are useful intermediates for the synthesis of several antiviral, antitumor, and antibiotic agents, and recently, they have also been used as organocatalysts for C-C addition. Alcohol dehydrogenases (ADH) are enzymes involved in the transformation of prochiral ketones to chiral hydroxyl compounds. While significant scientific effort has been paid to the use of aliphatic and exocyclic ketones as ADH substrates, reports on (hetero)cyclic carbonyl compounds as substrates of these enzymes are scarce. In the present study, 109 bacteria and 36 fungi were screened, resulting in 10 organisms belonging to both kingdoms capable of transforming cyclic and heterocyclic ketones into the corresponding alcohols. Among them, Erwinia chrysanthemi could quantitatively reduce cyclododecanone and Geotrichum candidum could stereoselectively reduce N-Boc-3-piperidone and N-Boc-3-pyrrolidinone to their corresponding (S)-alcohols; however, the anti-Prelog isomer was obtained when acetophenone was the substrate.
- Bianchi, Paola,Varela, Romina Fernández,Bianchi, Dario A.,Kemppainen, Minna,Iribarren, Adolfo M.,Lewkowicz, Elizabeth
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p. 137 - 144
(2017/06/21)
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- PROCESS FOR THE SYNTHESIS OF STABLE AMORPHOUS IBRUTINIB
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Disclosed herein is a new route of synthesis and a new stable amorphous form of ibrutinib. Also disclosed are pharmaceutical compositions, oral dosage forms and the use of the amorphous ibrutinib in the treatment of mantle cell lymphoma or chronic lymphocytic leukemia.
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Page/Page column 17; 20
(2017/09/05)
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- A Bruton tyrosine kinase (Btk) method for the preparation of
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The invention relates to a preparation method of a Bruton's tyrosine kinase (Btk) intermediate (VI). The method comprises the following steps: carrying out benzylamine ring opening on a compound (S)-(+)-5-substituted-2-tetrahydrofurancarboxylic acid to obtain a product, and reacting the product with acyl chloride or acid anhydride to obtain a compound II; reducing the compound II, carrying out salt formation on the reduced compound II by using 1R-(-)-camphorsulfonic acid, and degrading the obtained salt to obtain a compound III; and reducing the compound III, carrying out salt formation on the reduced compound III and AcOH to obtain a compound V, and carrying out Boc addition on the compound V under an alkaline condition to obtain the target compound VI.
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- (S)-1-Boc-3-hydroxypiperidine synthesis method
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The invention discloses a (S)-1-Boc-3-hydroxypiperidine synthesis method. The method utilizes (R)-glyceraldehyde acetonide as a raw material. Through witting reaction-based acetonitrile group addition, palladium-carbon-based double bond reduction, deprotection, p-tosyl selective addition, Raney nickel-based hydrogenation reduction of the nitrile group along with ring closing, and piperidine ring nitrogen Boc protection, the (S)-1-Boc-3-hydroxypiperidine is obtained. The method utilizes the cheap chiral raw material as a starting material, produces a chiral product and prevents large material consumption and a yield loss of a splitting technology.
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- Chiral-1-tert-butoxy-carbonyl-3-hydroxy-piperidine, and the preparation of chiral turning method
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The invention relates to preparation of chirality-1-t-butyloxycarboryl-3-hydroxy piperidine and a method for chirality turning. The preparation mainly comprises the following steps: resolving N-benzyl-3-hydroxy piperidine as a raw material to obtain a (S) or (R)-1-benzyl-3-hydroxy piperidine camphorsulfonic acid salt, performing alkali freedom to obtain (S) or (R)-1-benzyl-3-hydroxy piperidine, performing palladium carbon hydrogenation debenzylation/t-butyloxycarboryl protection to obtain (S) or (R)-1-t-butyloxycarboryl-3-hydroxy piperidine, acylating substituting sulfonyl chloride of (R) or (S)-1-substituting-3-hydroxy piperidine as a raw material to obtain (R) or (S)-1-substituting-3-hydroxy piperidine sulfonate, substituting by using substituting carboxylate to obtain (S) or (R)-1-substituting-3-hydroxy piperidine carboxylic ester, and performing alkaline hydrolysis to obtain (S) or (R)-1-substituting-3-hydroxy piperidine. The synthesis route is gentle in reaction condition, and is applicable to industrial large-scale production.
