Patent Application
20080051589
The invention is based, in part, on the discovery of an efficient synthetic route for the interconversion of taxane molecules that may be realized using reductive deoxygenation, hydrolysis and acylation. In particular, the method of the invention has utility in transforming mixtures of taxane molecules into one specific taxane molecule.
As used herein, an “alkoxy group” means a linear, branched, or cyclic saturated hydrocarbon attached to an oxygen atom. Preferably, an alkoxy group has between one and six carbon atoms. An alkoxy group also refers to substituted alkoxy groups, which may include substituents such as alkanoyloxy groups, alkenyl groups, alkyl groups, alkylsilyl groups, alkylsulfonyl groups, alkylsulfoxy groups, alkylthio groups, alkynyl groups, amino groups such as mono- and di-alkylamino groups and mono- and di-arylamino groups, amide groups, aryl groups, arylalkyl groups, carboxy groups, carboxyalkoxy groups, carboxyamide groups, carboxylate groups, haloalkyl groups, halogens, hydroxyl groups, nitrile groups, nitro groups, phosphate groups, siloxy groups, sulfate groups, sulfonamide groups, sulfonyloxy groups, and combinations of these. Preferred examples of alkoxy groups include, among others, methoxy, ethoxy, propoxy, cyclopropoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, tert-butoxy, cyclobutoxy, pentoxy, isopentoxy, neo-pentoxy, cyclopentoxy, hexoxy, and cyclohexoxy.
As used herein, an “alkyl group” means a linear, branched, or cyclic saturated hydrocarbon. Preferably, an alkyl group has between one and six carbon atoms. An alkyl group also refers to substituted alkyl groups, which may include substituents such as alkanoyloxy groups, alkenyl groups, alky groups, alkylsilyl groups, alkylsulfonyl groups, alkylsulfoxy groups, alkylthio groups, alkynyl groups, amino groups such as mono- and di-alkylamino groups and mono- and di-arylamino groups, amide groups, aryl groups, arylalkyl groups, carboxy groups, carboxyalkoxy groups, carboxyamide groups, carboxylate groups, haloalkyl groups, halogens, hydroxyl groups, nitrile groups, nitro groups, phosphate groups, siloxy groups, sulfate groups, sulfonamide groups, sulfonyloxy groups, and combinations of these. Preferred substituents are alkoxy groups, amino groups such as dialkylamino groups, diarylamino groups, carboxylic acid-containing groups, haloalkyl groups, halogens, hydroxyl groups, nitrile groups, nitro groups, and sulfonic acid groups. Examples of preferred alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, cyclopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, cyclobutyl, pentyl, isopentyl, neo-pentyl, 1-ethylpropyl, cyclopentyl, hexyl, and cyclohexyl.
As used herein, an “aryl group” means a phenyl group or naphthyl group, which is optionally substituted. Examples of substituents on aryl groups include, but are not limited to, alkanoyloxy groups, alkenyl groups, alkoxy groups, alkylsilyl groups, alkylsulfonyl groups, alkylsulfoxy groups, alkylthio groups, alkynyl groups, amino groups such as mono- and di-alkylamino groups and mono- and di-arylamino groups, amide groups, aryl groups, arylalkyl groups, carboxy groups, carboxyalkoxy groups, carboxyamide groups, carboxylate groups, haloalkyl groups, halogens, hydroxyl groups, nitrile groups, nitro groups, phosphate groups, siloxy groups, sulfate groups, sulfonamide groups, sulfonyloxy groups, and combinations of these. Preferred substituents are alkoxy groups, alkyl groups, amino groups such as dialkylamino groups and diarylamino groups, carboxylic acid-containing groups, haloalkyl groups, halogens, hydroxyl groups, nitrile groups, nitro groups, and sulfonic acid groups.
As used herein, an “arylalkyl group” means an aryl group attached to an alkyl group. An example of an arylalkyl group is a benzyl group.
As used herein, a “basic baccatin III structure” means a compound having the formula as shown in
As used herein, an “ester group” means a linear, branched, or cyclic substituent having an ester functionality, i.e., —C(O)O—. Examples of ester groups include acyl groups such as actyl and benzoy, which are bound to a hydroxyl group.
As used herein, an “ether group” means a linear, branched, or cyclic substituent having an ether functionality, i.e., —COC—. An examples of an ether group includes, but is not limited to, HOCH2CH2OC(CH2OH)H—.
As used herein, a “glycoside group” or a “glycosyl group” means any of a number of sugar derivatives that contain a non-sugar group bonded to an oxygen or nitrogen atom and that on hydrolysis yield a sugar such as glucose. An example of a preferred gylcosyl group is xylosyl.
