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HK1091211B - Nucleotide analog compositions - Google Patents

Nucleotide analog compositions Download PDF

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Publication number
HK1091211B
HK1091211B HK06111839.6A HK06111839A HK1091211B HK 1091211 B HK1091211 B HK 1091211B HK 06111839 A HK06111839 A HK 06111839A HK 1091211 B HK1091211 B HK 1091211B
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Hong Kong
Prior art keywords
crystals
adefovir dipivoxil
crystalline
xrd
typically
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HK06111839.6A
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Chinese (zh)
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HK1091211A1 (en
Inventor
M‧N‧阿里米利
T‧T‧K‧李
L‧V‧马内斯
E‧J‧普里斯博
L‧M‧舒尔茨
D‧E‧凯利
J‧D‧小芒格
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吉尔利德科学股份有限公司
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Publication of HK1091211A1 publication Critical patent/HK1091211A1/en
Publication of HK1091211B publication Critical patent/HK1091211B/en

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Description

Nucleotide analogue composition
This application is a divisional application of a patent application having an international filing date of 23/7/1998, an application number of 98803744.0, entitled "nucleotide analogue composition".
The invention relates to a nucleotide analogue 9-2- [ (di (pivaloyloxy) methoxy ] phosphinyl ] methoxy ] ethyl ] adenine (adefovir dipivoxil or AD for short) and application thereof. The invention also relates to methods of synthesizing AD.
AD is the bis-pivaloyloxymethyl ester of the parent compound 9- [ 2- (phosphonomethoxy) ethyl ] adenine ("PMEA") which has antiviral activity in animals and humans. AD and PMEA are described, for example, in the following documents: U.S. Pat. Nos. 4,724,233 and 4,808,716, EP 481214, Benzaria et al, Nucleotides and Nucleotides (1995)14(3-5): 563-565, Holy et al, Collection, Czech, chem, Commun (1989)54: 2190. 2201 and52: 2801-53: 2753-2777, Starrett et al, Antiviral Res. (1992) 19: 267 and 273 and j.med.chem. (1994) 37: 1857-1864. To date, AD has only been provided in an amorphous (i.e., amorphous) form. There has been no report of making it into a crystalline substance.
Methods for crystallizing organic compounds are described in the following documents: landgree, the organ Practice in the Organic Laboratory 2 nd edition, pages 43-51, D.C. Heathane Co., Mass., Lexington, 1977; myerson, Handbook of Industrial crystallization, pages 1-101, Butterworth-Heinemann, Mass., 1993.
The present invention provides one or more compositions or methods that meet one or more of the following objectives.
It is a primary object of the present invention to provide compositions containing new forms of AD crystals having the properties required for large scale synthesis or formulation into therapeutic formulations.
It is a further object of the present invention to provide an AD having good melting point and/or flow properties or bulk density properties, which AD facilitates the manufacture and formulation of AD containing compositions.
It is a further object of the present invention to provide shelf-stable AD.
It is a further object of the present invention to provide AD that can be easily filtered and dried.
It is a further object of the present invention to provide high purity AD having a purity of at least about 97% (w/w) (preferably, at least about 98%).
It is another object of the present invention to eliminate or minimize the production of by-products during the synthesis of AD.
It is a further object of the present invention to provide a method for AD purification that avoids expensive and time consuming column chromatography.
The present invention achieves its primary object by providing the following compounds: crystalline AD, particularly anhydrous crystalline form (hereinafter referred to as "form 1") and hydrated form (C)20H32N5O8P1·2H2O, hereinafter referred to as "form 2"), methanol solvated form (C)20H32N5O8P1·CH3OH，Hereinafter referred to as "form 3"), fumarate salt or complex (C)20H32N5O8P1·C4H4O4Hereinafter referred to as "form 4"), hemisulfate salt or complex, hydrobromide salt or complex, hydrochloride salt or complex, nitrate salt or complex, methanesulfonic acid (CH)3SO3H) Salt or complex, ethanesulfonic acid (C)2H5SO3H) A salt or complex, a β -naphthalenesulfonate or complex, an α -naphthalenesulfonate or complex, (S) -camphorsulfonate or complex, a succinate or complex, a maleate or complex, an ascorbate or complex, a nicotinate or complex.
Examples of the invention include (1) crystalline form 1 AD having an X-ray powder diffraction ("XRD") spectrum, expressed in degrees 2 Θ, substantially having one or more (in any combination) peaks at about 6.9, about 11.8, about 12.7, about 15.7, about 17.2, about 20.7, about 21.5, about 22.5, and about 23.3, using Cu-ka radiation; (2) an AD of crystalline form 2 having, expressed in degrees 2 Θ, an X-ray powder diffraction ("XRD") spectrum using Cu-ka radiation substantially having one or more (in any combination) peaks at about 8.7-8.9, about 9.6, about 16.3, about 18.3, about 18.9, about 19.7, about 21.0, about 21.4, about 22.0, about 24.3, about 27.9, about 30.8, and about 32.8; (3) an AD of crystalline form 3 having an XRD spectrum, expressed in degrees 2 Θ, using Cu-ka radiation substantially having one or more (in any combination) peaks at about 8.1, about 8.7, about 14.1, about 16.5, about 17.0, about 19.4, about 21.1, about 22.6, about 23.4, about 24.2, about 25.4, and about 30.9; and crystalline form 4 AD having an XRD spectrum, expressed in degrees 2 theta, using Cu-ka radiation substantially having one or more (in any combination) peaks at about 9.8, about 15.2, about 15.7, about 18.1, about 18.3, about 21.0, about 26.3, and about 31.7.
Examples of the present invention include AD crystals having the crystal morphology shown in any one or more of fig. 4-10.
In other embodiments, the present invention provides methods for preparing AD crystals by forming crystals from a crystallization solution comprising about 6-45% AD and about 55-94% crystallization solvent, wherein the AD crystals are formed from the crystallization solutionThe mixed solvent is selected from (1) a mixture of acetone and di-n-butyl ether of about 1: 10v/v to about 1: 3v/v, (2) a mixture of ethyl acetate and di-n-propyl ether of about 1: 10v/v to about 1: 3v/v, (3) a mixture of tert-butyl alcohol and di-n-butyl ether of about 1: 10v/v to about 10: 1v/v, (4) a mixture of dichloromethane and di-n-butyl ether of about 1: 10v/v to about 1: 3v/v, (5) a mixture of ethyl ether and di-n-propyl ether of about 1: 10v/v to about 10: 1v/v, (6) a mixture of tetrahydrofuran and di-n-butyl ether of about 1: 10v/v to about 1: 3v/v, (7) a mixture of ethyl acetate and di-n-butyl ether of about 1: 10v/v to about 1: 3v/v, and, (8) A tetrahydropyran to di-n-butyl ether mixture of about 1: 10v/v to about 1: 3v/v, (9) an ethyl acetate to ethyl ether mixture of about 1: 10v/v to about 1: 3v/v, (10) t-butyl methyl ether, (11) ethyl ether, (12) di-n-butyl ether, (13) t-butyl alcohol, (14) toluene, (15) isopropyl acetate, (16) ethyl acetate, (17) a mixture consisting essentially of the following (a) and (B): (A) a first crystallization solvent of the formula R1-O-R2Wherein R is a hydrogen atom, a nitrogen1Is alkyl of 1, 2, 3, 4, 5 or 6 carbon atoms, R2Is alkyl of 2, 3, 4, 5 or 6 carbon atoms, or R1And R2The two are linked together to form a 5, 6, 7 or 8 membered ring, provided that the dialkyl ether is not methyl ethyl ether and (B) a second crystallization solvent selected from (a) R1-O-R2Wherein the second dialkyl ether is different from the first dialkyl ether but is not methyl ethyl ether, (b) toluene, (c) tetrahydrofuran, (d) t-butanol, (e) ethyl acetate, (f) dichloromethane, (g) propyl acetate, and (h) isopropanol.
Examples of the invention include pure crystalline AD (e.g., form 1 and/or form 2). Examples of the invention also include compositions comprising crystalline AD (e.g., form 1 and/or form 2) and one or more compounds (e.g., pharmaceutically acceptable excipients or compounds present in a reaction mixture comprising crystalline AD).
Examples of the present invention include a method of preparing AD in which AD is dissolved in methanol to form crystals.
Another example is crystalline AD: it is suitable for pharmaceutical compositions or pharmaceutical uses comprising, for example, one or more of the AD forms 1, 2, 3 and/or 4 and a pharmaceutically acceptable carrier for treating viral conditions known to be effective with PMEA in humans or animals [ retroviral infections (HIV, SIV, FIV) or hepatitis B or other hepatitis viruses (hepadnaviruses) infections, or DNA virus infections (human cytomegalovirus or herpes viruses, such as HSV1 or HSV2) ].
The present invention provides a method for preparing AD in crystalline form 2, in which method AD crystals are formed in the presence of water.
In another example, a method of preparing AD comprises contacting PMEA with oxymethylpivalate in N-methylpyrrolidone (NMP, 1-methyl-2-pyrrolidone) and trialkylamine (e.g., Triethylamine (TEA)), and recovering AD.
In yet another example, a PMEA composition is provided having less than about 2% salt, which composition is useful in a process comprising the steps of: contacting with PMEA containing less than about 2% salt.
In yet another example, the AD product is obtained by a process comprising the steps of: wet granules are prepared from a mixture containing liquid, form 1 AD and the desired excipients, and the wet granules are optionally dried.
Fig. 1 is an XRD pattern of the crystal of form 1. FIG. 2 is a thermogram obtained by differential scanning calorimetry of the crystal of form 1. Fig. 3 is a fourier transform infrared absorption spectrum of the crystal of form 1. FIGS. 4 to 10 are photographs of the form 1 crystal at 100 times magnification. Fig. 4-10 are copies of photographs taken at 128% magnification. Fig. 11 is an XRD pattern of the crystal of form 2. FIG. 12 is a thermogram obtained by differential scanning calorimetry of the crystal of form 2. Fig. 13 is a fourier transform infrared absorption spectrum of the crystal of form 2. Fig. 14 is an XRD pattern of the form 3 crystal. FIG. 15 is a thermogram obtained by differential scanning calorimetry of the crystal of form 3. Fig. 16 is an XRD pattern of the crystal of form 4. FIG. 17 is a thermogram obtained by differential scanning calorimetry of the crystal of form 4. Figure 18 is an XRD pattern of AD hemisulfate crystals. FIG. 19 is an XRD pattern of AD hydrobromide crystals. Figure 20 is an XRD pattern of AD nitrate crystals. Figure 21 is an XRD pattern of AD mesylate salt crystals. Figure 22 is an XRD pattern of AD ethanesulfonate crystal. Fig. 23 is an XRD pattern of AD β -naphthalenesulfonate crystals. Fig. 24 is an XRD pattern of AD α -naphthalenesulfonate crystals. FIG. 25 is an XRD pattern of AD (S) -camphorsulfonate crystals. Figure 26 is an XRD pattern of AD succinate crystals.
Herein, temperature refers to degrees Celsius (. degree. C.) unless otherwise indicated. Room temperature means about 18-23 ℃.
In this context, alkyl refers to linear, branched and cyclic saturated hydrocarbons. Unless otherwise stated to the contrary, "alkyl" or "alkyl moiety" herein refers to a hydrocarbon containing 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 n-, secondary-, tertiary-, or cyclic moieties. "C1-10The term "alkyl" refers to an alkyl group containing 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms. Examples thereof are-CH3、-CH2CH3、-CH2CH2CH3、-CH(CH3)2、-CH2CH2CH2CH3、-CH2CH(CH3)2、-CH(CH3)CH2CH3、-C(CH3)3、-CH2CH2CH2CH2CH3、-CH(CH3)CH2CH2CH3、-CH(CH2CH3)2、-C(CH3)2CH2CH3、-CH(CH3)CH(CH3)2-、-CH2CH2CH(CH3)2、-CH2CH(CH3)CH2CH3、-CH2C(CH3)3、-CH2CH2CH2CH2CH2CH3、-CH(CH3)CH2CH2CH2CH3、-CH(CH2CH3)(CH2CH2CH3)、-C(CH3)2CH2CH2CH3、-CH(CH3)CH(CH3)CH2CH3、-CH(CH3)CH2CH(CH3)2、-C(CH3)(CH2CH3)2、-CH(CH2CH3)CH(CH3)2、-C(CH3)2CH(CH3)2、-CH(CH3)C(CH3)3Cyclopropyl, cyclobutyl, cyclopropylmethyl, cyclopentyl, cyclobutylmethyl, 1-cyclopropyl-1-ethyl, 2-cyclopropyl-1-ethyl, cyclohexyl, cyclopentylmethyl, 1-cyclobutyl-1-ethyl, 2-cyclobutyl-1-ethyl, 1-cyclopropyl-1-propyl, 2-cyclopropyl-1-propyl, 3-cyclopropyl-1-propyl, 2-cyclopropyl-2-propyl and 1-cyclopropyl-2-propyl.
Herein, unless stated to the contrary, "alkoxide" means a hydrocarbon attached to an oxygen atom, said hydrocarbon being a hydrocarbon containing 1, 2, 3, 4, 5 or 6 carbon atoms as defined herein for alkyl groups. Examples thereof are-OCH3、-OCH2CH3、-OCH2CH2CH3、-OCH(CH3)2、-OCH2CH2CH2CH3、-OCH2CH(CH3)2、-OCH(CH3)CH2CH3、-OC(CH3)3、-OCH2CH2CH2CH2CH3、-OCH(CH3)CH2CH2CH3、-OCH(CH2CH3)2、-OC(CH3)2CH2CH3、-OCH(CH3)CH(CH3)2、-OCH2CH2CH(CH3)2、-OCH2CH(CH3)CH2CH3、-OCH2C(CH3)3、-OCH(CH3)(CH2)3CH3、-OC(CH3)2(CH2)2CH3、-OCH(C2H5)(CH2)2CH3、-OC(CH2)3CH(CH3)2、-O(CH2)2C(CH3)3、-OCH2CH(CH3)(CH2)2CH3and-OCH2CH2CH2CH2CH2CH3。
"trialkylamine" means substituted with three C1-6A nitrogen atom substituted with an alkyl moiety, the alkyl moieties being independently selected. Examples thereof are nitrogen substituted by 1, 2 or 3 alkyl moieties as follows: -CH3、-CH2CH3、-CH2CH2CH3、-CH(CH3)2、-CH2CH2CH2CH3、-CH2CH(CH3)2、-CH(CH3)CH2CH3、-C(CH3)3、-CH2CH2CH2CH2CH3、-CH(CH3)CH2CH2CH3、-CH(CH2CH3)2、-C(CH3)2CH2CH3、-CH(CH3)CH(CH3)2、-CH2CH2CH(CH3)2、-CH2CH(CH3)CH2CH3、-CH2C(CH3)3、-CH2CH2CH2CH2CH2CH3、-CH(CH3)CH2CH2CH2CH3、-CH(CH2CH3)(CH2CH2CH3)、-C(CH3)2CH2CH2CH3、-CH(CH3)CH(CH3)CH2CH3、-CH(CH3)CH2CH(CH3)2、-C(CH3)(CH2CH3)2、-CH(CH2CH3)CH(CH3)2、-C(CH3)2CH(CH3)2or-CH (CH)3)C(CH3)3。
As used herein, "heteroaryl" includes, for example and without limitation, Leo a. pattern, in Principles of modern Heterocyclic Chemistry (published 1968 by w.a. benjamin, new york), especially chapters 1, 3, 4, 6, 7 and 9; the Chemistry of Heterocyclic Compounds, materials of monograms (published by John Wiley & Sons, N.Y., 1950 to now), especially volumes 13, 14, 16, 19 and 28; and j.am.chem.soc., (1960) 82: 5566 those heterocycles described in.
