HK1147496A - Co-crystals and pharmaceutical compositions comprising the same - Google Patents

Description

Co-crystals and pharmaceutical compositions comprising the same

Cross-referencing

Priority of U.S. application No.60/969,023, filed on 30/8/2007, the contents of which are incorporated herein by reference in their entirety.

Background

Hepatitis c virus ("HCV") is an attractive human medical problem. HCV is considered to be the causative agent of most non-A, non-B Hepatitis, with an estimated global human prevalence of 3% [ A.Alberti et al, "Natural History of Hepatitis C," J.hepatology, 31(suppl.1), pp.17-24 (1999) ]. Approximately four million individuals are infected in The United states alone [ M.J. Alter et al, "The epidemic of Viral Hepatitis in The United states," gastroenterol. Clin.North Am., 23, 437-455 (1994); M.J. Alter "hepatiss C Virus Infection in the United States," J.Heatology, 31(suppl.1), pp.88-91 (1999) ].

Only about 20% of infected individuals develop acute clinical hepatitis after the first exposure to HCV, while others appear to spontaneously eliminate the infection. However, in almost 70% of cases, The virus establishes a chronic infection that lasts for decades [ S.Iward, "The Natural Course of viral Hepatitis," FEMS Microbiology Reviews, 14, p.201-204 (1994); lavanchy, "Global scientific and Control of heparin C," J.viral Hepatitis, 6, pp.35-47 (1999) ]. This often leads to recurrent and progressive worsening liver inflammation, often leading to more severe disease states such as cirrhosis and Hepatocellular Carcinoma [ m.c. kew, "hepatotis C and Hepatocellular cartioma", femsiobiology Reviews, 14, page 211-220 (1994); saito et al, "heparin CVirus Infection Associated with the Development of heparin Carcinoma," Proc. Natl. Acad. Sci. USA, 87, p 6547-. Unfortunately, there is no widely effective treatment for impairing the progression of chronic HCV.

The HCV genome encodes a polyprotein of 3010-3033 amino acids [ Q.L.Choo, et al, "Genetic Organization and division of the Hepatitis C Virus," Proc.Natl.Acad.Sci.USA, 88, pp.2451-2455 (1991); kato et al, "molecular cloning of the Human Hepatitis C Virus Genome From Japan Patientswitch Non-A, Non-B Hepatitis," Proc.Natl.Acad.Sci.USA, 87, pp.9524-9528 (1990); takamizawa et al, "Structure and Organization of the Hepatitis C Virus Genome Isolated From Human carriers," J.Virol., 65, p.1105-. The HCV Nonstructural (NS) proteins are presumed to provide the necessary catalytic mechanisms for viral replication. NS proteins are derived from the proteolytic Cleavage of polyproteins [ R.Bartenschlager et al, "No structural Protein 3 of the Hepatitis C Virus Encodes a series-Type Protein requiring for Cleavage at the NS3/4and NS4/5Junctions," J.Virol., 67, page 3835-cake 3844 (1993); grakoui et al, "Characterization of the Hepatitis C Virus-Encoded spring protease: determination of protein-Dependent protein clearance Sites, "J.Virol., 67, pages 2832-2843 (1993); grakoui et al, "Expression and differentiation of Hepatitis C Virus Polyprotein Cleavage Products," J.Virol., 67, pages 1385-1395 (1993); tomei et al, "NS 3 is a serine protease requiring for processing of hepatis C virus polyprotein", J.Virol., 67, pp.4017-4026 (1993) ].

HCV NS protein 3(NS3) is essential for viral replication and infectivity [ Kolykhalov, J.virology, Volume 74, pp.2046-2051 "Mutations at the HCV NS3 series Protease Catalytic activity of HCV RNA in Chimpanzes ]. Mutations in the NS3 protease of Yellow Fever Virus are known to reduce the infectivity of the Virus [ Chambers, T.J., et al, "where is the Protein N-tertiary Domain of nonstructural Protein NS3 From Yellow wine river Virus is a series Protein disposal for Site-Specific cleavage in the Viral Polyprotein", Proc. Natl.Acad.Sci.USA, 87, page 8898. 8902 (1990) ]. The first 181 amino acids of NS3 (1027-1207 residues of the viral polyprotein) have been shown to contain the Serine protease domain of NS3 that processes all 4 downstream sites of the HCV polyprotein [ C.Lin et al, "Hepatitis C Virus NS3 spring protease: trans-clean Requirementsand Processing tools ", J.Virol, 68, pp 8147-8157 (1994) ].

The HCV NS3serine protease and its associated cofactor NS4A contribute to the processing of all viral enzymes and are therefore considered essential for viral replication. This processing appears to be similar to that performed by the human immunodeficiency virus aspartyl protease, which is also involved in the processing of viral enzymes. HIV protease inhibitors that inhibit viral protein processing are potent human anti-viral agents, suggesting that interference with this phase of the viral life cycle may result in a therapeutically active agent. Therefore, hcv ns3serine protease is also an attractive target for drug development.