As used herein, a “halogen” means fluorine, chlorine, bromine, and/or iodine.
As used herein, a “heterocyclic group” is a saturated, unsaturated, or aromatic cyclic compound that contains at least one atom other than carbon, e.g., oxygen, nitrogen, or sulfur, in a ring. Examples of heterocyclic groups include furyls such as 2-furan, morpholino, piperadino, piperazino, N-methylpiperazino, pyrrollyl, pyridyl, and thiophene.
As used herein, a “hydroxyl protecting group” means a substituent of a hydroxyl group that is employed to block or protect the hydroxyl functionality, often while reacting other functional groups on the molecule, but not always. Examples of hydroxyl protecting groups are well known in the art and are described in J. W. Barton, “Protecting Groups in Organic Chemistry”,: J. G. W. McOmie, ed., Plenum Press, New York, N.Y., 1973, and in T. W. Green and P. G. M. Wuts, “Protective Groups in Organic Synthesis”, John Wiley & Sons, New York, N.Y., 1999. Each of these references is incorporated herein by reference in their entirety.
As used herein, an “oxo-group” means a substituent derived from the oxidation of a glycoside group such as a xyloside as described in U.S. Pat. No. 5,356,928.
As used herein, “taxane or taxane molecule” means a molecule that contains a basic baccatin m structure with a (2R,3S)—C6H5CH(Rx)CH(OH)C(O)— group forming an ester with the hydroxyl group located at the C-13 position of the basic baccatin III structure. The group represented by Rx can be an amino group, a salt of an amino group (e.g., an ammonium salt), an amino group which is protected with an amino protecting group, or a substituent which may be converted into an amino group. Various isomers, homologues, and analogues of the basic baccatin m structure, and of the (2R,3S)—C6H5CH(Rx)CH(OH)C(O)— group also are included in the definition of a taxane molecule. For example, a 10-deacetylbaccatin III structure is contemplated within the scope of a taxane molecule. Included within the definition of a taxane or taxane molecule are taxol A (paclitaxel), taxol B (cephalomanninc), taxol C, taxol D, taxol E, taxol F, taxol G, docetaxel (TAXOTERE®), (see, e.g.,
As used herein, a “vinyl group” means a linear or branched substituent having a carbon-carbon double bond. Examples of vinyl groups include, but are not limited to, 1-methyl-1-propenyl (CH3CH=C(CH3)—), and 2-methyl-1-propenyl ((CH3) 2C=CH—).
Throughout the description, where compositions are described as having, including, or comprising specific components, or where processes are described as having, including, or comprising specific process steps, it is contemplated that compositions of the present invention also consist essentially of, or consist of, the recited components, and that the processes of the present invention also consist essentially of, or consist of the recited processing steps. Further, it should be understood that the order of steps or order for performing certain actions are immaterial so long as the invention remains operable. Moreover, two or more steps or actions may be conducted simultaneously.
Suitable starting materials for use in the present invention include, but not limited to, any taxane molecule with a C13 side chain containing an amide group. Taxanes containing an N-acylated phenylisoserine side chain at the C13 position may serve as a starting material for this invention. In addition the starting material may be a mixture of two or more of these taxanes. These compounds may have varying substitution patterns; for instance, the C10 position may contain an acetate or a hydroxyl, and the C7 position may contain a hydroxyl or a xylosyl group. Examples of these compounds may be found in
In a particular embodiment, the invention provides a method for converting a taxane molecule having the formula as shown in
A more preferred starting material of the interconversion reaction of the invention is a taxane molecule having the formula as shown in
The starting taxanes used in the present invention may or may not need to be OH— protected. In some cases OH protection may be desired and may be carried out such that one or more of the active groups, including but not limited to OH groups at C7, C10, and C2′; are protected with a hydroxyl protecting group. It has been shown that protection at the C2′ hydroxyl group is especially important because this dramatically reduces the amount of side chain cleavage observed in the subsequent reductive deoxygenation step.