Examples of heterocycles include, but are not limited to, pyridyl, thiazolyl, tetrahydrothienyl, thiooxytetrahydrofenyl, pyrimidyl, furyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, tetrazolyl, benzofuryl, thionaphthyl, indolyl, indolinyl (indolinyl), quinolyl, isoquinolyl, benzimidazolyl, piperidyl, 4-piperidonyl, pyrrolidinyl, 2-pyrrolidinonyl, pyrrolinyl, tetrahydrofuryl, tetrahydroquinolyl, tetrahydroisoquinolinyl, decahydroisoquinolyl, octahydroisoquinolyl, azonyl, triazinyl, 6H-1, 2, 5-thiadiazinyl, 2H, 6H-1, 5, 2-dithiazinyl, thianthrenyl, pyranyl, isobenzofuryl, chromenyl, xanthenyl, phenothiazinyl (phenoxathinyl), 2H-pyrrolyl, thienyl, pyrrolyl, 2-pyrrolidinyl, pyrrolyl, tetrazolyl, benzofuranyl, thiofuranyl, thienyl (phenoxathinyl), thienyl, Isothiazolyl, isoxazolyl, pyrazinyl, pyridazinyl, indolizinyl, isoindolyl, 3H-indolyl, 1H-indazolyl, purinyl, 4H-quinolizinyl, 2, 3-naphthyridinyl, 1, 5-naphthyridinyl, quinoxalinyl, quinazolinyl, 1, 2-naphthyridinyl, pteridinyl, 4 aH-carbazolyl, beta-carbolinyl, phenanthridinyl, acridinyl, pyrimidinyl, phenanthrolinyl, phenazinyl, phenothiazinyl, furazanyl, phenazinyl, isobenzodihydropyranyl, chromanyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperazinyl, indolinyl, isoindolinyl, quinuclidinyl, morpholinyl, oxazolidinyl, benzotriazolyl, benzisoxazolyl, oxindolyl, benzoxazolinyl, and isatinoyl (isatinoyl).
By way of illustration and not limitation, carbon-bonded heterocycles are bonded at the following positions: the 2, 3, 4, 5 or 6 position of pyridine, the 3, 4, 5 or 6 position of pyridazine, the 2, 4, 5 or 6 position of pyrimidine, the 2, 3, 5 or 6 position of pyrazine, the 2, 3, 4 or 5 position of furan, tetrahydrofuran, thiophene, pyrrole or tetrahydropyrrole, the 2, 4 or 5 position of oxazole, imidazole or thiazole, the 3, 4 or 5 position of isoxazole, pyrazole or isothiazole, the 2, 3 position of aziridine, the 2, 3 or 4 position of azetidine, the 2, 3, 4, 5, 6, 7 or 8 position of quinoline or the 1, 3, 4, 5, 6, 7 or 8 position of isoquinoline. More typically, carbon-bonded heterocycles include 2-pyridyl, 3-pyridyl, 4-pyridyl, 5-pyridyl, 6-pyridyl, 3-pyridazinyl, 4-pyridazinyl, 5-pyridazinyl, 6-pyridazinyl, 2-pyrimidinyl, 4-pyrimidinyl, 5-pyrimidinyl, 6-pyrimidinyl, 2-pyrazinyl, 3-pyrazinyl, 5-pyrazinyl, 6-pyrazinyl, 2-thiazolyl, 4-thiazolyl, or 5-thiazolyl.
The following are illustrative, but not limiting, of the present invention, and the nitrogen-bonded heterocycle is bonded at the following positions: aziridine, azetidine, pyrrole, pyrrolidine, 2-pyrroline, 3-pyrroline, imidazole, imidazolidine, 2-imidazoline, 3-imidazoline, pyrazole, pyrazoline, 2-pyrazoline, 3-pyrazoline, piperidine, piperazine, indole, indoline, 1-position of 1H-indazole, 2-position of isoindoline or isoindoline, 4-position of morpholine, 9-position of carbazole or β -carboline. More typically, the nitrogen-binding heterocyclic ring includes 1-aziridinyl, 1-azetidinyl, 1-pyrrolyl, 1-imidazolyl, 1-pyrazolyl and 1-piperidinyl.
In this context, AD that is "crystalline material," "crystalline," or "crystal" refers to solid AD in which substantially all of the constituent molecules are ordered in a certain three-dimensional spatial pattern (i.e., lattice). Crystalline or crystalline AD may comprise one or more types of constituents, e.g. AD.fumaric acid or AD.2H2And O. Crystalline material (i.e., crystals) may occur in one or more crystal habits (e.g., platelets, rods, platelets, or needles).
Unless explicitly stated or can be determined from the context, percentage (%) herein refers to weight (w/w) percentage. Thus, a solution containing at least about 40% AD refers to a solution containing at least about 40% (w/w) AD. Solid AD with 0.1% water means that 0.1% (w/w) water is associated in the solid.
Crystalline AD that is substantially free of amorphous AD refers to a solid composition in which greater than about 60% of the AD is present as crystalline material. Such compositions typically contain at least about 80% (typically at least about 90%) of one or more crystalline forms of AD, with the remainder of the AD being amorphous AD.
The compositions of the present invention may optionally contain salts of the compounds described herein, including pharmaceutically acceptable salts containing, for example, uncharged moieties or monovalent anions. These salts also include those derived from a combination of suitable anions such as inorganic or organic acids. Suitable acids include those having sufficient acidity to form stable salts, preferably those having low toxicity. For example, the acid can be generated by certain organic and inorganic acids (e.g., HF, HCl, HBr, HI, H2SO4、H3PO4) Or acid addition of organic sulfonic acids, organic carboxylic acids and basic centers (usually amines) to form the salts of the invention. Examples of organic sulfonic acids include C6-16Arylsulfonic acids, C6-16Heteroaryl sulfonic acids and C1-16Alkylsulfonic acids, such as benzenesulfonic acid, α -naphthalenesulfonic acid, β -naphthalenesulfonic acid, (S) -camphorsulfonic acid, methanesulfonic acid, ethanesulfonic acid, n-propanesulfonic acid, isopropylsulfonic acid, n-butanesulfonic acid, sec-butanesulfonic acid, isobutylsulfonic acid, tert-butanesulfonic acid, pentanesulfonic acid, and hexanesulfonic acid. Examples of the organic carboxylic acids include C1-16Alkyl radical, C6-16Aryl carboxylic acids and C4-16Heteroaryl carboxylic acids, such as acetic, glycolic, lactic, pyruvic, malonic, glutaric, tartaric, citric, fumaric, succinic, malic, maleic, hydroxymaleic, benzoic, hydroxybenzoic, phenylacetic, cinnamic, salicylic and 2-phenoxybenzoic acids. Such salts also include salts of the compounds of the invention with one or more amino acids. Suitable amino acids are many, especially the natural amino acids found as protein components, although amino acids typically have a basic or acidic group (e.g., lysine, arginine, or glutamic acid), a neutral group (e.g., glycine, serine, threonine, alanine, isoleucine, or leucine)Amino acid) side chain. Salts are generally biocompatible or pharmaceutically acceptable or non-toxic, especially non-toxic to mammalian cells. The biologically toxic salts are generally used with synthetic intermediates for the compounds of the invention. Salts of AD are typically crystalline, as described herein for form 4.
Examples include compositions that occur instantaneously upon carrying out the methods or operations of the present invention. For example, when contacting a sodium alkoxide with a solution of 9- (2-hydroxyethyl) adenine, the composition at the beginning of mixing will contain negligible amounts of sodium alkoxide. The composition is typically in the form of a heterogeneous mixture prior to sufficient agitation to mix the solutions. Such compositions typically contain negligible amounts of reaction products and primarily reactants. Likewise, as the reaction proceeds, the ratios between reactants, products and byproducts may vary. These transient compositions are intermediates generated during processing and are specifically included as examples of the present invention.
The present invention includes compositions comprising a mixture of two or more different crystal types (i.e., forms), such as form 1 and form 2 crystals, form 1, form 2 and form 4 crystals, or form 2 and form 4 crystals. Mixtures of form 1 and form 2 AD crystals may be present in pharmaceutical formulation or manufacture, typically, such mixtures have form 1 AD accounting for at least about 70%, typically at least about 90%, but in some cases such mixtures may have form 2 and/or amorphous AD up to about 70%.
Crystalline form of AD
As described by Starrett et al, j.med.chem., (1994) 19: 1857 AD prepared and recovered by the process described in 5-1864 and recovered by elution from a silica gel column with a mixture of methanol (about 4%) and methylene chloride (about 96%) and rotary evaporation at about 35 ℃ under reduced pressure precipitated in amorphous form.
The present inventors have identified several different crystalline AD morphologies. The present inventors have characterized their properties by several methods, typically by XRD and DSC thermograms. XRD is generally used by the practitioner to characterize or identify the composition of crystals (see, e.g., united states pharmacopoeia, volume 23, 1995, methods 941, pages 1843 to 1845, published by u.s.pharmaceutical Convention, Rockville, maryland, tout et al, X-Ray Structure Determination, a Practical Guide, published 1968 by MacMillan, new york). The diffraction patterns obtained from crystalline compounds are often characteristic for a given crystalline form, although weak or very weak diffraction peaks may not always appear in the same diffraction patterns obtained from successive lots of crystals. Especially in the case where there is a significant amount of other crystal forms in the sample (e.g., the form 1 crystal has been partially hydrated to form 2 crystal). The relative intensities of the bands, especially at low-angle X-ray incidence (low 2 θ), may vary due to dominant orientation effects arising from differences in, for example, crystal habit, particle size, and other measurement conditions. Thus, the relative intensities of the diffraction peaks are not ultimately characteristic of the crystal morphology being addressed. Rather, it is more important to note the relative positions of the peaks rather than their amplitudes to determine whether AD crystallization is one of the morphologies described herein. Each XRD peak in the different samples is generally within about 0.3-12 theta degrees, which is a broad peak. A broader XRD peak may consist of two or more peaks close together. For isolated peaks, a peak is typically found within about 0.12 θ degrees in continuous XRD analysis. Assuming that the XRD spectrum of one compound is measured with the same instrument in successive XRD analyses, the differences in XRD peak positions are mainly due to differences in sample preparation or purity of the sample itself. When we identify a separate XRD spike at a given location (e.g. about 6.9), this means that the peak is 6.9 ± 0.1. When we identify a broad XRD peak at a given position at a given 2 theta value, this means that the peak is + -0.3 at that 2 theta value.
It should be noted that in the present invention, there is no need to rely on all bands observed in the highly purified control sample; even one band may be characteristic for a given AD crystal morphology, e.g. 6.9 for morphology 1. Identification should focus on the location and overall pattern of the bands, especially by selecting bands that are unique to each crystalline form.
Additional diagnostic techniques that may optionally be used to identify crystalline AD include Differential Scanning Calorimetry (DSC), melting point determination, and infrared absorption spectroscopy (IR). DSC measures the thermal transition temperature when crystals absorb or release heat due to changes in their crystal structure or melting of the crystals. In continuous analysis, the thermal transition temperature and melting point are typically within about 2 ℃, usually within about 1 ℃. When we say that a compound has a given value of DSC peak or melting point, this means that the DSC peak or melting point is within ± 2 ℃. DSC provides an alternative method to distinguish between different crystalline forms of AD. Different crystalline morphologies can be identified, at least in part, by their different transition temperature characteristics. IR measures the absorption of infrared light by specific chemical bonds associated with groups in the molecule that vibrate in response to light. DSC and/or IR may thus provide physicochemical information that can be used to describe AD crystallization.
Form 1
Single crystal X-ray crystallography was used to characterize form 1 AD. The cell constants and orientation matrix obtained by least squares refinement using measured positions for 3242 reflections with I > 10 σ in the range 3.00 < 2 θ < 45.00 ° are as follows: a is 12.85，b＝24.50，c＝8.28β is 100.2 °, Z is 4, and space group Cc.
The XRD pattern of form 1 typically peaks at about 6.9 (typically at about 6.9 and about 20.7, or more typically at about 6.9, about 15.7 and about 20.7, and generally at least at about 6.9, about 11.8, about 15.7 and about 20.7). Typically, the XRD peak at about 6.9 or, generally, (1) the peak plus one or two additional peaks or (2) the peak plus one or two additional peaks at about 6.9 plus differential scanning calorimetry data or melting point data is sufficient to distinguish the crystals of form 1 from other forms or to identify form 1 itself. The spectrum of form 1 typically has peaks at about 6.9, about 11.8, about 12.7, about 15.7, about 17.2, about 20.7, about 21.5, about 22.5, and about 23.3. The XRD pattern of form 1 typically peaks at any one (or combination) of about 6.9 and/or 11.8 and/or 15.7 and/or 17.2 and/or 20.7 and/or 23.3. Fig. 1 is a crystal X-ray diffraction pattern of typical form 1. It is to be understood, however, that fig. 1-26 are merely illustrative and that the characteristic representations of other crystalline AD formulations may differ from those described above.
Form 1 AD is anhydrous and contains little or no detectable water. Generally, the form 1 crystals generally contain less than about 1% (typically less than about 0.5%, usually less than about 0.2%) water. Moreover, the form 1 crystals generally contain less than about 20% (typically less than about 10%, often less than about 1%, and often less than about 0.1%) of amorphous AD. Frequently, form 1 crystals are free of amorphous AD, as can be determined by DSC, XRD, or polarized light microscopy at 100 x magnification. Form 1 AD is typically substantially free of crystallization solvent, i.e., contains typically less than about 1%, usually less than about 0.6%, and does not contain lattice bound solvent molecules, if suitably recovered from the crystallization bath.
Form 1 crystals generally have a median volume, as measured by light scattering, of about 25 to about 150 μm, typically about 30 to about 80 μm. The form 1 formulations generally all contain crystals between about 1-200 μm in length, with a typical maximum dimension of each crystal in the formulation being about 60-200 μm. In some form 1 formulations, about 1-10% of the crystals in the formulation have a maximum dimension above 250 μm. The crystals of form 1 shown in FIGS. 4-10 typically have a platelet, lath, pin or irregular habit. Aggregates of form 1 crystals also appear, typically in the range of about 25 to 150 μm in diameter.
The form 1 crystal has a DSC endothermic transition at about 102 deg.C (see FIG. 2), and has a groupThe IR spectrum shown above in FIG. 3. The crystalline form 1 preparations have a bulk density of about 0.15 to about 0.60g/mL, usually about 0.25 to about 0.50g/mL, and a surface area of about 0.10 to about 2.20m2In terms of/g, typically about 0.20 to 0.60m2(ii) in terms of/g. Thus, the AD of form 1 is characterized by the XRD spectral peaks expressed in degrees 2 θ at any one of (or a combination of) about 6.9 and/or about 11.8 and/or about 15.7 and/or about 20.7 using Cu-ka radiation and the endothermic transition as measured by differential scanning calorimetry at 102 ℃. The characteristics of form 1 AD can also be characterized by distinct XRD spectral peaks expressed in degrees 2 θ at about 6.9 ± 0.1, about 11.8 ± 0.1, about 15.7 ± 0.1, 17.2 ± 0.1, about 20.7 ± 0.1 using Cu-ka radiation and endothermic transition peaks measured by differential scanning calorimetry at 102.0 ± 0.2 ℃ and/or endothermic onset temperatures at 99.8 ± 0.2 ℃.