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- Synthetic method of (S)-N-Boc-3-hydroxypiperidine
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The invention discloses a synthetic method of (S)-N-Boc-3-hydroxypiperidine. The synthetic method comprises: hydrogenating 3-hydroxypyridine to obtain 3-hydroxypiperidine; carrying out heating and refluxing on 3-hydroxypiperidine and a resolving agent namely D-pyroglutamic acid in ethanol solution, and cooling to separate out solids so as to obtain (S)-3-hydroxypiperidine D-pyroglutamate; and adding (S)-3-hydroxypiperidine D-pyroglutamate into di-tert-butyl dicarbonate ester under an alkaline condition, and carrying out refining after reaction to obtain (S)-N-Boc-3-hydroxypiperidine. According to the synthetic method disclosed by the invention, by optimizing a synthetic route of (S)-N-Boc-3-hydroxypiperidine and adopting the low-cost and recyclable resolving agent to reduce the production cost, the yield is improved while ensuring the product purity.
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- Development of a biocatalytic process to prepare (S)- N -boc-3-hydroxypiperidine
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(S)-N-Boc-3-hydroxypiperidine (S-NBHP) is a useful synthon for the synthesis of pharmaceutical intermediates including ibrutinib, the API of the newly approved drug Imbruvica, for the treatment of lymphoma. To our knowledge, there are no published biotransformation methods scalable to prepare S-NBHP. We report here the development of an efficient process catalyzed by recombinant ketoreductase (KRED) to reduce N-Boc-piperidin-3-one to obtain optically pure S-NBHP. The process has been optimized and demonstrated to have commercial potential with 100 g/L of substrate concentration, product of >99% ee with under 5% of enzyme loading (w/w).
- Ju, Xin,Tang, Yuanyuan,Liang, Xiaoliang,Hou, Maoqi,Wan, Zhonghui,Tao, Junhua
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p. 827 - 830
(2014/07/08)
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- A practical and enantiospecific synthesis of (-)-(R)- and (+)-(S)-piperidin-3-ols
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A highly enantiospecific, azide-free synthesis of (-)-(R)- and (+)-(S)-piperidin-3-ol in excellent yield was developed. The key step of the synthesis involves the enantiospecific ring openings of enantiomerically pure (R)- and (S)-2-(oxiran-2-ylmethyl)-1H-isoindole-1,3(2H)-diones with the diethyl malonate anion and subsequent decarboxylation.
- Babu, Meruva Suresh,Raghunadh, Akula,Ramulu, Konda,Dahanukar, Vilas H.,Syam Kumar, Unniaran K.,Dubey, P. Kumar
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p. 1507 - 1515
(2015/02/19)
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- Daucus carota mediated-reduction of cyclic 3-oxo-amines
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Carrots (Daucus carota) were used to reduce cyclic amino-ketones in high yields and enantiomeric excesses. This cheap, eco-compatible, and efficient reducing reagent allows the easy access to precursors of biologically active products.
- Lacheretz, Romain,Pardo, Domingo Gomez,Cossy, Janine
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supporting information; experimental part
p. 1245 - 1248
(2009/08/08)
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- A new asymmetric synthetic route to substituted piperidines
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An asymmetric synthesis of substituted piperidines has been described. β-Cyclodextrin- or oxazaborolidine-catalyzed asymmetric reduction of α-azido aryl ketones to the corresponding alcohols has been employed as the key step along with ring closing metathesis and selective dihydroxylation.
- Reddy, M. Somi,Narender,Rao, K. Rama
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p. 331 - 336
(2007/10/03)
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- QUINAZOLINE DERIVATIVES AS ANTITUMOR AGENTS
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A quinazoline derivative of the formula (I): (A chemical formula should be inserted here - please see paper copy enclosed) Formula I wherein the substituents are as defined in the text for use in the production of an anti proliferative effect which effect is produced alone or in part by inhibiting erbB2 receptor tyrosine kinase in a warm blooded animal such as man.