It should be understood that a hydroxyl-protecting group may remain on an end product. Examples of preferred hydroxyl protecting groups include acetate (Ac), benzoyl (Bz), trimethylsilyl (TMS), triethylsilyl (TES), 2,2,2-trichloroethoxycarbonyl (Troc), and ethoxylethyl ether (EE). Procedures for preparing the protected taxanes differ depending on the choice of protecting groups. In one preferred embodiment, the C2′ and C7 hydroxyl groups are protected by treating the starting taxane in anhydrous THF with a hindered base such as diisopropylethylamine, followed by a catalytic amount of N, N-dimethylaminopyridine (DMAP), and then chlorotrimethylsilane from 0-25° C. In another embodiment, the starting taxane in anhydrous THF is treated with ethylvinyl ether and a catalytic amount of pyridinium para-toluenesulfonate. In both cases, typically a slight excess of protecting reagent is used per hydroxyl group being protected. A non-limiting example of preparing a protected taxanes amide is shown in
In the present invention, the hydroxyl protected or unprotected taxane amide may be reductively deoxygenated to produce a protected or unprotected taxane imine or iminium compound. If, in the starting taxane amide, RN is hydrogen, then a taxane imine may be formed. In one alternative embodiment, the imine compound generally has the formula shown in
Throughout this invention the use of taxane imine compound includes, but is not limited to, taxane imines and taxane iminium compounds, including those described above. In one alternative embodiment, this reaction may be achieved by contacting the taxane amide with a suitable reducing agent, including but not limited to, a transition metal containing compound. The reductive deoxygenation may be effected using reagents known in the art. For example, the reductive deoxygenation step is preferably carried out using a zirconium hydride compound such as zirconocene chloride hydride (bis(cyclopentadienyl) zirconium chloride hydride), also known as Schwartz's reagent. A non-limiting example of this reaction is shown in
Other transition metal containing compounds or transition metal reducing agents, include, but are not limited to, titanium-containing reducing agents, hafnium-containing reducing agents, niobium-containing reducing agents, and molybdenum-containing reducing agents. Analogues and derivatives of Schwartz's reagent may also be used non-limiting examples of these reagents are shown in
In one alternative embodiment, the amount of Schwartz's reagent used can vary from about 0.1 molar equivalent up to about 10 molar equivalents per mole of starting material. Preferably, about 3 or more molar equivalents of Schwartz's reagent are used. More specifically, the reductive deoxygenation reaction may consume 2 molar equivalents of Schwartz's reagent and an additional equivalent may be added to help drive the reaction to completion. Also, an additional equivalent of Schwartz's reagent may be needed if any hydroxyl groups are left unprotected as free hydroxyl groups will react and neutralize one equivalent of Schwartz's reagent. In one alternative embodiment, the Schwartz's reagent may be added to the taxane amide solution either as a dry powder or as a slurry in an appropriate solvent. Preferably, the solvent is anhydrous. Tetrahydrofuran is a preferred solvent. In one preferred embodiment, the taxane amide solution in anhydrous THF, and is added to a slurry of Schwartz's reagent also in anhydrous THF.
The reductive deoxygenation preferably is carried out in an inert environment, e.g., under a nitrogen or argon atmosphere. In a preferred embodiment, the Schwartz's reagent slurry and the taxane amide solution is cooled below ambient temperature prior to mixing. The pre-reaction slurry/solution temperature is preferably less than about 15° C., and more preferably less than about 10° C.
After addition of the taxane amide solution to the Schwartz's reagent slurry, hydrogen gas may be generated and the reaction solution will warm slightly as the reaction is mildly exothermic. Following complete addition, agitation of the reaction solution continues at a reduced temperature until the reaction is deemed complete. The reaction time is usually about 1-4 hours, but it may be longer. The reaction is deemed complete when the starting materials are substantially converted to a taxane imine. Preferably, the reaction is stopped when less than 10% of the starting material remains based on HPLC area of the starting taxane amide and the taxane imine.
Subsequent to completion of the reductive deoxygenation reaction, most of the transition metal and or transition metal by-products may be removed before proceeding with the next reaction. For example, in one alternative embodiment, all or substantially all of the transition metal or metal by-products may be removed such that the resultant mixture comprises less than 10,000 parts/million, preferably less than 5,000 parts/million, or more preferably less than 1000 parts per million of transition metal or transition metal by-products. For the present invention, removal of the transition metal or any metal by-products is optional.
It is understood that the removal of transition metal or any transition metal by-products may be performed separately or in conjunction with any other step described herein. Also, such removal step may be performed at various times during the process of the present invention.
Techniques for removing transition metal compounds or transition metal by-products are known in the art, including, but not limited to, complexation, precipitation, filtration, chelation, centrifugation, electrochemical methodology, chromatography, or any combination thereof. In one alternative embodiment, a chelating agent comprising a chelating agent effective to chelate a transition metal may be used to remove the transition metal compound or transition metal by-product. Such chelating agents may include, but are not limited to, ethylene diamine tetra acetic acid (EDTA), ethylene glycol (bis) aminoethyl ether tetra acetic acid (EGTA), 1,2-bis- (o-aminophenoxy)ethane, N, N, N′, N′-tetra-acetic acid (BAPTA), N, N, N′, N′-tetrakis-(2-pyridylmethyl)ethylenediamine (TPEN), nitrilotriacetic acid, TIRON®, and analogues and derivatives thereof.