Form 2
The XRD pattern of form 2 (an example of which is shown in fig. 11) typically peaks at about 22.0 (typically at about 18.3 and about 22.0, or more typically at about 9.6, about 18.3 and about 22.0, and generally at least at about 9.6, about 18.3, about 22.0 and about 32.8). Any 3 or 4 of these 4 XRD characteristic peaks, or typically (1)4 peaks or (2) 2 or 3 of these peaks plus differential scanning calorimetry data or melting point data, is sufficient to distinguish the crystallinity of form 2 from other forms or to identify form 2 itself. The XRD pattern of form 2 typically has peaks at about 8.7-8.9, about 9.6, about 16.3, about 18.3, about 18.9, about 19.7, about 21.0-21.3, about 21.4, about 22.0, about 24.3, about 27.9, about 30.8 and about 32.8.
The form 2 crystals are AD dihydrate and they are generally substantially free of detectable crystallization solvent other than water. The form 2 crystals generally contain less than about 30% (typically less than about 10%, often less than about 1%, and often less than about 0.1%) of amorphous AD. Crystalline generally does not contain amorphous AD, as can be determined by DSC, XRD, or polarized light microscopy at 100 x magnification. Form 2 crystals typically have a median volume, as measured by light scattering, of about 15 to about 85 μm, typically about 25 to about 80 μm. The form 2 formulations typically all contain crystals between about 1-300 μm in length. The form 2 crystals had a DSC endothermic transition at about 73 ℃ (see fig. 12) and had an IR spectrum substantially as shown in fig. 13.
Thus, the AD of form 2 is characterized by the XRD spectral peaks expressed in degrees 2 θ at any one of (or a combination of) about 9.6 and/or about 18.3 and/or about 22.0 and/or about 32.8 using Cu-ka radiation and the endothermic transition as measured by differential scanning calorimetry at 73 ℃. The characteristics of form 2 AD can also be characterized by distinct XRD spectral peaks expressed in degrees 2 θ at 9.6 ± 0.1, 18.3 ± 0.1, 22.0 ± 0.1, 24.3 ± 0.1 and 32.8 ± 0.1 using Cu-ka radiation and endothermic transition peaks measured by differential scanning calorimetry at 72.7 ± 2 ℃ and/or endothermic onset temperature at 69.5 ± 2 ℃.
Form 3
The XRD pattern of form 3 (e.g., as shown in fig. 14) typically peaks at about 8.1 (typically at about 8.1 and about 25.4, or more typically at about 8.1, about 19.4 and about 25.4). Typically, any 1 or 2 of these 3 XRD characteristic peaks, or generally (1) 3 or 4 of these peaks or (2) 2 or 3 of these peaks plus differential scanning calorimetry data or melting point data, is sufficient to distinguish the crystallinity of form 3 from other forms or to identify form 3 itself. Form 3 AD has an endothermic transition at about 85 ℃ as measured by differential scanning calorimetry (figure 15). The spectrum of form 3 typically has peaks at about 8.1, about 8.7, about 14.1, about 16.5, about 17.0, about 19.4, about 21.1, about 22.6, about 23.4, about 24.2, about 25.4 and about 30.9.
Unlike forms 1 and 2, form 3 crystals contain about 1 equivalent of methanol in the crystal lattice. Methanol is generally derived from the crystallization solvent. Form 3, however, is substantially free of other detectable solvents or water. Form 3 crystals generally contain less than about 20% (typically less than about 10%, often less than about 1%, and often less than about 0.1%) of amorphous AD. Crystalline does not contain amorphous AD, as can be determined by DSC, XRD or polarized light microscopy at 100 x magnification. The form 3 crystals typically have a median volume, as measured by light scattering, of about 20 to about 150 μm, typically about 30 to about 120 μm. The form 3 formulations typically all contain crystals between about 1-300 μm in length.
Form 4
The XRD pattern of form 4 (e.g., as shown in fig. 16) typically peaks at about 26.3 (typically at about 26.3 and about 31.7, or typically at about 26.3, about 31.7 and about 15.2, or typically at about 26.3, about 31.7, about 15.2 and about 21.0). Typically, these 4 XRD characteristic peaks, or typically (1) 3 of these peaks or (2) 2 or 3 of these peaks plus differential scanning calorimetry data or melting point data, are sufficient to distinguish the crystallinity of form 4 from other forms or to identify form 4 itself. Form 4 AD has an endothermic transition at about 121 ℃ and about 148 ℃ as measured by differential scanning calorimetry (fig. 17). The spectrum of form 4 typically has a peak at any one of (or a combination of) about 9.8, about 15.2, about 15.7, about 18.1, about 18.3, about 21.0, about 26.3 and about 31.7. Thus, the AD of form 4 is characterized by the XRD spectral peaks expressed in degrees 2 θ at any one of (or a combination of) about 15.2 and/or about 21.0 and/or about 26.3 and/or about 31.7 using Cu-ka radiation and the endothermic transition as measured by differential scanning calorimetry at about 121.3 ℃ and about 148.4 ℃. The characteristics of form 4 AD can also be characterized by distinct XRD spectral peaks expressed in degrees 2 θ at 9.8 ± 0.1, 18.1 ± 0.1, 21.0 ± 0.1, 26.3 ± 0.1 and 31.7 ± 0.1 using Cu-ka radiation and endothermic transition peaks measured by differential scanning calorimetry at 121.3 ± 2 ℃ and 148.4 ± 2 ℃.
Crystalline salts of organic and inorganic acids
Figures 18-26 are XRD spectra obtained from crystalline salts or complexes of AD with organic and inorganic acids. These salts are hemisulfate (FIG. 18), hydrobromide (FIG. 19), nitrate (FIG. 20), methanesulfonic acid (CH)3SO3H) Salt or complex (FIG. 21), ethanesulfonic acid (C)2H5SO3H) A salt or complex (fig. 22), a β -naphthalenesulfonate or complex (fig. 23), an α -naphthalenesulfonate or complex (fig. 24), (S) -camphorsulfonate or complex (fig. 25), and a succinate or complex (fig. 26). These XRS spectra show a number of peaks that characterize the compound and allow one skilled in the art to distinguish each compound from other crystalline forms.
Figure 18 is a hemisulfate salt or complex having characteristic XRD peaks at degrees 2 theta of any one (or combination) of about 8.0, about 9.5, about 12.0, about 14.6, about 16.4, about 17.0, about 17.5-17.7, about 18.3, about 19.0, about 20.2, about 22.7, about 24.1, and about 28.2. The salt or complex has a melting point of about 131-. It can be characterized by 4 characteristic peaks at about 12.0, about 14.6, about 16.4, about 17.5-17.7. Thus, its properties can be further characterized as having 3 or 4 of these XRD peaks and having a melting point of about 131-. The characteristics of the AD hemisulfate can also be characterized by distinct XRD spectral peaks expressed in degrees 2 theta at 8.0 + -0.1, 12.0 + -0.1, 14.6 + -0.1, 16.4 + -0.1 and 17.5-17.7 + -0.3 and a melting point of 131-.
Figure 19 is a hydrobromide salt or complex having characteristic XRD peaks at degree 2 theta of any one (or combination) of about 13.2, about 14.3, about 15.9, about 17.8, about 20.7, about 21.8, about 27.2 and about 28.1. The salt or complex decomposes upon heating at about 196-. Thus, its characteristics can be characterized by having 4 characteristic XRD peaks at about 13.2, about 14.3, about 17.8 and about 28.1. One skilled in the art can further characterize the compound as having 3 or 4 of these XRD peaks and decomposing upon heating at about 196-. The AD hydrobromide can also be characterized by distinct XRD spectral peaks expressed in degrees 2 theta at 13.2 + -0.1, 14.3 + -0.1, 17.8 + -0.1, 20.7 + -0.1 and 27.2 + -0.1 and a decomposition point of 196-.
Figure 20 is a nitrate or complex having characteristic XRD peaks at degrees 2 theta of any one (or combination) of about 8.0, about 9.7, about 14.1, about 15.2, about 16.7, about 17.1, about 18.3, about 18.9, about 19.4, about 20.0, about 21.2, about 22.3, about 23.2, about 24.9, about 27.6, about 28.2, about 29.4 and about 32.6. The salt or complex decomposes upon heating at about 135-136 deg.C. Thus, its characteristics can be characterized by having 4 characteristic XRD peaks at about 14.1, about 23.2, about 29.4 and about 32.6. One skilled in the art can further characterize the compound as having 3 or 4 of these XRD peaks and having a melting point of about 131-. The characteristics of the AD nitrate can also be characterized by distinct XRD spectral peaks expressed in degrees 2 theta at 8.0 + -0.1, 14.1 + -0.1, 23.2 + -0.1, 29.4 + -0.1 and 32.6 + -0.1 and decomposition points at 135-.
Figure 21 is a mesylate salt or complex having characteristic XRD peaks in degrees 2 theta of any one of (or a combination of) about 4.8, about 15.5, about 16.2, about 17.5, about 18.5, about 20.2, about 24.8, about 25.4, and about 29.5. The melting point of the salt or complex is about 138-. Thus, its characteristics can be characterized by having 4 characteristic XRD peaks at about 4.8, about 15.5, about 20.2 and about 24.8. One skilled in the art can further characterize the compound as having 3 or 4 of these XRD peaks and having a melting point of about 138-. The characteristics of the AD methanesulfonate salt were also characterized by distinct XRD spectral peaks expressed in degrees 2. theta. at 4.8. + -. 0.1, 15.5. + -. 0.1, 16.2. + -. 0.1, 20.2. + -. 0.1 and 24.8. + -. 0.1 and a melting point of 138 ℃ 139. + -. 2 ℃ using Cu-K.alpha.radiation.
Figure 22 is an ethanesulfonate or complex having characteristic XRD peaks at degrees 2-theta of any one of about 4.4, about 8.8, about 18.8, about 23.0-23.3, and about 27.3 (or combinations thereof). The melting point of the salt or complex is about 132-133 ℃. Thus, its characteristics can be characterized by having 4 characteristic XRD peaks at about 4.4, about 8.8, about 18.8 and about 27.3. One skilled in the art can further characterize the compound as having 3 or 4 of these XRD peaks and having a melting point of about 132-. The characteristics of the AD ethanesulfonate can also be characterized by distinct XRD spectral peaks expressed in degrees 2. theta. at 4.4. + -. 0.1, 8.8. + -. 0.1, 18.8. + -. 0.1, 23.0-23.3. + -. 0.3 and 27.3. + -. 0.1 and a melting point of 132-.
Figure 23 is a beta-naphthalenesulfonate salt or complex having characteristic XRD peaks in degrees 2-theta at any one of (or combination of) about 9.8, about 13.1, about 16.3, about 17.4, about 19.6, about 21.6-22.3, about 23.4, about 24.1-24.5, and about 26.6. The melting point of the salt or complex is about 156-157 ℃. Thus, its characteristics can be characterized by having 4 characteristic XRD peaks at about 13.1, about 17.4, about 23.4 and about 26.2. One skilled in the art can further characterize the compound as having 3 or 4 of these XRD peaks and having a melting point of about 156-157 ℃. The characteristics of the beta-naphthalenesulfonates of AD can also be characterized by using the distinct XRD spectral peaks expressed in degrees 2 θ at 9.8 + -0.1, 13.1 + -0.1, 17.4 + -0.1, 23.4 + -0.1, and 26.2 + -0.1 and the melting point at 157 + -2 deg.C of Cu-Ka radiation.
Figure 24 is an alpha-naphthalene sulfonate or complex having characteristic XRD peaks at degrees 2 theta of any one (or combination) of about 8.3, about 9.8, about 11.5, about 15.6, about 16.3, about 16.7-17.4, about 19.6, about 21.0, about 22.9, about 23.7, about 25.0, and about 26.1. The melting point of the salt or complex is about 122-128 ℃. Thus, its characteristics can be characterized by having 4 characteristic XRD peaks at about 9.8, about 15.6, about 19.6 and about 26.1. One skilled in the art can further characterize the compound as having 3 or 4 of these XRD peaks and having a melting point of about 122-128 ℃. The characteristics of the alpha-naphthalenesulfonates of AD can also be characterized by using the distinct XRD spectral peaks expressed in degrees 2 θ at 9.8 + -0.1, 15.6 + -0.1, 19.6 + -0.1, 21.0 + -0.1 and 26.1 + -0.1 and the melting point at 128 + -2 deg.C for Cu-Ka radiation.
Fig. 25 is an (S) -camphorsulfonate or complex having characteristic XRD peaks at degree 2 theta of any one (or combination) of about 5.4, about 6.5, about 13.7, about 15.5, about 16.8-17.2, about 19.6, about 20.4-20.7, about 21.2, about 23.1, about 26.1, about 27.5, about 28.4, about 31.3, and about 32.2. The melting point of the salt or complex is about 160-161 ℃. Thus, its characteristics can be characterized by having 4 characteristic XRD peaks at about 5.4, about 6.5, about 13.7 and about 16.8-17.2. One skilled in the art can further characterize the compound as having 3 or 4 of these XRD peaks and having a melting point of about 160-. The characteristics of the (S) -camphorsulfonic acid salt of AD can also be characterized by using Cu-Ka radiation, distinct XRD spectrum peaks expressed in degrees 2 theta of 5.4 + -0.1, 6.5 + -0.1, 13.7 + -0.1, 16.8-17.2 + -0.3 and 19.6 + -0.1 and a melting point of 160-.
Figure 26 is a succinate salt or complex having characteristic XRD peaks at degrees 2 theta of any one (or combination) of about 4.7, about 9.5, about 10.6, about 14.9, about 16.3, about 17.4, about 17.9, about 19.9, about 20.8, about 22.1, about 23.9-24.2, about 26.5, about 27.6 and about 28.2. The melting point of the salt or complex is about 122-124 ℃. Thus, its characteristics can be characterized by having 4 characteristic XRD peaks at about 4.7, about 9.5, about 14.9 and about 17.4. One skilled in the art can further characterize the compound as having 3 or 4 of these XRD peaks and having a melting point of about 122-124 ℃. The characteristics of the AD succinate can also be characterized by distinct XRD spectral peaks expressed in degrees 2 theta at 9.5 + -0.1, 14.9 + -0.1, 16.3 + -0.1, 17.4 + -0.1 and 23.9-24.2 + -0.3 and a melting point of 122- + 124 + -2 ℃ using Cu-Ka radiation.
Examples of the invention include compositions comprising a crystalline salt (e.g., a salt of AD as characterized above) and a pharmaceutically acceptable excipient. Other examples include methods of preparing pharmaceutical compositions by contacting a crystalline salt (such as the above-characterized salt of AD) with a pharmaceutically acceptable excipient. Other examples include products resulting from the process of contacting a crystalline salt (such as the above-characterized salt of AD) with a pharmaceutically acceptable excipient.
Method for synthesizing AD
The following scheme a is a representative process flow diagram for the preparation of AD and form 1 AD crystals.
Diagram A
The scale of the process steps shown in diagram a and described below can be increased or decreased by those skilled in the art, as desired.