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Page/Page column 132
(2008/06/13)
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- ANTIPARASITIC TERPENE ALKALOIDS
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The present invention relates to novel terpene alkaloids and their use as antiparasitic agents. The present invention also relates to an antiparasitic agent which comprises a terpene alkaloid compound of this invention as an effective ingredient in an antiparasitic formulation. More particularly, the present invention relates to derivatives of the terpene alkaloid (1S,2R,4aS,5R,8R,8aR)-2-(acetyloxy)-8a-hydroxy-3,8-dimethyl-5-(1-methylethenyl)-1,2,4a,5,6,7,8,8a-octahydronaphthalen-1-yl (2S,3aR,9bR)-6-chloro-9b-hydroxy-5-methyl-1,2,3,3a,5,9b-hexahydropyrrolo[2,3-c][2,1]benzoxazine-2-carboxylate. Pharmaceutical compositions comprising the same are also disclosed.
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- Synthesis of piperidinyl and pyrrolidinyl butyrates for potential in vivo measurement of cerebral butyrylcholinesterase activity
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Biochemical changes in postmortem brains of Alzheimer's disease patients include decreased acetylcholinesterase and choline acetyl transferase activity, indicating reduced activity of the central cholinergic system, while butyrylcholinesterase (BChE) activity increases. A method that can measure regional BChE activity in the brain in vivo may be useful for investigating the relationship between BChE and Alzheimer's disease. Seven compounds, either piperidinyl or pyrrolidinyl butyrates, were synthesized as BChE substrate radiotracers to map central BChE activity in vivo by positron emission tomography (PET). 14C-labeled compounds were assayed to determine their hydrolysis rates by BChE and the partition coefficient. The five esters of secondary alcohols had lipophilic properties sufficient to pass readily through the blood-brain barrier while the metabolites were sufficiently hydrophilic to be retained in the brain. The esters showed moderate hydrolysis rates by BChE and high specificity for BChE relative to acetylcholinesterase, while two esters of primary alcohols were hydrolyzed too rapidly to estimate reliably the local cerebral BChE activity. From these results, we conclude that one or more of these five esters, when labeled with 11C, would be a useful tracer for quantification of BChE activity by PET.
- Kikuchi, Tatsuya,Fukushi, Kiyoshi,Ikota, Nobuo,Ueda, Takao,Nagatsuka, Shin-Ichiro,Arano, Yasushi,Irie, Toshiaki
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- Unprecedented oxidation of a phenylglycinol-derived 2-pyridone: Enantioselective synthesis of polyhydroxypiperidines
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Equation presented The phenylglycinol-derived 2-pyridone 1 undergoes m-CPBA oxidation steroselectively leading to the chiral nonracemic unsaturated bicyclic hydroxylactam 2, from which the enantioselective synthesis of (3R,5R)-3,4,5-trihydroxypiperidine (16) and the formal synthesis of the azasugar epiisofagomine are described. The enantioselective synthesis of (S)-N-Boc-3-hydroxypiperidine and (3R,4S)-3,4-dihydroxypiperidine is also reported.
- Amat, Mercedes,Llor, Nuria,Huguet, Marta,Molins, Elies,Espinosa, Enrique,Bosch, Joan
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p. 3257 - 3260
(2007/10/03)
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- Carbamic acid derivatives and method for preparing the same
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Anti-dementia medicaments comprising a carbamic acid derivative or their pharmaceutically acceptable acid addition salts having an anti-amnesic activity as an active ingredient are provided. The carbamic acid derivatives are represented by a general formula (1) STR1 wherein Ar denotes an aromatic heterocyclic ring which may have at least one substituent or its benzene-condensed ring, or phenyl group which may have at least one substituent, R1 denotes hydrogen atom or lower alkyl group, R2 denotes lower alkyl group which may be substituted with halogen atom, phenyl group which may have at least one substituent, naphthyl group, or five- or six-membered heterocyclic ring and its benzene-condensed ring, X and Y, which may be the same or different, denote sulfur atom or oxygen atom.
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