In one alternative embodiment, the cooled reductive deoxygenation reaction solution is added to an excess of N,N-bis(2-hydroxyethyl)glycine (bicine) as an aqueous solution while maintaining the solution at an ambient temperature. About 2 or more equivalents of bicine preferably may be used based on the amount of transition metal present. Subsequently, the reaction may be worked-up, which may include additional treatment of the original reductive deoxygenation reaction solution with additional aqueous bicine solution. In a preferred embodiment, the aqueous bicine layer may be back extracted with an organic solvent, preferably THF or ethyl acetate, to recover any taxane imine in the bicine layer. A non-limiting example of this reaction is shown in
Following conversion to an imine compound, hydrolysis of the imine functionality produces an amine or amine salt at the C-3′ position. Any common acid may be used to effect the hydrolysis of the imine compound. Such acids may include, but are not limited to: (i) hydrofluoric acid, hydrochloric, hydrobromic acid, or hydroiodic acid; (ii) nitric acid and or other nitrogen containing acids; (iii) sulfuric acid or other sulfur containing acids; (iv) carboxcylic acids, except for trifluro acetic acid; (v) phosphoric acid or other phosphate containing acids; (vi) tartaric acid; (vii) perchloric acid; (viii) p-tolulene sulfuric acid; (ix) picric acid. Preferably, the acid is sulfuric acid. The acid may be aqueous and/or in solution with a protic solvent, e.g. ethanol and/or methanol. In one alternative embodiment of the present invention, the amine salt may have the formula shown in
Hydrolysis of the imine compound may be carried out at or about ambient temperature. Also, hydrolysis may be carried out on an isolated or purified taxane imine. Preferably, an aqueous solution of acid is added directly to the reductive deoxygenation reaction solution after zirconium removal. The amine hydrolysis step may be employed before or after the addition of the anti-solvent (see below) to produce the taxane amine from the reaction mixture.
In one embodiment, the presence of the amine salt was confirmed using ES-MS/MS mass spectroscopy (Micromass Quattro LC Mass Spectrometer) and electrospray ionization mass spectrum.
In addition to hydrolyzing the taxane imine, the acid can also serve to remove the hydroxyl protecting groups if the protecting groups used are sensitive to acid. Examples of acid sensitive protecting groups include, but are not limited to, trimethylsilyl, triethylsilyl, and ethoxyethyl ether. Therefore, care should be taken in the selection of an appropriate hydrolysis reagent to avoid unwanted removal of protecting groups that may be present on the taxane molecule. However, it may be desirable to remove certain protecting groups on a taxane molecule to facilitate conversion of a “protected” taxane molecule to a known taxol derivative, e.g., removal of a silyl or ether protecting group from the C7, C10, and/or C2′ hydroxyl groups. Accordingly, if an acid is to perform additional functions in the reaction, the amount of acid used in the reaction should be appropriately adjusted.
The taxane amine salt may be solidified by adding an appropriate amount of a less polar solvent, preferably methyl tert-butyl ether. Other solvents, such as dichloromethane, heptane, hexane, toluene, or trifluorotoluene may also be used. Once solidified, the taxane amine salt can be easily filtered thus providing purification. Essentially all non-amine by-products or unreacted starting material remain in the filtrate and are thus removed.
The taxane amine salt may be converted into paclitaxel or other useful taxanes by reacting the amine in various ways. For instance, the appropriate amine salt may be converted into paclitaxel by reacting it with benzoyl chloride or some other benzoylating agent. This may be accomplished by first dissolving the amine salt in a solvent, preferably THE and then neutralizing the amine salt with excess base, preferably an inorganic base, most preferably sodium carbonate, followed by addition of a slight excess of a benzoylating agent, preferably benzoyl chloride. A non-limiting example of such a reaction is shown in
It has been shown however, that when the benzoylating step procedure is followed the production of a specific impurity has been found. The structure of this impurity has not been conclusively established but was found to have a molecular weight of 1104 Daltons. This impurity has also been found to be difficult to remove in downstream purification steps. The formation of this impurity was found to be dependent on the pH of the benzoylation step; therefore the moderation of base used to neutralize the amine salt was important. In a preferred alternative embodiment, the amine salt is dissolved in THF and then treated with benzoyl chloride followed by phosphate buffer of about pH 7 such that the final pH of the benzoylation solution was about 4 to 6. Using the phosphate buffer in place of the excess base dramatically reduced the formation of this impurity.