Method for synthesizing diethyl p-toluenesulfonyloxymethylphosphonate
In one example, the synthesis of diethyl p-toluenesulfonyloxymethylphosphonate shown in step 1 of scheme A is described as follows. A mixture of diethyl phosphite (0.8kg), paraformaldehyde (0.22kg) and triethylamine (0.006kg) in toluene (2.69kg) was heated to 87 ℃ (84-110 ℃) in a reactor under an inert atmosphere (e.g. nitrogen) with stirring for 2 hours, then heated to reflux and maintained at reflux for 1 hour until the reaction was complete. The completion of the reaction was monitored by TLC (detection of traces of diethyl phosphite) and1H-NMR spectrum was confirmed by whether or not a diethyl phosphite peak was observed at delta 8.4-8.6ppm at more than 1%. The above solution was cooled to about 1 ℃ (-2 ℃ to 4 ℃) and p-toluenesulfonyl chloride (1.0kg) was added, after combustion triethylamine (0.82kg) was slowly added below 10 ℃ (for about 3-6 hours in an exothermic reaction). The resulting mixture is warmed to 22 ℃ (19-25 ℃) and stirred for at least 5 hours (typically about 16-24 hours) until the reaction is complete. The completion of the reaction was monitored by TLC (detection of traces of p-toluenesulfonyl chloride) and was monitored by1H-NMR (bimodal p-toluenesulfonyl chloride (. delta.7.9 ppm) was no longer detected) was confirmed. The solid was filtered off and washed with toluene (0.34 kg). The washings and the filtrate were combined and washed 2 times with water (1.15kg each), or if desired with water (1.15kg), a 5% aqueous solution of sodium carbonate (3.38kg) and 2 times with water (1.15kg each). If emulsification occurs, brine may be added to the first organic/water mixture. The organic phase is distilled under vacuum at a temperature below 50 [ Loss On Drying (LOD) not exceeding 10% and water content not exceeding 0.5% as determined by Karl Fischer titration ] to yield about 85-95% pure toluene-free oily title compound. Upon cooling, the oily compound became viscous.
Method for synthesizing 9- (2-hydroxyethyl) adenine
In a fruitIn the examples, the synthesis of 9- (2-hydroxyethyl) adenine shown in step 2 of scheme A is described as follows. Sodium hydroxide (6g) was added to a slurry of adenine (1.0kg) and molten ethylene carbonate (0.72kg, m.p.37-39 ℃) in DMF (2.5kg) in a reactor with an inert atmosphere (e.g. nitrogen) and the mixture was heated to 125 ℃ (95 ℃ to reflux) with stirring until the reaction was complete (about 3-9 hours if the mixture temperature was 110 ℃ to reflux and about 15-48 hours if the mixture temperature was 95-110 ℃). The reaction was monitored by HPLC for completion (residual adenine not exceeding 0.5%). The mixture was cooled to below 50 ℃ and diluted with toluene (3.2 kg). The resulting slurry was cooled to 3 ℃ (0-6 ℃) and stirred for at least 2 hours. The slurry was filtered and the filter cake was washed 2 times with cold (0-5 ℃) toluene (0.6 kg each). Vacuum drying the filter cake at 35-70 deg.C1Toluene by H-NMR or LOD of not more than 2%), optionally ground, to give the title compound as a white to off-white powdery solid.
Method for synthesizing 9- [ 2- (diethylphosphonomethoxy) ethyl ] adenine
The compound is prepared from sodium alcohol (C)1-6Alkyl) and 9- (2-hydroxyethyl) adenine. The sodium alkoxide (typically sodium t-butoxide or sodium isopropoxide) is contacted with 9- (2-hydroxyethyl) adenine in a solvent such as DMF at about 20-30 ℃ for about 1-4 hours. Syntheses with 1 molar equivalent of 9- (2-hydroxyethyl) adenine and about 1.2-2.2 molar equivalents of sodium alkoxide generally yield good results.
In one example, the synthesis of 9- [ 2- (diethylphosphonomethoxy) ethyl ] adenine shown in step 3 of scheme A is described as follows. A slurry of 9- (2-hydroxyethyl) adenine (1.0kg) and DMF (4.79kg) was heated to about 130 deg.C (125 ℃ C. and 135 ℃ C.) for 30-60 minutes in a reactor with an inert atmosphere (e.g., nitrogen). The reactor contents were rapidly cooled to about 25 deg.C (20-30 deg.C) with vigorous stirring, and sodium t-butoxide (0.939kg) was added in portions over about 1-3 hours while maintaining vigorous stirring and maintaining the contents temperature at about 25 deg.C (20-30 deg.C). When having already finishedAfter all the sodium tert-butoxide has been added, stirring is carried out and the temperature is maintained for about 15 to 45 minutes. The reactor was then cooled to about-10 ℃ (-13 ℃ to 0 ℃) and a solution of diethyl p-toluenesulfonyloxymethylphosphonate (2.25 kg, calculated on purity) in DMF (1.22kg) was added over about 5-10 hours. The mixture is maintained at about-5 ℃ (-10 ℃ to 0 ℃) until the reaction is complete, which is typically about 0.5 to 2 hours after the last portion of diethyl p-toluenesulfonyloxymethylphosphonate is added. The completion of the reaction was monitored by HPLC [ residual 9- (2-hydroxyethyl) adenine does not exceed 3% ]. Glacial acetic acid (0.67kg) was added to control the temperature of the reaction vessel below 20 ℃. The mixture was stirred at about 22 deg.C (15-25 deg.C) for about 15-45 minutes. The cooled mixture was concentrated in vacuo until the distillation was complete, then the contents were cooled to below 40 ℃. Dichloromethane (16.0kg) was added and the contents stirred at 20 ℃ (15-25 ℃) for at least 1 hour. Such as DMF content relative to total solids [ NaOTs (sodium toluene sulphonate), NaOAc, Et2PMEA > 20% (from)1H-NMR), the mixture is concentrated in vacuo until the distillation is complete, the contents are cooled to below 40 ℃, dichloromethane (16kg) is added, and the reactor contents are stirred at about 20 ℃ (15-25 ℃) for at least 1 hour. Diatomaceous earth (0.5kg) was added and the contents of about 20 deg.C (15-25 deg.C) were stirred for at least 1 hour. The solid was filtered off and washed 3 times with dichloromethane (1 kg each). The filtrate and rinse, which did not exceed 80 ℃, were concentrated in vacuo until the distillation was complete, the reactor contents were cooled to 40 ℃, dichloromethane (5.0kg) was added to the residue and the contents of about 25 ℃ (20-40 ℃) were stirred to dissolve the solids. The resulting solution not exceeding 80 ℃ is concentrated in vacuo until the distillation is complete. Dichloromethane (7.0kg) was added and the contents of about 25 c (20-40 c) were stirred to dissolve the solids. If the DMF content is greater than 12% compared to diethyl PMEA, the mixture not exceeding 80 deg.C is concentrated in vacuo, the contents are cooled to below 40 deg.C, dichloromethane (7.0kg) is added and the contents at about 25 deg.C (20-40 deg.C) are stirred to dissolve the solids. The mixture was washed by adding water (0.8kg) to the mixture and stirring at about 25 deg.C (22-30 deg.C) for about 15-45 minutes. Allowing the mixture to stand for 4 hours, allowing the mixture to phase separate, separating the aqueous and organic phases. The aqueous phase was back-extracted 2 times with dichloromethane (1.5 kg each) as follows: stirring for 15-45 minutes, maintaining the solution at about 25 deg.C (22-30 deg.C), and allowing the phases to separate for at least 2 hours. The combined organic phases, which do not exceed 80 ℃, are then concentrated in vacuo until the distillation is complete. Toluene (3.0kg) was added thereto, and the mixture was stirred at about 25 ℃ C (22-30 ℃ C.) for about 15-45 minutes, and the obtained mixture at not more than 80 ℃ C was subjected to vacuum azeotropy. Toluene (3.0kg) was added and the mixture was heated to about 80 deg.C (75-85 deg.C), stirred for about 15-45 minutes, cooled to less than 30 deg.C over about 60-90 minutes, and then cooled to about 0 deg.C (-3 deg.C to 6 deg.C). After gentle stirring at about 0 ℃ for at least 12 hours, the resulting slurry was filtered and the filter cake was rinsed 3 times with cold (about 0-6 ℃) toluene (about 0.2kg each). The wet cake was dried under vacuum at about 50 deg.C (35-65 deg.C) and the dried product was ground. The drying of the product was monitored as a function of the water removal (water content not exceeding 0.3% as determined by Karl Fischer titration). An inert atmosphere is maintained throughout step 3.
Method for synthesizing PMEA
In one example, the synthesis of PMEA as shown in step 4 of scheme a is described as follows. A mixture of diethyl PMEA (1.00kg), acetonitrile (2.00kg) and trimethylbromosilane (1.63kg) was heated to reflux temperature in a reactor under an inert atmosphere (e.g., nitrogen) and maintained under stirring for about 1-3 hours until the reaction was complete. By using31P-NMR or HPLC (no diethyl PMEA was detected and no more than 2% monoethyl PMEA was detected) monitored for completion of the reaction. The solution was vacuum distilled to a semi-solid at ≤ 80 deg.c, then placed in water (2.00kg), warmed to about 55 deg.c (52-58 deg.c) and stirred for about 30-60 minutes to dissolve all solids. The resulting mixture was cooled to about 22 deg.C (19-25 deg.C), adjusted to pH3.2 with aqueous sodium hydroxide, the contents were heated to about 75 deg.C (72-78 deg.C) until the consistency became less (about 15-120 minutes), cooled to about 3 deg.C (0-6 deg.C), and stirred for at least 3 hours (3-6 hours). The slurry was filtered and the filter cake was rinsed with water (1.00 kg). The wet cake was suspended in water (3.75kg) and the suspension was heated to about 75 deg.C (72-78 deg.C) with vigorous stirring. After about 2 hours of stirring, the slurry was cooled to about 3 deg.C(0-6 ℃) and stirring for at least 2 hours. The slurry was filtered and the filter cake was rinsed 2 times with water (0.50 kg each) and acetone (1.00kg each), respectively. The isolated solid was dried under vacuum at a temperature not exceeding about 90 ℃ to a low water content (water content not exceeding 0.5% as determined by Karl Fischer titration) to yield a white crystalline PMEA. The product was ground to fine particle size.
Method for synthesizing AD
A representative method of preparing AD involves suspending 1 molar equivalent of PMEA in NMP in a volume ratio of about 1: 5.68-56.8, and then adding about 2-5 molar equivalents (often about 2.5-3.5, usually about 3 molar equivalents) of triethylamine ("TEA") to the solution with mild to moderate stirring. Next, about 3 to about 6 molar equivalents (often about 4.5 to about 5.5 molar equivalents, usually about 5 molar equivalents) of chloromethyl pivalate is added to give a reaction mixture. The preparation of the reaction mixture is usually carried out at room temperature. The reaction mixture is heated and maintained at a temperature below 66 c (typically about 28-65 c, usually at about 55-65 c) to allow the reaction to proceed for about 2-4 hours. The time required to heat the reaction mixture to about 28-65 ℃ is not critical and can vary depending on the volume of the reaction mixture and the capacity of the apparatus used to heat the mixture. Mild to moderate agitation keeps the solids in suspension during the reaction and minimizes splashing of reactants around in the reaction vessel. The product resulting from the present process contains AD produced by the process of reacting the listed reactants, typically under the given conditions.
In one example, the conversion of PMEA to AD shown in step 5 of diagram a is described as follows. A mixture of 1-methyl-2-pyrrolidone (3.15kg), PMEA (1.00kg), triethylamine (1.11kg) and chloromethyl pivalate (2.76kg) was heated to about 60 + -3 deg.C (not more than 66 deg.C) in a reactor under an inert atmosphere (e.g., nitrogen) with moderate stirring for 4 hours (1-4 hours) or less until the temperature reached31P-NMR or HPLC (mono (POM) PMEA not exceeding 15%) showed the reaction was complete. The mixture was diluted with isopropyl acetate (12.00kg) and cooled to 25 + -3 deg.C and stirred for about 30 minutes. Filtering off solidsWashed with isopropyl acetate (5.0 kg). The organic phases were combined and washed with water (3.70 kg each) by gentle stirring at 25 + -3 deg.C for about 15-45 minutes, repeated 2 times. The water washes were combined and back extracted with isopropyl acetate (4.00 kg each) by stirring at 25 + -3 deg.C for 15-45 minutes, repeated 2 times. The combined organic phases were washed with water (1.80kg) at 25 + -3 deg.C by stirring for 15-45 minutes, and then the organic phases were concentrated in vacuo at about 35 + -5 deg.C (not more than 40 deg.C) to about 40% of the original volume. After polish filtration (1 μm filter) and rinsing with 1.5kg of isopropyl acetate, the organic phase is concentrated further under vacuum at 35. + -. 5 ℃ C. (not more than 50 ℃ C.) until a pale oil is obtained. The pale oil typically contains about 6-45% (typically about 30-42%) AD.
Method for crystallizing AD
The crystallization of AD from organic oils is generally carried out by the following method: (1) using a relatively small volume of NMP (i.e., less than about 10mL/g PMEA) compared to the amount of PMEA as a reactant and/or (2) vacuum distillation for a sufficient period of time (i.e., at least about 4-6 hours) in the AD synthesis reaction to minimize the amount of isopropyl acetate that remains bound to the organic oil after vacuum distillation. The conglomerate of reaction starting materials (e.g., NMP or PMEA) in oil may comprise about 2-20% of the crystallization solution, but is generally less than about 1-2%. When crystals are prepared from organic oils, there is about 20-45% (often about 30-42%, usually about 35-42%) of AD in the oil prior to addition of the crystallization solvent.
AD can be crystallized, if desired, from, for example, a supersaturated solution. In such supersaturated solutions, nucleation occurs and easily leads to crystal formation. The rate of nucleation generally increases with increasing supersaturation and temperature. Supersaturated solutions are typically prepared by changing the temperature (usually by lowering it), evaporating the solvent, or changing the solvent composition (e.g., by adding a miscible non-solvent or poor solvent). Combining these methods (e.g., evaporation under reduced pressure, cooling the solution while increasing the solute concentration) also produces a supersaturated AD solution.
Crystalline AD is prepared by forming AD crystals in an AD composition, usually from a solution of AD in a crystallization mixture containing at least about 6% (typically at least about 30%, usually at least about 35%) AD. Typically, the crystallization is carried out by preparing a solution of AD containing about 6-45% AD and about 55-94% of a crystallization solvent. The upper limit of solubility of AD in most crystallization solvents at room temperature is about 10-41%. AD cannot be freely dissolved in certain crystallization solvents, such as those having a solubility of less than about 0.3mg/mL in di-n-butyl ether, and the addition of such solvents to AD solutions can increase the saturation or supersaturation of the solution. An organic solution containing an amount of AD close to the upper limit of solubility of AD in the crystallization solvent is generally used. The lower amount (about 6%) is the minimum amount of AD required in a solution that continuously produces crystals some solvents (such as methanol or dichloromethane) may contain more than about 50% AD.
The crystallization temperature is not critical and may vary as the crystallization process generally proceeds spontaneously over a temperature range. Crystallization above about 35 c (especially about 45-50 c) may result in decreased yield and/or increased impurities associated with crystallization. Crystallization is typically carried out at about-5 ℃ to about 50 ℃ (often about 0-35 ℃, usually about 4-23 ℃). It may be desirable to operate at less than about-5 ℃ to increase the crystallization yield or increase the rate of crystal formation, but low crystallization temperatures may increase by-products. Thus, in general, it is more convenient and economical to use solvents that are near ambient temperatures (about 15-23℃.) or typical cooling temperatures (0-4℃.) that most cooling devices or methods can easily achieve. When the solution contains a lower concentration (i.e., about 10-20%) of AD, crystallization at a lower temperature (i.e., about 0-15 ℃) tends to increase the crystallization yield.