Once the benzoylation step is deemed complete (<2% amine remaining by HPLC area) the crude paclitaxel may be worked up by separating and removing the aqueous phosphate layer, washing the organic layer with saturated NaCl solution, drying the organic layer with a drying agent, preferably magnesium sulfate (MgSO4), and precipitating the dried organic layer into 4-6X hexane or heptane. The product precipitate can then be easily filtered and dried and is adequate for further purification. Following this procedure it was found that after drying, the formation of two impurities was observed. Both of these impurities had molecular weights of 871 Daltons and were determined to be oxetane ring-opened paclitaxel derivatives. These derivatives are well known in the literature and are known to form in the presence of eletrophilic reagents such as acid chlorides. Therefore, residual amounts of benzoyl chlorides were determined to be the cause of these impurities.
In a preferred alternative embodiment, about 0.5 molar equivalents of amines, preferably primary amines, most preferably ammonium hydroxide was added to the benzoylation reaction solution after the reaction was deemed complete to react with any residual benzoyl chloride. The aqueous layer is then separated and removed and the organic layer is washed twice more with the appropriate amine followed by a saturated NaCl solution wash each time. The organic layer is then dried with a drying agent, preferably magnesium sulfate (MgSO4), and precipitating the dried organic layer into 4-6X hexane or heptane. Following this protocol, the formation of the MW 871 impurities was dramatically reduced.
The end product and/or isolated intermediates may be analyzed using analytical techniques known in the art such as infrared (IR) spectroscopy, nuclear magnetic resonance (NMR) spectroscopy, e.g., 1H—NMR and 13C—NMR, high performance liquid chromatography (HPLC), e.g., reversed phase HPLC, and/or mass spectrometry (MS), e.g., electrospray ionization MS (ES MS) and matrix-assisted laser adsorption ionization MS (MALDI-MS). Combinations of these techniques also may be used, e.g., HPLC-MS.
In an embodiment of the above-described methodology, the invention is directed to methods of interconverting mixtures of taxane molecules, e.g., from biomass or biomass extracts, to a particular taxane molecule, e.g., paclitaxel. The methodology for interconversion of mixtures of taxane molecules into a specific taxane molecule generally is the same as for the interconversion of a single taxane molecule into another single taxane molecule.
The following materials may be used in the present invention as described below:
STEP 1: To a solution of taxane amides (500 g, 586 mmol) in 1.5 L dry THF under nitrogen, 7.2 g DMAP (590 mmol) is added, followed by 338 mL N-ethyl diisopropyl amine (Hünig's Base; 1.47 mol) and slow addition of TMS chloride (1.47 mol) over about an hour. The reaction is stirred for another hour at RT. HPLC (XTerra RP C-18 Column; Water-acetonitrile gradient with 0.05% formic acid) should show no taxanes at this time. The reaction is cooled to 0° and then filtered. The salt is washed with 500 mL dry THF.
STEP 2: The solution from STEP 1 is cooled to 0° under nitrogen and added slowly to a slurry of 469 g (1.97 mol) Bis(cyclopentadienyl)zirconium chloride hydride (Schwartz's Reagent) in 1 L dry THF under nitrogen. There is hydrogen evolution at this point. Thus, the reaction vessel should be large enough to accommodate some foaming. The reaction is stirred at 0° for about 4 hours while monitoring by HPLC, and is then taken to ca. 90% completion, and quenched before significant side-products form.
STEP 3: The reaction is quenched with aqueous bicine solution (1 M; 2.8 L); the layers are separated and the organic phase is washed with a second 2.8 L of bicine solution. The combined aqueous layers are extracted once with 500 mL ethyl acetate, and the combined organic phase is treated with 350 ML 10% sulfuric acid solution with good stirring, for one hour.
STEP 4: The hydrolyzed mixture from STEP 3 is added to 10 L MTBE, stirred well for 30 minutes, and filtered. The solid amine salt is washed with 2×2 L 4:1 LMTBE:THF. The LMTBE and LMTBE:THF solutions are saved to recover unreacted taxanes. The solid is dissolved in 2 L THF, and then 200 mL water is added, followed by 317 g solid NaHCO3. Caution: There is some foaming at this point. Preferably, the sodium bicarbonate is added slowly. The mixture is stirred well for 15 min after the addition of the sodium bicarbonate. Then, 68 mL Benzoyl chloride is added, and the reaction is stirred well for 1 hour. The reaction is then treated with 3 L brine, and the layers separated. The brine solution is washed once with 500 mL ethyl acetate. The combined organic layers are dried over MgSO4 and filtered. The resulting paclitaxel solution is then added to 15 L heptane, and the precipitate is filtered and vacuum-dried overnight, providing a 75-80% yield (i.e., approximately 375-400 g). The paclitaxel is 70% to 90% pure, or 75% to 95% pure, or 80% to 90% pure, or 90% to 95% pure. If desired, the paclitaxel may be purified further.