Heating the solution containing AD and crystallization solvent above room temperature (preferably about 35 c) appears to aid crystallization, possibly due to an increased nucleation rate. The time for heating the crystallization mixture to about 35 ℃ is not critical and can vary depending on the capacity of the apparatus used, and is generally between about 20 and 45 minutes. The heating is then stopped and the temperature is reduced by cooling or allowing the temperature to drop back for about 10-120 minutes. During this time, crystals form and continue to form for at least about 4-36 hours. Crystallization is usually initiated immediately or shortly after the crystallization mixture reaches 35 ℃. Crystallization was carried out by allowing the temperature to drop to 0-23 ℃ after the solution reached 35 ℃. Crystallization is carried out with mild to moderate stirring or without stirring, typically with moderate stirring, generally giving good results.
Regardless of the solvent used, crystal formation can generally occur in about 5 minutes to about 72 hours, generally giving good results in about 10-16 hours. The crystallization time is not critical and can vary, although shorter crystallization times (about 30-90 minutes) can result in reduced recovery of AD. When a crystallization solvent is added to a reaction mixture containing another organic solvent (e.g., NMP), crystallization usually starts immediately when the temperature reaches about 35 ℃ or less, and the solution becomes cloudy.
The crystallization is carried out in customary laboratory or manufacturing plant apparatus, such as round-bottomed flasks, conical flasks, stainless steel reactors or glass-lined reactors. Mechanical agitation and temperature control devices of standard laboratory or commercial manufacturing scale are typically used in the crystallization.
When using a crystallization system comprising two different solvents, the most polar solvent is generally added to the AD first, followed by the least polar solvent. If insoluble components are present, they can optionally be removed from the solution after the first crystallization solvent has been added, for example by filtration or centrifugation. For example, when form 1 crystals are prepared from an organic solution containing AD and other components from the AD synthesis reaction using acetone and di-n-butyl ether, acetone is typically added first. Likewise, n-butanol can be added prior to the addition of di-n-butyl ether or ethyl acetate can be added prior to the addition of di-n-butyl ether. The solution containing the first polar solvent may become cloudy due to precipitation of the mono (POM) PMEA that may be present. The mono (POM) PMEA may then be removed from the solution by standard physical methods such as filtration or centrifugation, followed by addition of a second solvent such as di-n-butyl ether.
The crystallization solvent used to prepare the form 1 crystals generally contains less than about 0.2% water. When a significant amount (i.e., about 1-2%) of water is present in the crystallization solvent, a different amount of form 2 crystals are produced during the crystallization process and these form 2 crystals are also recovered along with the form 1 crystals. If desired, the amount of water in the crystallization reaction can be reduced by conventional means, including the use of anhydrous reagents or drying the solvent with molecular sieves or other known drying agents. Optionally, the amount of water that may be present in the organic solution containing AD (e.g., from an AD synthesis reaction with a solvent comprising by-products and organic oils as described above) can be reduced by using an azeotropic co-solvent (e.g., isopropyl acetate) to reduce the water prior to addition of the crystallization solvent.
In one example, the crystallization of form 1 AD shown in step 6 of diagram a is described as follows. The pale oil containing AD described above was dissolved in acetone (1.0kg) and heated to 35. + -. 3 ℃ and diluted in about 4 parts with di-n-butyl ether (5.00kg) while maintaining a temperature of about 32-38 ℃ and moderate stirring. The clear solution was cooled to about 25-30 ℃ over about 30-60 minutes (no more than 90 minutes), seeded with a small amount of form 1 AD crystals (about 5g) and then cooled to 22 ± 3 ℃ over 30-60 minutes (no more than 90 minutes) while maintaining moderate stirring. Moderate stirring is continued for at least about 15 hours at 22 ± 3 ℃. The resulting slurry was filtered and the filter cake washed with a premix of acetone (0.27kg) in di-n-butyl ether (2.4kg) (1: 9 v/v). If desired, premixed acetone (0.57kg) and di-n-butyl ether (4.92kg) can be added and the wet solid further purified by maintaining the temperature of the wet solid at 22. + -. 3 ℃ for about 15-24 hours with stirring. The solid was then filtered off and the filter cake was washed with premixed acetone (0.27kg) and di-n-butyl ether (2.4 kg). The filter cake, maintained at ≤ 35 deg.c (about 25-35 deg.c), is vacuum dried for about 1-3 days (LOD not exceeding 0.5%) to yield AD as a white to off-white powdery solid in form 1. The dried product was ground.
The present invention includes a method of preparing the form 2 crystal. The form 2 crystals can be conveniently prepared by hydrating the form 1 crystals, although hydrates can be obtained by crystallizing AD from a crystallization solvent containing a certain amount of water. The amount refers to an amount of water that does not interfere with crystallization but provides the water needed for hydration. The water may be present in the form of ice, liquid water or water vapor. Typically, water is physically contacted with the form 1 crystals under conditions such that form 2 crystals are formed. If desired, the form 1 crystals can be contacted with water vapor in a gas having a relative humidity of at least about 75% (e.g., air, carbon dioxide, or nitrogen) to completely convert the form 1 crystals to form 2 crystals. The form 1 crystals are generally contacted with air having a relative humidity of at least about 75% at about 18-30 deg.C (or typically at room temperature) for about 1-10 days to completely convert them to form 2. However, the form 1 crystal at room temperature was substantially non-hygroscopic in air having a relative humidity of 54%, and the water content did not increase after standing for 13 days.
The hydration of form 1 crystals to form 2 crystals results in a composition comprising a mixture of form 1 and form 2 AD crystals, wherein the proportion of form 1 AD crystals is about 100% to 0% and the remainder of the AD is form 2 crystals. Thus, during the conversion, the proportion of form 2 crystals increased from 0% to 100%. These compositions may include various formulations (e.g., tablets).
As indicated above, the form 2 crystals can also be prepared by the following method: AD is crystallized in the presence of water (e.g., about 2-5% water in the crystallization solvent used to prepare form 1 crystals). As with the former form 1 crystal, the formation of the crystal takes substantially about 4 to 36 hours at, for example, about 0 to 23 ℃. Such formulations may contain some form 1 crystals, but if desired, any residual form 1 crystals may be converted to form 2 crystals by exposure to moisture or by adding sufficient water to the crystallization solvent, as described above.
The method of preparing form 3 crystals is generally to grow the crystals in anhydrous methanol solution of AD. AD in methanol may be obtained by mixing sufficient amorphous or crystalline AD in methanol at room temperature for about 10-15 minutes or optionally dissolving solid AD to obtain a solution of at least about 100-150mgAD/mL methanol. The solubility of AD in methanol at room temperature was greater than 600 mg/mL. Crystallization is then carried out at about-5 ℃ to about 25 ℃ (typically at about 0-23 ℃) for about 4-48 hours.
The crystals obtained with isopropyl acetate as the only crystallization solvent are usually predominantly rod crystals with a few needle crystals, which may be longer, i.e. up to about 500 μm in length. FIG. 8 is a bar of about 20-500 μm in length obtained by crystallization in isopropyl acetate at about 15 ℃.
Optionally, crystallization from supersaturated and saturated or certain unsaturated AD solutions can be promoted or accelerated by adding AD seeds to the solution (although the addition of seeds is not mandatory). For example, form 1 AD can be obtained by adding a small amount of crystalline form 1 AD to the above organic solution (e.g., organic oil to which crystallization solvent has been added) without heating to 35 ℃. The crystals added as seed crystals promote the formation of form 1 crystals. Similarly, form 2 and form 3 crystals can be obtained by adding seed crystals to a suitable solution having various crystal forms, for example, form 2 crystals can be obtained from an organic solution containing ethyl acetate and about 2% water, or form 3 crystals can be obtained from a saturated solution of AD in anhydrous methanol. The amount of crystals used as seed crystals can be varied as desired to obtain optimum results. Generally, it is sufficient to add about 0.1 to 1.0g of crystals per liter of AD recrystallization solution.
If desired (e.g., to improve the purity of the crystals), crystalline AD can be recrystallized.
For example, form 1 AD is recrystallized in essentially the same manner as described above for the preparation of form 1 crystals. For example, recrystallization from acetone and di-n-butyl ether is carried out by dissolving crystalline AD in acetone at a ratio of about 0.2 to 0.4g/mL at about 20 to 35 ℃ and then, if necessary, removing insoluble components, for example, by filtration or centrifugation of the solution, which is usually turbid. The insoluble component is typically a mono (POM) PMEA. Next, the solution is warmed to about 35-40 ℃ and about 5.2-6.2mL (usually about 5.7mL) of warm (about 35-40 ℃) di-n-butyl ether is added for every 0.2-0.4g of crystals that were originally used for recrystallization. The recrystallized mixture was then allowed to cool to room temperature over about 4-4.5 hours. If a smaller volume is used (e.g., about 1-3L), the recrystallization mixture will cool rapidly to room temperature. The time for cooling the mixed liquor is not critical and can vary.
Recrystallization generally begins immediately after the addition of di-n-butyl ether and mixing is complete, and recrystallization may be allowed to proceed for about 4 to 36 hours (typically about 6 to 24 hours). After recrystallization at room temperature for about 4-36 hours, some additional crystallization is usually obtained by cooling the recrystallization mixture to about 4-10 c and allowing the mixture to stand at this reduced temperature for about 1-6 hours. Typically, the amount of AD used in recrystallization will be sufficient to form a saturated or near saturated solution, i.e., about 0.4g/mL when the solvent is acetone. AD was completely dissolved in acetone in about 2-8 minutes with moderate stirring. The material that remains undissolved after this initial mixing period is removed and discarded, and the second less polar solvent of the solvent pair is then added to the mixture containing the first crystallization solvent.
If desired, the form 1 crystals can be recrystallized from a single solvent such as acetone. In this example, sufficient crystals are dissolved in a solvent at room temperature to give a saturated or near saturated solution, and then the insoluble components are removed. Then, in the same manner as the above recrystallization using acetone and di-n-butyl ether solvent, the mixture was warmed to 35 ℃ and allowed to cool.
The recrystallization of the form 2 crystals will proceed in the same manner as the recrystallization of the form 1 crystals, but form 2 crystals dissolved in a recrystallization solvent will be used. If desired, the form 1 crystals obtained by recrystallization can be converted to form 2 crystals, as described herein for the form 1 crystals to form 2 crystals. The form 2 crystal may be recrystallized into the form 1 crystal. In this case, if desired, molecular sieves or other solvent drying means may be used to limit the amount of water present after dissolution of the form 2 crystals in the first solvent and during recrystallization. The form 2 crystals can also be recrystallized from a solvent containing about 1-2% water to directly obtain form 2 crystals.
The form 3 crystals were recrystallized from methanol in the same manner as the preparation of the form 3 crystals described herein. Crystals were prepared with a saturated or nearly saturated (i.e., at least about 0.6g/ml) methanol solution of AD.
Salts can be prepared, if desired, by acid addition of certain organic and inorganic acids to the basic center in adenine in AD. The acid addition salts are generally prepared by standard methods involving dissolving the AD free base in an aqueous, hydro-alcoholic or hydro-organic solution containing the selected acid or counter ion of the acid, optionally crystallizing it, and optionally simultaneously evaporating, stirring or cooling the solution. The free base is usually reacted in an organic solution containing an acid or counter ion, in which case the salt is usually isolated directly, or may be added to the solution as a seed crystal or concentrated to facilitate precipitation of the salt. Examples include those containing AD, a solvent (usually a crystallization solvent), and a sulfonic acid (e.g., C)6-16Arylsulfonic acids, C4-16Heteroaryl sulfonic acids or C1-16Alkyl sulfonic acid). Examples also include solutions comprising AD, a solvent (typically a crystallization solvent), and a carboxylic acid (e.g., a tricarboxylic acid, a dicarboxylic acid, a monocarboxylic acid, any of these carboxylic acids containing from about 1 to about 12 carbon atoms).
Pharmaceutical formulations and routes of administration
The compositions of the invention containing crystalline AD (typically crystalline form 1, hereinafter referred to as the active ingredient) are administered by any route suitable for the condition being treated, including oral, rectal, nasal, topical (including ocular, buccal and sublingual), vaginal and parenteral (including subcutaneous, intramuscular, intravenous, intradermal, intrathecal and epidural) administration. Typically, the compositions of the present invention are administered orally, but compositions containing crystalline AD can be administered by any of the other methods described above.
Although AD may be administered as a pure compound, it is preferably in the form of a pharmaceutical preparation. The formulations of the present invention comprise AD and one or more pharmaceutically acceptable excipients or carriers ("desirable excipients"), and optionally, other therapeutic ingredients. Excipients must be "desirable" in that they are compatible with the other ingredients of the formulation and not injurious to the patient.
The formulations may be those suitable for topical or systemic administration, for example those suitable for oral, rectal, nasal, buccal, sublingual, vaginal or parenteral (including subcutaneous, intramuscular, intravenous, intradermal, intrathecal and epidural) administration. The formulations are in unit dosage form and are prepared by any of the methods well known in the pharmaceutical art. These methods include the step of admixing the active ingredient with the carrier or excipient that constitutes one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then, if necessary, drying or shaping the product.
Formulations of the present invention suitable for oral administration may be provided in the following forms; discrete units such as sachets, sachet or tablet each containing a predetermined amount of active ingredient; powder or granules; solutions or suspensions in aqueous or non-aqueous liquids; or an oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The active ingredient may also be provided in the form of a bolus, electuary or paste.
The formulations of the present invention include compositions comprising AD and desirable excipients. These excipients include binders, diluents, disintegrants, preservatives, dispersants, glidants (anti-adherents) and lubricants. These compositions may also contain unit dosage forms, including tablets and capsules, as desired. If desired, these compositions may be tablets containing about 5 to 250mg (typically about 5 to 150mg) of AD, including tablets containing about 60 or 120mg of AD per tablet. These tablets may contain, if desired, about 1-10% binder, about 0.5-10% disintegrant, about 5O-60% diluent, or about 0.25-5% lubricant. These compositions may also be wet granules containing a liquid (e.g., water), AD, and one or more desirable excipients selected from the group consisting of binders, diluents, dispersants, and disintegrants.
Tablets may be formed by compression or molding, optionally with one or more accessory ingredients or excipients. Tablets typically contain about 5-250mg (usually about 30-120mg) AD per tablet (usually predominantly form 1 AD), for example about 60mg or 120mg of form 1 AD per tablet, with only a limited amount (usually less than about 20%) of form 2 crystalline, other crystalline types or amorphous AD present. Compressed tablets may be prepared by compressing, in a suitable machine, the AD in a flowable form such as a powder or granules, optionally mixed with a binder, disintegrant, lubricant, inert diluent, preservative, surfactant or dispersing agent. Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound, usually moistened with a liquid diluent. If desired, the tablets may be coated and printed, embossed or scored, and may be formulated so as to provide sustained or controlled release of the active ingredient therein. Examples include products made by the following process: a mixture containing crystalline AD (typically crystalline form 1 or form 2) and desirable excipients (e.g., dried wet granules containing, for example, lactose, pregelatinized starch, croscarmellose sodium, talc, and magnesium stearate) is compressed.
The compositions containing crystalline AD and excipients may also contain L-carnitine or a salt of L-carnitine (e.g. L-carnitine-L-tartrate (2: 1)). The in vivo release of pivalic acid from the pivaloyloxymethyl moiety of AD appears to decrease the concentration of L-carnitine in the patient. Tablets containing L-carnitine-L-tartrate and AD reduce the effect of pivalic acid in reducing L-carnitine in patients taking AD. The clinician will understand the amount of L-carnitine that needs to be added after considering the degree of depletion of L-carnitine in the patient.
Typical formulation ingredients for tablets or related dosage forms include one or more binders, diluents, disintegrants or lubricants. These excipients may increase the stability of the formulation, aid in tablet compression during manufacture, or aid in disintegration of the formulation after ingestion. Tablets are generally prepared by the following method: one or more excipients are mixed with AD and wet granulated, and the resulting granules are then wet milled and dried to a loss on drying of less than about 3%. If desired, there may be about 1-10% binder (e.g., pregelatinized starch or povidone which can improve the processability of the tablet). Optionally, there may be about 0.5-5% of a disintegrant (e.g., croscarmellose sodium or microcrystalline cellulose) to facilitate dissolution of the tablet. If desired, there may be about 40-60% diluent (e.g., mono-or di-saccharides) to mask the physical properties of AD or to facilitate tablet dissolution. Optionally, there may be about 0.25-10% of a lubricant (such as magnesium stearate, talc or silicon dioxide) to facilitate tablet ejection during manufacture. Optionally, the tablet may contain a scavenger (e.g., lysine or gelatin) to capture formaldehyde that may be released during storage of AD. Excipients are described, for example, in the following sections of the Handbook of pharmaceutical Excipients (2 nd edition 1994), compiled by the American pharmaceutical Association: a piece of pregelatinized starch (page 491-493), a piece of croscarmellose sodium (page 141-142), a piece of lactose monohydrate (page 252-261), a piece of talcum powder (page 519-521) and a piece of magnesium stearate (page 280-282).
Typical containers storing form 1 AD formulations can limit the amount of water in the container. A desiccant (such as silica gel or activated carbon or both) is typically included in the unit formulation or dose. The container is typically induction sealed. Encapsulation of silica gel alone is sufficient to keep the AD-containing tablets dry when stored at ambient temperature. There are two pivaloyloxymethyl moieties in one molecule of AD. Thus, silica gel is suitable as a single desiccant for compounds such as therapeutic agents containing one or more pivaloyloxymethyl moieties. The water permeability of the container is described, for example, in chapter 23 "container-permeate" of the United states Pharmacopeia convention of Rockville, Md.
For infections of the eye or other external tissues (e.g. mouth and skin), it is preferred that the formulation is administered in the form of a topical ointment or cream containing, for example, 0.01 to 10% by weight of the active ingredient (including amounts of active ingredient between 0.1 and 5% in 0.1% by weight increments such as 0.6%, 0.7% by weight, etc.), preferably 0.2 to 3% by weight, most preferably 0.5 to 2% by weight. When formulated as an ointment, the active ingredient may be used with a paraffinic or water-miscible ointment base. Alternatively, the active ingredient may be formulated as a cream with an oil-in-water cream base.
If desired, the cream-based aqueous phase may comprise, for example, at least 30% by weight of a polyhydric alcohol [ i.e., an alcohol having two or more hydroxyl groups such as propylene glycol, 1, 3-butylene glycol, mannitol, sorbitol, glycerol, and polyethylene glycols (including PEG400) and mixtures thereof ]. Topical formulations may suitably contain compounds which promote absorption or penetration of the active ingredient through the skin or other site of action. Examples of such transdermal enhancers include dimethyl sulfoxide and related analogs.
The oily phase of the emulsions of the invention may be constituted in known manner by known ingredients. Although the oil phase may consist of emulsifiers alone, it is preferred to consist of a mixture of at least one emulsifier with a fat or oil or both. Preferably, the oil phase contains a hydrophilic emulsifier and a lipophilic emulsifier as a stabilizer. Both oils and fats are also preferred. At the same time, the emulsifier with or without stabilizer constitutes the emulsified wax, while the wax with oil and fat constitutes the emulsified ointment base, which forms the oily dispersed phase of the cream.
Emulsifiers and emulsion stabilizers suitable for use in the formulations of the present invention include Tween60、Span80. Cetostearyl alcohol, benzyl alcohol, tetradecanol, glyceryl monostearate, and sodium lauryl sulfate.
The oils or fats suitable for use in the formulation are selected based on whether the desired cosmetic properties are obtained. Thus, it is preferred that the cream be a non-greasy, non-staining and washable product with a suitable consistency to avoid leakage from the cartridge or other container. Straight or branched chain, mono-or dibasic alkyl esters such as di-isoadipate, isocetyl stearate, propylene glycol diester of coconut fatty acid, isopropyl myristate, nonyl oleate, isopropyl palmitate, butyl stearate, 2-ethylhexyl palmitate or a blend of branched chain esters known as Crodamol CAP may be used, the last three esters being preferred. They may be used alone or in combination depending on the desired properties. Alternatively, high melting point greases such as white soft paraffin and/or liquid paraffin or other mineral oils may be used.
Formulations suitable for topical administration to the eye also include eye drops wherein the active ingredient is dissolved or suspended in a suitable carrier, especially an aqueous solvent suitable for the active ingredient. The concentration of active ingredient in these formulations is preferably from 0.01% to 20%, in some embodiments from 0.1% to 10%, and in other embodiments, about 1.0% by weight.
Formulations suitable for topical administration in the mouth include lozenges comprising the active ingredient in a flavoured base, usually sucrose and acacia or tragacanth; lozenges comprising the active ingredient in an inert base (such as gelatin and glycerin, or sucrose and acacia); and mouthwashes containing the active ingredient in a suitable liquid carrier.
Formulations for rectal administration may be presented as suppositories with suitable bases containing, for example, cocoa butter or a salicylate.
Formulations wherein the carrier is a solid, suitable for nasal or inhalation administration include powders having a particle size, for example, in the range 1 to 500 μm (including particles having a particle size in the range 20 to 500 μm, in 5 μm increments, such as 30 μm, 35 μm, etc.). Formulations wherein the carrier is a liquid, suitable for administration, for example, as a nasal spray or nasal drops, include aqueous or oily solutions of the active ingredient. Formulations suitable for aerosol administration may be prepared by conventional methods and may be administered with other therapeutic agents. Inhalation-type therapeutic agents can be easily administered by means of a metered dose inhaler.
Formulations suitable for vaginal administration may be presented as pessaries, plugs, creams, gels, pastes, foams or sprays containing the active ingredient together with a suitable carrier as is known in the art.
Formulations suitable for parenteral administration are sterile and include aqueous and non-aqueous injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; aqueous and non-aqueous sterile suspensions, which may contain suspending agents and thickening agents. The formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials with elastomeric stoppers, and may be stored in a freeze-dried condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described. Preferred unit dosage formulations are those containing a daily or unit daily divided dose (as described above) of the active ingredient or a suitable fraction thereof.
In addition to the ingredients specifically mentioned above, the formulations of the present invention may contain other agents conventional in the art having regard to the type of formulation in question, for example, formulations suitable for oral administration may contain flavoring agents.
The present invention also provides a veterinary composition comprising at least one active ingredient as defined above and a veterinary carrier.
Veterinary carriers are substances useful for the purpose of administering the composition to cats, dogs, horses, rabbits and other animals, and may be solids, liquids or gases which are inert or desirable in the veterinary art and which are compatible with the active ingredient. These veterinary compositions may be administered orally, parenterally or by any other desired route.
The compounds of the present invention may be used to provide controlled release pharmaceutical formulations comprising a matrix or absorbent material and, as an active ingredient, one or more compounds of the present invention, wherein the release of the active ingredient is controlled and modulated to reduce the frequency of administration or to improve the pharmacokinetic or toxicity profile of the compound. Controlled release formulations suitable for oral administration, wherein the discrete units contain one or more compounds of the invention, may be prepared by conventional methods.
All documents cited herein are incorporated herein by reference in their entirety.
The following examples are further illustrative of the present invention and are not intended to be limiting.
Example 1: preparation of form 1 crystals
To a 500mL single neck round bottom flask equipped with a magnetic stir bar was added PMEA (27.3g, 100 mmol). N-methylpyrrolidone (109.3mL) and triethylamine (50.6g, 69.8mL, 500mmol) were added under nitrogen and the resulting suspension was stirred vigorously. Chloromethyl pivalate (75.2g, 72.0mL, 500mmol) was added and the stirred suspension was placed in a 45 ℃ oil bath for 18.5 hours. The resulting thick, pale yellow suspension was diluted with isopropyl acetate (1.0L) and stirred for 1 hour. The solids were filtered off by filtration ("Kimax sintered glass plate funnel No. C") and washed with isopropyl acetate (250 mL). The washings and the filtrate were combined, and the resulting organic phase was extracted with water (200 mL. times.2). The aqueous extracts were combined and back-extracted with isopropyl acetate (250 mL. times.2). All organic phases were combined to give 1975 mL. Isopropyl acetate was added to bring the total volume of the organic phase to 2.0L. For internal control of this experiment, the organic phase was divided into two 1.0L aliquots. One of the portions was washed with brine and treated with sodium sulfate, while the other portion was treated without these steps (see below).
A1.0L aliquot of the organic phase used for the novel process was concentrated directly using a standard (Buchi) rotary evaporator to an oil which was completely transparent and no salt was visible during the entire process, at a bath temperature of 45 ℃ and a vacuum of 50-70 mmHg. the weight of the oil was 32.4 g. The oil was diluted with acetone (25mL) to give a completely clear solution without visible precipitation of the salt. After standing at room temperature for about 3 hours, the solution was still completely transparent. This solution was put in an oil bath set at 45 ℃ and di-n-butyl ether (140mL) was gradually added to maintain the internal temperature at about 40 ℃. The flask was then removed from the oil bath, allowed to cool to room temperature and stirred at room temperature for about 16 hours, as a result, form 1 AD precipitated. The solid product was collected by filtration ("Kimax sintered glass plate funnel No. M"). And the solid was washed with a solution of 10% acetone in 90% di-n-butyl ether (v/v, 40mL) and dried in a vacuum oven (ambient temperature, nitrogen purge, 28 "vacuum). 12.2g (48.8% of theory yield on the 50mmol reaction scale) of a white solid were obtained, the purity of the AD determined by HPLC to be 99.8%, based on external standard.
The remaining 1.0L of organic phase was used as a control for the above results and was treated as follows. The organic phase was washed with brine (25mL), dried over sodium sulfate (25g, 12 hours dry time) and concentrated as before. 27.4g of an oil are obtained which is crystallized from acetone (25mL) and butyl ether (135mL) as described above. The resulting solid was collected by filtration and dried as described above to give 12.3g (48.9% of theory) of a white solid with a purity of 98.7% for the AD as determined by HPLC based on external standard.
Example 2: preparation of form 1 crystals
9.7kg of NMP were added to 3kg of PMEA in a 30-gallon enamelled steel reaction vessel (product of Pfaudler, Rochester, N.Y., model No. P20-30-150-. The moderate agitation used was sufficient to keep the solid PMEA in suspension and to prevent splashing of the reactor contents on the walls. Then 5.6kg of TEA and 8.3kg of chloromethyl pivalate were added. Then, 2.7kg of NMP was further added, and the residual material from the transfer line feeding the reactor was washed. The temperature was adjusted to about 48 ℃ and maintained at 38-48 ℃ for 18 hours with moderate stirring. After the reaction is complete, 48kg of isopropyl acetate are added to the reactor at room temperature and filtered (Tyvek)TMFilter, diameter 15.5 ", product of Kavon Filter Products, Wall, N.Y.,the model is as follows: no.1058-D) the resulting mixture was maintained at 43-48 ℃ for 1 hour with moderate stirring before removing the solids. The contents of the reactor were washed out with 12kg of isopropyl acetate and filtered. The filtrate was transferred to a 50-gallon enamel steel reaction vessel (product of Pfaudler, Inc., model: No. P24-50-150-. In the subsequent step, the temperature is allowed to fall back to ambient temperature.
The mixture was then washed with 22kg of water and vigorously stirred for about 1.5-2 minutes. Stirring was stopped and the phases were allowed to separate completely (about 10 minutes). The lower aqueous phase (about 26L) was transferred to a 30 gallon enamel steel reaction vessel. 22kg of water was then added to the organic phase remaining in the 50 gallon reactor and the reactor contents were vigorously stirred for about 1.5-2 minutes. Stirring was stopped and the phases were allowed to separate completely (about 1 hour 40 minutes). The lower aqueous phase was transferred to a 30 gallon enamel steel reaction vessel, which contained two portions of aqueous rinse. To a 30 gallon reactor water wash was added 24kg of isopropyl acetate and the reactor contents were vigorously stirred for about 1.5-2 minutes, then stirring was stopped for a sufficient time to allow complete phase separation (about 10 minutes). The upper organic phase was retained and mixed with the organic phase previously retained in the 50 gallon reactor. To the water wash in the 30 gallon reactor was added 24kg of isopropyl acetate and the reactor contents were vigorously stirred for about 1.5-2 minutes, then stirring was stopped for a sufficient time to allow complete phase separation (about 20 minutes). The upper organic phase was retained and mixed with the organic phase previously retained in the 50 gallon reactor. The organic phases were combined and then washed with brine solution (7kg water, 3.9kg NaCl) and vigorously stirred for about 1.5-2 minutes, followed by stopping stirring to allow complete separation of the phases (about 5 minutes). The brine phase was discarded. 18kg of sodium sulfate was added to the reactor and the mixture was stirred vigorously for about 1.5-2 minutes and then allowed to stand for about 1 hour. At this time, the weight of the organic phase was 98.5 kg.
The reactor contents were gently stirred and filtered through a bag Filter (product of American Felt and Filter Co., model No. RM C S/S122). The AD containing organic phase was shifted to a clean 50 gallon reactor and vacuum distilled at 33-41 c and 26-30 "Hg vacuum to remove volatile organics until 50-55L condensate was collected. The organic phase was removed from the 50 gallon reactor to a clean 30 gallon reactor by vacuum filtration through a cartridge filter (Memtec America, model: No.910044) with a cotton gauze wrapped cartridge and washed with 8.6kg isopropyl acetate. The solution was kept at 5 ℃ overnight and then concentrated in vacuo at 26-41 ℃ for 3 hours to give about 7-9L of oil. To the oil was added 5.4kg of acetone to give a clear solution. The solution was then stirred and warmed to 43 ℃ and 27kg of room temperature di-n-butyl ether were added over about 4 minutes, and the solution was then warmed to 43 ℃ again. 15kg of di-n-butyl ether was added over about 4 minutes and the temperature of the solution was returned to 43-44 ℃ and then allowed to fall back to 20 ℃ over about 7 hours and 15 minutes. During this process, AD crystals are formed in the reactor. The crystals were recovered by filtration (Nutche filter) and dried. 2.40kg AD (45.1%) were obtained.