This example demonstrates the conversion of a taxane amide mixture composed mainly of taxol C and taxol E into taxol A (paclitaxel)
A taxane amide mixture composed of 2.6% taxol A, 4.8% taxol B, 54.1% taxol C, 0.2% taxol D, 32.0% taxol E, and 4.2% taxol F (100 g., ˜117 mmol) was dissolved in 500 mL anhydrous THF. Then solid Schwartz's reagent (137 g., 533 mmol) was added at 5° C. and the reaction was stirred at that temperature under nitrogen and was monitored by HPLC. After 4 hours the reaction was stopped by stirring it with 800 mL 1 M bicine solution for 30 minutes. The organic layer was stirred with another 800 mL 1 M bicine for 30 minutes. The combined aqueous layers were stirred with 150 mL ethyl acetate for 20 minutes. The THF and ethyl acetate layers were combined and treated with 70 mL 10% H2SO4 for 1 hour followed by the slow addition of 1800 mL methyl tert-butyl ether (MTBE) and an additional 30 minutes of stirring. The taxane amine salt was filtered and washed with MTBE:THF 3:1 and then dissolved in 200 mL THF. Then 100 mL of water, 36 grams of NaHCO3 (429 mmol), and 14 mL of benzoyl chloride (234 mmol) was added. The reaction was complete after 1 hour and was washed with 100 mL water, then 100 mL saturated NaCl solution, and finally dried with MgSO4. The filtered solution was next precipitated in 1200 mL of heptane. After filtration and vacuum drying, the crude paclitaxel was obtained (51 grams, purity 76.5%, yield 44%).
This example demonstrates the conversion of a taxane amide mixture composed mainly of taxol A and taxol B into taxol A (paclitaxel) using ethoxyethyl ether protection.
A taxane amide mixture composed of 51.4% taxol A, 28.2% taxol B, 7.6% taxol C, 0.5% taxol D, 1.7% taxol E, 3.3% taxol F, and 0.6% taxol G (481.6 g., ˜578.8 mmol) was dissolved in 2.5 L anhydrous THF followed by the addition of 3.63 grams of pyridinium para-toluenesulfonate (14.5 mmol) and 138.4 mL ethylvinyl ether (1.45 mol). The reaction was stirred at room temperature and after 16 hours was complete. The reaction solution was next cooled down to 5° C. and 2.03 mL of triethylamine was added followed by 441.8 grams of solid Schwartz's reagent (1.72 mol). The reaction was stirred at 5° C. and monitored by HPLC. After 4 hours the reaction was stopped by stirring it with 2.58 L 1 M bicine solution for 30 minutes. The organic layer was stirred with another 2.58 L 1 M bicine for 30 minutes. The combined aqueous layers were stirred with 500 mL ethyl acetate for 20 minutes. The THF and ethyl acetate layers were combined and treated with 336.7 mL 10% H2SO4 for 1 hour followed by the slow addition of 7.5 L methyl tert-butyl ether (MTBE) and an additional 1 hour of stirring. The taxane amine salt was filtered and dissolved in 1 L THF. Another 85 mL 10 H2SO4 was added and stirred for 30 minutes followed by the slow addition of 3 L MTBE followed by 30 minutes of stirring. The amine salt was filtered and washed with 3×1 L MTBE:THF 3:1. The amine salt was then dissolved in 1 L THF. Then 200 mL water followed by 212.7 grams NaHCO3 (2.53 mol) was added cautiously. Next 80 mL benzoyl chloride (687 mmol) was added and the reaction was stirred at room temperature for 1 hour. The reaction solution was next washed with water, dried with MgSO4, filtered, and precipitated into 6 L heptane. Filtration and vacuum drying yielded the crude paclitaxel (507 grams, purity 78.2%, yield 80%).
This example demonstrates the conversion of a taxane amide mixture composed mainly of taxol C and taxol E into taxol A (paclitaxel) using ethoxyethyl ether protection.