Example 3: preparation of form 1 crystals
546.3g PMEA (2 moles) was added to a 12L three-necked round bottom flask at room temperature followed by 2.18L NMP. Slow mechanical stirring (sufficient to suspend the solid PMEA but not splash the flask contents) was started to suspend the PMEA, then 1.39L of TEA was added to the flask, followed by 1.44L of pivaloyloxymethyl chloride. The flask was purged with nitrogen and the reaction was heated to 60 ℃ over 30-45 minutes. The reaction was held at 60 ℃ and gently stirred for 2-2.5 hours. The completion of the reaction was confirmed by HPLC. When the yield of AD reached 65-68% as determined by area normalization, 7.48L of cold (0-3 deg.C) isopropyl acetate was added to the flask and the reaction was stopped. The stirring was increased to moderate stirring (with moderate vortex but no splashing of the contents) and the mixture was maintained at room temperature for 30 minutes with moderate stirring, at which time solids (e.g., TEA. HCl, mono (POM) PMEA) precipitated out of solution.
Next, the reaction mixture was filtered through a glass fritted funnel (40-60 μm) and the filter cake was washed with 2.51L of isopropyl acetate at room temperature.
The filtrate was then extracted 2 times with 2.0L of drinking water at room temperature. The aqueous phases were combined and back-extracted 2 times with 2.51L of isopropyl acetate (room temperature). All organic phases were combined and extracted 1 time with 985mL of drinking water. The organic phase was separated and concentrated at a temperature of 35-39 c and under vacuum of about 30mmHg to give 1.24kg of a yellow oil.
The oil was transferred to a 12L three-necked flask and cooled to room temperature over about 30 minutes. To the flask was added 628mL of room temperature acetone followed by 3.14L of di-n-butyl ether. Slow stirring was started and the solution was heated to 35 ℃ over about 5-20 minutes. When the temperature reached 35 ℃, heating was stopped and the temperature did not continue to rise. The solution was cooled to below 30 deg.C (20-29 deg.C) over about 30 minutes. During cooling, form 1 crystals formed in the crystallized mixture while maintaining slow stirring, and stirring was continued slowly at room temperature for 14-20 hours. The crystals are then filtered off (Tyvek)TMFilter) and the filter cake was washed with 2L of a 10% acetone/90% di-n-butyl ether (v/v) solution. The filter cake was dried in a nitrogen purged oven at room temperature until its weight reached constant (about 2 days).
The yield of AD of form 1 obtained is 50-54% of the theoretical value calculated from PMEA, and the purity by HPLC according to area normalization is 97-98.5%.
Example 4: preparation of form 1 crystals
273.14g of PMEA (1 mole) was added to a 3L three-necked round bottom flask at room temperature followed by 1.09L of NMP. Slow mechanical stirring (sufficient to suspend the solid PMEA but not splash the flask contents) was started to suspend the PMEA, then 0.418L of TEA (3 equivalents) was added to the flask, followed by 0.72L of pivaloyloxymethyl chloride (5 equivalents). The flask was purged with nitrogen and the reaction was heated to 60 ℃ over 30-45 minutes. The reaction was held at 60 ℃ and gently stirred for 2-2.5 hours. The completion of the reaction was confirmed by HPLC. When the yield of AD is determined to reach 68-70% by an area normalization methodIn this case, 3.74L of cold (0-3 ℃ C.) isopropyl acetate was added to the flask to terminate the reaction. The stirring was increased to moderate stirring (with moderate vortex but no splashing of the contents) and the mixture was allowed to stand at room temperature for 30 minutes with moderate stirring, at which time solids (e.g., TEA HCl, mono (POM) PMEA) precipitated out of solution. The reaction mixture was filtered through a glass fritted funnel (40-60 μm) and the filter cake was washed with 1.26L of isopropyl acetate at room temperature. Subsequently, the filtrate was extracted with 1.01L of drinking water at room temperature for 2 times. The aqueous phases were combined and back-extracted 2 times with 1.26L of isopropyl acetate (room temperature). All organic phases were combined and extracted 1 time with 492mL of drinking water. The organic phase was separated and concentrated at a temperature of 35-39 c and under vacuum of about 30mmHg to give 0.6kg of a yellow oil. The oil was transferred to a 3L three-necked flask and cooled to room temperature over about 30 minutes. To the flask was added 314mL of acetone (room temperature), followed by 1.57L of di-n-butyl ether. Slow stirring was started and the solution was heated to 35 ℃ over about 5-20 minutes. When the temperature reached 35 ℃, heating was stopped and the temperature did not continue to rise. The solution was cooled to below 30 deg.C (20-29 deg.C) over about 30 minutes. During the cooling process, form 1 crystals were formed in the crystallized mixture while maintaining slow stirring. Then, 1.15L of di-n-butyl ether was added to the crystallized mixture (room temperature). Stirring was continued slowly at room temperature for about 16 hours. The crystals are filtered off (Tyvek)TMFilter), the filter cake was washed with 1L of a 10% acetone/90% di-n-butyl ether (v/v) solution, and then the solution was filtered off. The filter cake was dried in a nitrogen purged oven at room temperature until its weight reached constant (about 2 days).
The yield of AD of form 1 obtained is 55-58% of theoretical value calculated from PMEA and the purity measured by HPLC according to area normalization is 99-100%.
Example 5: preparation of AD crystals using isopropyl acetate as crystallization solvent
To a 500mL three-necked round bottom flask in PMEA (10.93g) equipped with a stirring device was added 43.7mL of NMP at room temperature under a nitrogen atmosphere. The mixture was stirred to suspend the PMEA. Subsequently, TEA (27.9mL) was added at room temperature followed by pivaloyloxymethyl chloride (28.9mL) at room temperature. The temperature was raised to 45 ℃ and the suspension was stirred at 45 ℃ for 12 hours. The resulting thick yellow suspension was diluted with isopropyl acetate (150mL) at room temperature and stirred vigorously at room temperature for 75 minutes. The solid was filtered through a "C" fritted glass plate funnel and washed with 50mL isopropyl acetate at room temperature. The filtrates were combined and washed 2 times with 40mL portions of deionized water. The combined aqueous washes were back-extracted with 40mL isopropyl acetate, 2 times. All organic phases were combined, washed 1 time with 20mL of deionized water, the aqueous and organic phases were allowed to separate and contact for 2 hours at 17 ℃. During this time, the formation of long rods was observed at the aqueous-organic phase interface. The crystals were collected by filtration using an "M" sintered glass plate funnel and dried to give 512mg of long rod crystals.
Example 6: analysis of AD by HPLC
Crystalline form 1 was analyzed by HPLC for AD to determine its purity, to isolate or identify byproducts, and to illustrate the use of byproducts as reference standards for AD. The level of compound present was analyzed by area normalization. HPLC analysis was performed within 12 hours of standard or sample preparation.
A liquid chromatograph equipped with a constant volume sample injector, a variable wavelength absorption detector and an electronic integrator was used, and the column used was an Alitech Mixer finger Exchange manufactured by Alltech corporation of Deerfield, IllinoisTMC8(7 μm, pore size 100)250mm x 4.6mm internal diameter) and the protective column used was also a product of Alltech (20mm x 4.6mm internal diameter, dry packed with Pellicular C8 particles). The water used is of chromatographic grade. The chemical used was chromatographic grade acetonitrile (Burdick, Muskegon, Mich.)&Product of Jackson corporation), monopotassium phosphate (KH) grade for anhydrous analysis2PO4Mallinckrodt, product of Paris, kentucky), water-free analytical grade dipotassium phosphate (K)2HPO4Mallinckrodt, part, kentucky) and a.c.s. reagent grade phosphoric acid (Mallinckrodt, part, kentucky). Prior to use, the aqueous potassium phosphate solution was filtered (0.45 μm nylon 66 membrane filter, product of Rainin, Woburn, Mass.) and degassed. Equivalents of these components and compounds may also be used. Equivalent devices and/or reagents may also be used to achieve similar results.
The mobile phase A consisted of potassium phosphate buffer at pH6.0 and acetonitrile 70: 30v/v, and was prepared by mixing 1400mL of 200mM potassium phosphate buffer (pH6.0) with 600mL of acetonitrile. The mobile phase B consisted of potassium phosphate buffer at pH6.0 and acetonitrile 50: 50v/v, and was prepared by mixing 1000mL of 200mM potassium phosphate buffer (pH6.0) with 1000mL of acetonitrile.
Before sample analysis, the HPLC column was equilibrated with mobile phase A at a flow rate of 1.2mL/min for 1 hour at room temperature. A5 uL sample of AD containing byproducts (approximately 1mg/mL solution) was analyzed at a flow rate of 1.2mL/min for 25 minutes as a protocol, and was converted to 100% mobile phase B after 19 minutes by using mobile phase A for the first 1 minute, followed by a linear gradient. The column was then held for 5 minutes at 100% mobile phase B.
The sample containing AD was prepared by accurately weighing about 25mg of AD sample or formulation and dissolving AD in a final volume of 25.0mL of sample solvent. The sample solvent was prepared by mixing 200mL of potassium phosphate buffer (3.40g of monopotassium phosphate/L of water, adjusted to pH3.0 with phosphoric acid) with 800mL of acetonitrile and equilibrating to room temperature. The compounds are identified by their elution time and/or retention time. In such gradient solvent systems, AD typically elutes at about 9.8 minutes. The mono (POM) PMEA eluted at about 6.7 minutes and PMEA eluted at about 3.5 minutes.
Example 7: physical characterization of the form 1 crystals
Form 1 crystals were analyzed by XRD by loading approximately 100-150mg of crystals onto an aluminum holder and then subjecting the aluminum to crystallizationThe rack was placed in a diffractometer (model GE XRD-5, automated operation by a Nicolet automated unit). Form 1 crystals were scanned at a scan rate of 0.05 deg./1.5 seconds between 4 and 35 degrees 2 theta by exposing the crystals to an X-ray generator operating at 40KV and-20 mA using a standard focused copper X-ray tube (variacan CA-8) with a graphite monochromator (product of ES Industries) and scintillation detector. The weighted average of the X-ray wavelengths used for the calculation is CuK alpha 1.541838. Form 1 AD crystals have characteristic XRD peaks (expressed in degrees 2 theta) at about 6.9, 11.8, 12.7, 15.7, 17.2, 20.7, 21.5, 22.5 and 23.3. A representative XRD pattern of form 1 is shown in figure 1.
The crystals of form 1 were also analyzed using a differential scanning calorimeter and the resulting thermogram is shown in FIG. 2 with a characteristic endothermic transition at about 102.0 ℃ and an onset temperature of about 99.8 ℃. Thermograms were obtained at a scanning rate of 10 ℃/min under nitrogen atmosphere. The samples were not sealed in containers in the DSC apparatus, but were analyzed in the DSC apparatus at ambient pressure. Calorimetric scans were obtained using a differential scanning calorimeter (TA Instruments, model 2910 DSC with model 2200 controller). Differential calorimetric profiles were obtained with about 5mg of AD. Differential scanning calorimetry has been described (see, for example, U.S. pharmacopoeia, volume 23, method 891, 1995, published by U.S. pharmacopoeia, Inc. of Rockville, maryland).
The melting point of the form 1 crystals was determined by conventional melting point analysis. The analysis was carried out using a CPU of Mettler FP90 equipped with a cell of FP81 type according to the manufacturer's instructions. The sample was equilibrated at an initial temperature of 63 deg.C for 30 seconds, and then the temperature was raised at a rate of 1.0 deg.C/min. The form 1 crystals melt between 99.1 and 100.7 ℃.
An infrared absorption (IR) spectrum of the form 1 crystals was obtained using a Perkin-Elmer model 1650 FT-IR spectrophotometer according to the manufacturer's instructions. Containing about 10 wt% (5mg) of form 1 crystals and about 90 wt%(50mg) dried (overnight at 60 ℃ under vacuum) potassium bromide (Aldrich, IR grade) in the form of translucent pellets was obtained by grinding the two powders together to give a fine powder. IR spectroscopy has been described (see, for example, U.S. pharmacopoeia, 23, 1995, methods 197, R.T. Morrison et al, organic chemistry, 3 rd edition, page 405-. Prior to scanning the sample, the spectrophotometer sample chamber is purged with high purity nitrogen gas at about 6 lb/in 2 for at least 5 minutes to reduce the interference of carbon dioxide absorption in the background scan to ≦ 3%. Infrared absorption Spectrum (KBr, cm) of the form 1 crystal-1) Characteristic bands are found at about 3325-. A representative infrared absorption spectrum of form 1 is shown in fig. 3.
The form 1 crystals are typically an opaque white or off-white powder when dried. The crystals obtained from a given formulation are generally polydisperse and have a crystal habit including platelets, needles, platelets and aggregates of platelets, needles and platelets. The crystals of form 1 are generally about 1 to 300 μm long, irregularly shaped, and have broken or angular edges. The form 1 crystals obtained from the preparation at low temperature (usually about 2-4 ℃) using acetone and di-n-butyl ether as crystallization solvents are usually aggregates containing a large proportion of needle crystals and some laths. FIGS. 4 to 7 are photographs of crystals of form 1 obtained by crystallization in acetone and di-n-butyl ether at a temperature of 15 ℃ or higher. These photographs show that the length of the platelets or plate-shaped and needle-shaped crystals is between about 10-250 um. FIG. 9 shows the crystals of form 1 obtained by crystallization from acetone and di-n-butyl ether at about 2 to 4 ℃. The photographs show that the diameter of the plate-shaped and needle-shaped crystal aggregates is between about 30-120 μm. Each crystal in the aggregate has a corner edge.
The water content of the form 1 crystals was found to be less than 1% by Karl-Fischer titration. The present inventors have conducted water content analysis essentially as described in the prior art (see, for example, the United states Pharmacopeia, pages 1619 to 1621, 1990 by Rockville, Md., USA).
Example 8: preparation of form 2 crystals
The form 1 crystals were converted to form 2 dihydrate by incubation at room temperature in an atmosphere with a relative humidity of 94% for 3 days. In the process of transformation from form 1 to form 2, a mixture of form 1 and form 2 crystals is obtained. While there was no detectable form 2 in the initial form 1 formulation. The crystallization of form 2 increased with time. At the end of the 3-day incubation, the final form 2 preparation contained no detectable crystals of form 1.
Example 9: physical characterization of the form 2 crystals
Form 2 crystals were analyzed by XRD in the same manner as form 1. Form 2 AD crystals have characteristic XRD peaks (expressed in degrees 2 theta) at about 8.7-8.9, 9.6, 16.3, 18.3, 18.9, 19.7, 21.0, 21.4, 22.0, 24.3, 27.9, 30.8 and 32.8. A representative XRD pattern of form 2 is shown in fig. 11.
The crystallization of form 2 was also analyzed by differential scanning calorimetry in the same manner as form 1, and the resulting thermogram is shown in FIG. 12, which has a characteristic endothermic transition at about 72.2 ℃ and an onset temperature of about 69.5 ℃.
The melting point of the form 2 crystals was determined by conventional melting point analysis. The analysis was carried out in the same manner as in form 1. The form 2 crystals melt between 70.9 and 71.8 ℃.
An IR spectrum of form 2 was obtained in the same manner as in form 1 (see fig. 13). The IR spectrum (KBr, cm)-1) Characteristic bands are found at about 3300-. These bands are similar to those of the crystal of form 1, but form 2 has another stretched band of water-related O-H bonds at about 3500.
The water content of the form 2 crystals was found to be 6.7% by Karl-Fischer titration. The present inventors have conducted water content analysis essentially as described in the prior art (see, for example, the United states Pharmacopeia, pages 1619 to 1621, 1990 by Rockville, Md., USA).
Example 10: preparation of form 3 crystals
A sufficient amount of form 1 crystals (about 250mg) was dissolved in anhydrous methanol (about 2mL) at room temperature to give a solution. The solution is obtained by mixing for about 10-15 minutes until the crystals are dissolved. The solution was allowed to stand without stirring for about 10 to 48 hours, and then form 3 crystals were recovered from the solution.
Example 11: physical characterization of the form 3 crystals
Form 3 crystals were analyzed by XRD in the same manner as form 1. Form 3 AD crystals were characterized as having XRD peaks expressed in degrees 2 theta substantially at about 8.1, 8.7, 14.1, 16.5, 17.0, 19.4, 21.1, 22.6, 23.4, 24.2, 25.4 and 30.9. A representative XRD pattern of form 3 is shown in fig. 14.
Example 12: synthesis and purification of PMEA
PMEA used for AD synthesis and crystallization was purified to improve product yield and purity. To a 12L three neck round bottom flask containing 548.8g of diethyl PMEA was added 637.5mL of acetonitrile at room temperature. The diethyl PMEA was dissolved by moderate stirring (with moderate vortex but no or substantially no splashing of the flask contents). The flask was purged with nitrogen and 803.8g of trimethylbromosilane was added slowly (about 2-5 minutes). The flask contents were heated at reflux temperature (65 ℃) for 2 hours until residual monoethyl PMEA ≦ 1% as determined by HPLC according to area normalization. Volatile substances are distilled off at the temperature of less than or equal to 80 ℃ and the mmHg of-20 mmHg. Then 1500mL of water (room temperature) was added to the flask. Next, the pH of the solution was adjusted to 3.2 with 25% w/v sodium hydroxide. The flask contents were then heated at 75 ℃ for 2 hours, then the contents were cooled to 3-4 ℃ over 15-20 minutes and held at 3-4 ℃ for 3-3.5 hours. The flask contents were then filtered through a sintered glass filter and the filter cake was washed with 150mL of cold (3-4 ℃ C.) water. The washed filter cake was transferred to a clean 12L three-neck flask, 2025mL of water was added to the flask and the flask was heated to 75 ℃ and held at that temperature for 2 hours. The heating was stopped and the flask was cooled and held at 3-4 ℃ for 3-3.5 hours. The flask contents were then filtered with a sintered glass plate filter and the filter cake was washed with 150mL of cold (3-4 deg.C) water, followed by 1050mL of acetone (room temperature). The filter cake was heated at 65-70 ℃ under-20 mmHg to make the weight constant. The PMEA yield was 85.4% and the purity was 99% as determined by area normalization or external standard HPLC analysis.
Example 13: single crystal X-ray crystallographic analysis of form 1
Approximately 200mg of the AD drug from lot 840-D-1 was dissolved in 200mg of acetone. The solution was heated to about 60 ℃. Di-n-butyl ether was added slowly to the solution at 60 deg.C (ambient temperature) until the initial trace of precipitation appeared. The mixture was then shaken and reheated to about 60 ℃ to form a clear and homogeneous solution. The solution was allowed to stand overnight, allowed to cool to ambient temperature, and held at ambient temperature for about 2 days. The crystals obtained are highly polydisperse, some up to 1mm in length. The supernatant was decanted and the residual crystals were washed 4 times in a cycle with a total of about 1mL of di-n-butyl ether to remove the residual supernatant. Crystals having a size of about 150X 200X 320 μm were analyzed by single crystal X-ray diffraction.
All measurements were carried out with graphite-loaded monochromatic Mo-Ka radiation (lambda. 0.71069)) By the Siemens SMART diffractometer of (1). With Paratone NTMHydrocarbon oil fixes crystals to glass fibersAnd (4) dimension. Data were collected at-135. + -. 1 ℃. Pictures of any hemisphere of inverted lattice space were collected with a w-scan (0.3 °/frame, 10 seconds per frame).
5967 integrated reflections (measuring maximum 2 theta at 51.6 deg.) were averaged to give 3205 Friedel unique reflections (Rint0.044). The structure is resolved with anisotropically refined non-hydrogen atoms. Hydrogen atoms are introduced at ideal positions. Full matrix least squares refinement according to 2438 observed reflections (with I > 3 σ and 306 variable parameters) converges at R-0.048 (Rw-0.054).
The cell constants and orientation matrices obtained by the least squares refinement using the measured positions of 3242 reflections with I > 10 σ in the range of 3.00 < 2 θ < 45.00 ° corresponding to the C-center monoclinic cell are specifically as follows: a is 12.85，b＝24.50，c＝8.28β is 100.2 °, Z is 4, and space group Cc.
The following table shows the data obtained from the study. Images of AD are shown in FIGS. 27 and 28.
Partial atomic coordinates of AD of form 1a
Atom(s) x y z
P1 1.0808 0.22760(05) 0.6554
O1 0.8826(03) 0.23934(12) 0.6880(04)
O2 1.1005(04) 0.26242(16) 0.5213(05)
O3 1.0440(03) 0.16716(14) 0.6037(05)
O4 1.0034(04) 0.12075(16) 0.3651(05)
O5 0.9271(05) 0.16940(19) 0.1501(06)
O6 1.1768(03) 0.21530(12) 0.7951(04)
O7 1.3179(03) 0.17817(13) 0.6942(04)
O8 1.3518(04) 0.13595(19) 0.9392(06)
N1 0.6976(04) 0.09182(15) 0.7806(05)
N2 0.6997(04) 0.06321(14) 0.3428(05)
N3 0.6929(04) 0.15993(15) 0.3987(05)
N4 0.6929(04) 0.17777(13) 0.6860(05)
N5 0.7041(04) -0.00364(15) 0.5388(05)
C1 0.6935(05) 0.14417(19) 0.8165(06)
C2 0.7000(04) 0.09175(17) 0.6147(06)
C3 0.7008(04) 0.04924(19) 0.4999(06)
C4 0.6945(05) 0.11621(19) 0.3029(06)
C5 0.6962(04) 0.14452(17) 0.5538(05)
C6 0.6968(05) 0.23782(18) 0.6890(06)
C7 0.8026(04) 0.25795(18) 0.7733(06)
C8 0.9855(05) 0.25344(20) 0.7701(07)
C9 0.9597(06) 0.1557(03) 0.4715(08)
aThe numbers in parentheses indicate the standard deviation of the last significant digit
Partial atomic coordinates of AD of form 1a(continuation)
Atom(s) x y z
C10 0.9798(05) 0.1318(02) 0.2018(07)
C11 1.0283(04) 0.08975(19) 0.1036(06)
C12 1.1460(06) 0.1018(03) 0.1244(10)
C13 1.0105(06) 0.0329(02) 0.1618(08)
C14 0.9783(07) 0.0959(03) -0.0773(08)
C15 1.2825(05) 0.22414(20) 0.7731(06)
C16 1.3473(05) 0.1340(02) 0.7942(09)
C17 1.3650(05) 0.0841(02) 0.6937(09)
C18 1.4337(07) 0.0440(03) 0.8045(12)
C19 1.4160(05) 0.1000(02) 0.5486(09)
C20 1.2561(06) 0.0599(03) 0.6340(11)
H1 0.6911 0.1572 0.9239
H2 0.6915 0.1239 0.1897
H3 0.7060 -0.0145 0.6494
H4 0.7044 -0.0304 0.4560
H5 0.6836 0.2511 0.5796
H6 0.6439 0.2511 0.7458
H7 0.8166 0.2445 0.8826
H8 0.8025 0.2967 0.7751
aThe numbers in parentheses indicate the standard deviation of the last significant digit
Partial atomic coordinates of AD of form 1a(continuation)
Atom(s) x y z
H9 0.9977 0.2379 0.8768
H10 0.9916 0.2920 0.7786
H11 0.9032 0.1380 0.5107
H12 0.9346 0.1884 0.4165
H13 1.1770 0.0992 0.2371
H14 1.1785 0.0762 0.0630
H15 1.1561 0.1377 0.0861
H16 0.9367 0.0263 0.1513
H17 1.0404 0.0072 0.0974
H18 1.0430 0.0293 0.2736
H19 0.9919 0.1315 -0.1138
H20 1.0079 0.0696 -0.1405
H21 0.9041 0.0903 -0.0902
H22 1.2855 0.2557 0.7074
H23 13266 0.2293 0.8768
H24 13999 0.0345 0.8938
H25 1.4441 0.0122 0.7442
H26 1.5002 0.0604 0.8454
H27 1.4811 0.1181 0.5869
H2.8 1.4288 0.0681 0.4897
H29 1.3701 0.1237 0.4784
H30 1.2125 0.0863 0.5708
H31 1.2623 0.0287 0.5684
H32 12254 0.0497 0.7257
aThe numbers in parentheses represent the standard deviation of the last significant digitDifference (D)
Fig. 29 is a powder X-ray diffraction pattern of form 1 AD: (a) observed values, (b) calculated values.
Example 14: preparation of form 4 crystals
Form 1 AD (10.05g) was dissolved in warm (about 35 ℃ C.) isopropanol (50mL) and filtered through a fritted glass funnel (M fritted glass, ASTM 10-15 μ M). The filtrate was added to a stirred isopropanol solution (35 ℃) containing dissolved fumaric acid (2.33g), and the resulting mixture was allowed to cool naturally to room temperature. Immediately after the AD solution is added to the isopropanol solution, form 4 crystals (i.e., AD fumaric acid (1: 1)) form spontaneously in the mixture. The crystals were allowed to form at room temperature for 2 days, recovered by filtration, and dried under vacuum at room temperature under a nitrogen atmosphere.
Example 15: preparation of form 4 crystals
Form 1 AD (1005.1g) was dissolved in warm (about 45 ℃ C.) isopropanol (3.0L). The warmed AD solution was added to a stirring solution of isopropanol (6.0L) (about 45 ℃) in a 12L flask containing dissolved fumaric acid (233.0g) over about 20 minutes with moderate stirring. The mixture was held at 40-45 ℃ for 10 minutes and the warming was stopped when a thick precipitate formed. Several minutes after the AD solution was added in its entirety, the mixture became cloudy, and after several minutes, the precipitate became thick, at which point stirring was stopped (the mixture was allowed to warm to 42 ℃). The precipitate was allowed to form for 1 hour. Slow stirring was started and continued for about 2 hours, then the 12L flask was immersed in water at room temperature and stirred slowly overnight to facilitate cooling of the mixture. By first filtration (Tyvek)TMFilter) and a second filtration (M sintered glass plate funnel) the precipitate was recovered and dried under vacuum at room temperature under nitrogen.
Example 16: preparation of crystalline AD from organic and inorganic acids
Form 1 AD (500mg, 1.0mmol) was dissolved in warm (< 40 ℃ C.) isopropanol (5 mL). Acid (1.0mmol) dissolved in 2mL (or a larger volume required to dissolve the acid) of isopropanol was added to the AD solution. The solution was stored at room temperature in a covered scintillation vial. In some cases, precipitated salt was observed shortly after capping the solution (about 1 minute). For other salts, a precipitate started to form up until several months after the solution was capped. The melting points of all 13 salts are shown in the table below. The XRD data (degrees 2 theta) for the 9 salts are also shown in the table below. The XRD data shows a large portion of the most intense peak of these salts.
*Doublet or acromion
**3-4 peaks in the broad peak
Example 17: AD formulations
Form 1 AD was formulated as tablets containing 30, 60 or 120mg AD per tablet with several excipients as described below.
1 part 2 (intragranular and extragranular) is incorporated into the dosage form during manufacture.
2 is sufficient to make a suitable wet granulate. The water is removed to a level where the Loss On Drying (LOD) does not exceed 3%.
The process for producing the tablet containing form 1 AD was to blend croscarmellose sodium, pregelatinized starch, and lactose monohydrate in a granulator. Water is added and the contents are mixed in a granulator until a suitable wet granulate is formed. The wet granulation is ground and dried in a dryer to a moisture content of no more than 3% loss on drying and the dry granulation is passed through a mill. The milled granules were mixed with extragranular excipients, lactose monohydrate, croscarmellose sodium and talc and blended in a blender to obtain a powder blend. Magnesium stearate was added, blended in a blender, and compressed into tablets. The tablets are filled into high density polyethylene or glass bottles (optionally with a silica gel desiccant) together with polyester fiber packaging.
Example 18: AD formulations
AD was formulated with several excipients into tablets each weighing 100mg and containing 25mg or 50mg of AD as follows. Tablets were made by wet granulation in a manner similar to that described above.

Claims (12)

1. Methanol solvated form C20H32N5O8P·CH3Crystalline adefovir dipivoxil in OH, further characterized by having peaks at 8.1 ± 0.1, 19.4 ± 0.1, 25.4 ± 0.1, and 30.9 ± 0.1 in an X-ray powder diffraction spectrum expressed in degrees 2 θ using Cu-K α radiation; and the DSC endothermic transition is 85 ℃ +/-2.
2. Crystalline adefovir dipivoxil as claimed in claim 1, further characterized by 1 equivalent of methanol in the crystal lattice.
3. Crystalline adefovir dipivoxil as claimed in claim 1, having a median size, as measured by light scattering, in the range of 20 to 150 μm.
4. Crystalline adefovir dipivoxil as claimed in claim 3, having a median size, as measured by light scattering, of from 30 to 120 μm.
5. Crystalline adefovir dipivoxil as claimed in claim 1, wherein the crystals have a length of 1 to 300 μm in the preparation thereof.
6. Crystalline adefovir dipivoxil as claimed in claim 1, having peaks at 8.1 ± 0.1, 8.7 ± 0.1, 14.1 ± 0.1, 16.5 ± 0.1, 17.0 ± 0.1, 19.4 ± 0.1, 21.1 ± 0.1, 22.6 ± 0.1, 23.4 ± 0.1, 24.2 ± 0.1, 25.4 ± 0.1 and 30.9 ± 0.1 in X-ray powder diffraction spectrum expressed in degrees 2 Θ using Cu-ka radiation.
7. Adefovir dipivoxil, wherein less than 20% of the adefovir dipivoxil is amorphous adefovir dipivoxil and the remainder is crystalline adefovir dipivoxil as claimed in any one of claims 1 to 6.
8. Adefovir dipivoxil, wherein less than 10% of the adefovir dipivoxil is amorphous adefovir dipivoxil and the remainder is crystalline adefovir dipivoxil as claimed in any one of claims 1 to 6.
9. Adefovir dipivoxil, wherein less than 1% of the adefovir dipivoxil is amorphous adefovir dipivoxil and the remainder is crystalline adefovir dipivoxil as claimed in any one of claims 1 to 6.
10. Adefovir dipivoxil, wherein less than 0.1% of the adefovir dipivoxil is amorphous adefovir dipivoxil and the remainder is crystalline adefovir dipivoxil as claimed in any one of claims 1 to 6.
11. A pharmaceutical composition comprising crystalline adefovir dipivoxil as claimed in any one of claims 1 to 6 or adefovir dipivoxil as claimed in any one of claims 7 to 10 and a pharmaceutically acceptable excipient.
12. Use of crystalline adefovir dipivoxil as claimed in any one of claims 1 to 6 or adefovir dipivoxil as claimed in any one of claims 7 to 10 for the preparation of an antiviral pharmaceutical composition.
HK06111839.6A 1997-07-25 2006-10-26 Nucleotide analog compositions HK1091211B (en)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US5377197P 1997-07-25 1997-07-25
US90074597A 1997-07-25 1997-07-25
US08/900,745 1997-07-25
US60/053,771 1997-07-25

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HK1091211A1 HK1091211A1 (en) 2007-01-12
HK1091211B true HK1091211B (en) 2013-10-11

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