A taxane amide mixture composed of 2.6% taxol A, 4.8% taxol B, 54.1% taxol C, 0.2% taxol D, 32.0% taxol E, and 4.2% taxol F (10 g., ˜11.7 mmol) was dissolved in 50 mL anhydrous THF followed by the addition of 75 mg of pyridinium para-toluenesulfonate (0.3 mmol) and 2.8 mL ethylvinyl ether (29.7 mmol). The reaction was stirred at room temperature and after 16 hours was complete. The reaction solution was next cooled down to −5° C. and 13.7 grams of solid Schwartz's reagent (53.4 mmol). The reaction was stirred at 5° C. and monitored by HPLC. After 6 hours the reaction was stopped by stirring it with 80 mL 1 M bicine solution for 30 minutes. The organic layer was stirred with another 80 mL 1 M bicine for 30 minutes. The combined aqueous layers were stirred with 5 mL ethyl acetate for 20 minutes. The THF and ethyl acetate layers were combined and treated with 7 mL 10% H2SO4 for 1 hour followed by the slow addition of 150 mL methyl tert-butyl ether (MTBE) and an additional 1 hour of stirring. The taxane amine salt was filtered and dissolved in 20 mL THF. Another 5 mL 10% H2SO4 was added and stirred for 30 minutes followed by the slow addition of 60 mL MTBE followed by 30 minutes of stirring. The amine salt was filtered and washed with 3×5 mL MTBE:THF 3:1. The amine salt was then dissolved in 20 mL THF. Then 5.2 grams NaHCO3 (61.9 mmol) was added cautiously. Next 1.4 mL benzoyl chloride (12 mmol) was added and the reaction was stirred at room temperature for 1 hour. The reaction solution was next washed with water, dried with MgSO4, filtered, and precipitated into 120 mL heptane. Filtration and vacuum drying yielded the crude paclitaxel (5.81 grams, purity 76%, yield 43.6%).
This example demonstrates the conversion of a taxane amide mixture composed mainly of taxol B into taxol A (paclitaxel) using trimethylsilyl ether protection.
A taxane amide mixture composed of 2.5% taxol A, 83.1% taxol B, 1.9% taxol C, 0.7% taxol D, 5.8% taxol E, 0.3% taxol F, 0.9% taxol G (10 g., ˜12 mmol) was dissolved in 30 mL anhydrous THF followed by the addition of 72 mg of N, N-dimethylaminopyridine (DMAP) (0.59 mmol), 5.13 mL diisopropylethyl amine (DIPEA) (29.43 mmol), and 3.72 mL chlorotrimethylsilane (TMS-C1) (29.31 mmol). The reaction was stirred at room temperature and after 1 hour was complete. The reaction mixture was then cooled to 0° C. and filtered (to remove amine hydrochloride salts). The filtrate was added to a slurry of 8.44 gram Schwartz's reagent (32.83 mmol) in 20 mL anhydrous THF at 0° C. under nitrogen. After 4 hours the reaction was stopped by stirring it with 50 mL 1 M bicine solution for 30 minutes. The organic layer was stirred with another 50 mL 1 M bicine for 30 minutes. The combined aqueous layers were stirred with 10 mL ethyl acetate for 20 minutes. The THF and ethyl acetate layers were combined and treated with 7 mL 10% H2SO4 for 1 hour followed by the slow addition of 210 mL methyl tert-butyl ether (MTBE) and an additional 30 minutes of stirring. The amine salt was filtered and dissolved in 30 mL THF and 2 mL water followed by 7.4 grams NaHCO3 (88.1 mmol) was added cautiously. Then 1.36 mL benzoyl chloride (12 mmol) was added and the reaction was stirred at room temperature for 15 minutes. The reaction solution was then washed with 20 mL saturated NaCl solution, dried over MgSO4, filtered, and precipitated into 120 mL heptane. Filtration and vacuum drying yielded the crude paclitaxel (9.28 grams, purity 87.8%, yield 86.6%).
This example demonstrates the conversion of a taxane amide mixture composed mainly of taxol A into taxol A (paclitaxel) using trimethylsilyl ether protection.
A taxane amide mixture composed of 79.2% taxol A, 17.4% taxol B, 1.6% taxol C, and 0.3% taxol E (2.5 g., ˜2.9 mmol) was dissolved in 7.5 mL anhydrous THF followed by the addition of 18 mg of N, N-dimethylaminopyridine (DMAP) (0.13 mmol), 1.54 mL diisopropylethyl amine (DIPEA) (8.8 mmol), and 1.1 mL chlorotrimethylsilane (TMS-C1) (8.7 mmol). The reaction was stirred at 0° C. and after 1 hour was complete. The reaction mixture was then filtered (to remove amine hydrochloride salts). The filtrate was added to a slurry of 2.26 gram Schwartz's reagent (8.8 mmol) in 3.3 mL anhydrous THF at 5° C. under nitrogen. After 4 hours the reaction was stopped by stirring it with 13 mL 1 M bicine solution for 30 minutes. The organic layer was stirred with another 13 mL 1 M bicine for 30 minutes. The combined aqueous layers were stirred with 2.5 mL ethyl acetate for 20 minutes. The THF and ethyl acetate layers were combined and treated with 1.75 mL 10% H2SO4 for 1 hour followed by the slow addition of 65 mL methyl tert-butyl ether (MTBE) and an additional 30 minutes of stirring. The amine salt was filtered and dissolved in 10 mL THF and 0.41 mL benzoyl chloride (3.5 mmol), 2.5 mL saturated NaCl solution, and 12.5 mL pH 7 phosphate buffer was added and the reaction was stirred at room temperature for 15 minutes. The reaction solution was then treated with 0.17 mL concentrated ammonium hydroxide and stirred for 15 minutes. The solution was then washed with 5 mL saturated NaCl solution. The separated organic layer was dried over 1.25 grams MgSO4, filtered, and precipitated into 73 mL heptane. Filtration and vacuum drying yielded the crude paclitaxel (2.3 grams, purity 80.9%, yield 76.5%).
This example demonstrates the conversion of N-methyl taxol A into N-methyl taxol C.
A sample of N-methyl taxol A (3.5 grams, 4.0 mmol) was dissolved in 10 mL anhydrous THF followed by the addition of 30 mg of N, N-dimethylaminopyridine (DMAP) (0.24 mmol), 2.0 mL diisopropylethyl amine (DIPEA) (11.5 mmol), and 1.4 mL chlorotrimethylsilane (TMS-C1) (11.4 mmol). The reaction was stirred at 0° C. and after 1 hour was complete. The reaction mixture was then filtered (to remove amine hydrochloride salts). The filtrate was added to a slurry of 3.0 gram Schwartz's reagent (11.8 mmol) in 5 mL anhydrous THF at 0° C. under nitrogen. After 4 hours the reaction was stopped by stirring it with 18 mL 1 M bicine solution for 30 minutes. The organic layer was stirred with another 18 mL 1 M bicine for 30 minutes. The combined aqueous layers were stirred with 7 mL ethyl acetate for 20 minutes. The THF and ethyl acetate layers were combined and treated with 2.2 mL 10% H2SO4 for 1 hour followed by the slow addition of 83 mL methyl tert-butyl ether (MTBE) and an additional 30 minutes of stirring. The amine salt was filtered and dissolved in 12 mL THF and 0.6 mL hexanoyl chloride (4.0 mmol), 3.1 mL saturated NaCl solution, and 17 mL pH 7 phosphate buffer was added and the reaction was stirred at room temperature for 15 minutes. The reaction solution was then treated with 0.22 mL concentrated ammonium hydroxide and stirred for 15 minutes. The solution was then washed with 6 mL saturated NaCl solution. The separated organic layer was dried over 1.7 grams MgSO4, filtered, and precipitated into 95 mL heptane. Filtration and vacuum drying yielded the crude paclitaxel (2.83 grams, yield 75%).
Throughout the description, where compositions are described as having, including, or comprising specific components, or where processes are described as having, including, or comprising specific process steps, it is contemplated that compositions of the present invention also consist essentially of, or consist of, the recited components, and that the processes of the present invention also consist essentially of, or consist of, the recited processing steps. Further, it should be understood that the order of steps or order for performing certain actions are immaterial so long as the invention remains operable. Moreover, two or more steps or actions may be conducted simultaneously.
The content of each of the patent and non-patent documents referred to herein is expressly incorporated herein by reference in its entirety.
The invention may be embodied in other specific forms without departing form the spirit or essential characteristics thereof The foregoing embodiments are therefore to be considered in all respects illustrative rather than limiting on the invention described herein. Scope of the invention is thus indicated by the appended claims rather than by the foregoing description, and all changes that come within the meaning and range of equivalency of the claims are intended to be embraced therein.
Also, the invention may suitably comprise, consist of or consist essentially of the elements or process steps described herein. Further, the invention described herein suitably may be practiced in the absence of any element or process step which is or is not disclosed herein.
This application claims priority from U.S. Provisional Application Ser. No. 60/401,191, filed Aug. 4, 2002. This application is incorporated herein by reference in its entirety.
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/US03/24666 | 8/4/2003 | WO | 00 | 5/30/2006 |
| Number | Date | Country | |
|---|---|---|---|
| 60401191 | Aug 2002 | US |