Until recently, the only established treatment for HCV disease was interferon therapy. However, interferons have significant side effects [ m.a. wlaker et al, "Hepatitis C viruses: anoverview of Current applications and Progress, "DDT, 4, pages 518-29 (1999); mordapour et al, "Current and Evalling therapeutics for Heapatitis C," Eur.J.Gastroenterol.Heapatol., 11, pp 1199-1202 (1999); H.L.A.Janssen et al, "Suicide Associated with Alfa-interference Therapy for Chonic viral Hepatitis," J.Heapatol., 21, pp.241-243 (1994); renault et al, "Side Effects of Alpha interference," sensines in Liver Disease, 9, pp.273-]And induce long-term remission in only a fraction (-25%) of cases O.Weiland, "interference Therapy in viral Hepatitis C Virus infection", FEMS Microbiol.Rev., 14, 279-288 (1994)]. Interferon in the form of PEG: (And) And ribavirin and interferonOnly recent introduction of combination therapyResulting in moderate improvement in remission rates and partial reduction in side effects. In addition, the prospects for effective anti-HCV vaccines remain uncertain.

Thus, there is a need for more effective anti-HCV treatments. Such inhibitors would have therapeutic potential as protease inhibitors, particularly as serine protease inhibitors, and more particularly as HCV NS3 protease inhibitors. In particular, such compounds are useful as antiviral agents, particularly as anti-HCV agents.

The HCV inhibitor VX-950, which has the structure shown below, is a desirable class of compounds. VX-950 is described in PCT publication No. WO02/18369, which is incorporated herein by reference in its entirety.

Summary of The Invention

The present invention relates generally to co-crystals (co-crystals) comprising the HCV inhibitor VX-950 and a co-crystal former, and compositions comprising the VX-950 co-crystal. The co-crystal former may be a pharmacologically inert excipient that alters the crystalline form of the solid drug by forming co-crystals, clathrates, or other crystalline solid forms. It is within the meaning of "co-form" (co-form) as used herein. In some cases, VX-950 and the co-crystal former together can form a multi-component single-phase crystalline solid or composition, i.e., a co-crystal. Specific VX-950 co-crystals are advantageous compared to their free form because they may have improved solubility, higher water solubility, or higher solid state physical stability than amorphous VX-950 dispersions. The particular VX-950 co-crystal provides reduced dosage form quality and thus reduced dosing burden, as VX-950 co-crystal also exhibits higher bulk density relative to the amorphous form. In addition, VX-950 co-crystals provide manufacturing advantages over amorphous forms that require spray drying, melt extrusion, lyophilization, or precipitation.

In one aspect, the invention provides VX-950 co-crystals comprising VX-950 and 2, 4-dihydroxybenzoic acid as a co-crystal former compound. In certain embodiments, the molar ratio of VX-950 and 2, 4-dihydroxybenzoic acid in the co-crystal ranges from about 5: 1 to about 1: 5 (e.g., about 1: 1). In certain embodiments, the co-crystal has at least two of the four X-ray powder diffraction peaks at about 7.6, 8.5, 9.6, and 11.9 degrees 2-theta, each with a standard deviation of about +/-0.3 deg. 2-theta. In certain embodiments, the co-crystal has a peak in its DSC thermogram at about 150.81 ℃ with a standard deviation of about +/-5 ℃.

In another aspect, the invention provides VX-950 co-crystals comprising VX-950 and 2, 5-dihydroxybenzoic acid as a co-crystal former compound. In certain embodiments, the molar ratio of VX-950 and 2, 5-dihydroxybenzoic acid in the co-crystal ranges from about 5: 1 to about 1: 5 (e.g., about 1: 1). In certain embodiments, the co-crystals each have at least two of the four X-ray powder diffraction peaks at about 8.3, 9.6, 11.7, and 17.1 degrees 2-theta, each with a standard deviation of about +/-0.3 degrees 2-theta (i.e., ° 2-theta). In certain other embodiments, the co-crystal has a peak in its DSC thermogram at about 163.48 ℃ with a standard deviation of about +/-5 ℃.

In another aspect, the invention provides a co-crystal comprising VX-950 and 3-methoxy-4-hydroxybenzoic acid as a co-crystal former compound. In certain embodiments, the molar ratio of VX-950 and 3-methoxy-4-hydroxybenzoic acid in the co-crystal ranges from about 5: 1 to about 1: 5 (e.g., about 1: 1). In certain embodiments, the co-crystals each have at least two of the four X-ray powder diffraction peaks at about 7.8, 8.6, 11.4, and 11.9 degrees 2-theta, each with a standard deviation of about +/-0.3 degrees 2-theta. In certain embodiments, the co-crystals each have a peak in their DSC thermogram at about 178.09 ℃ with a standard deviation of about +/-5 ℃.

In another aspect, the invention provides compositions each comprising VX-950, a co-crystal former selected from the group consisting of 2, 4-dihydroxybenzoic acid, 2, 5-dihydroxybenzoic acid, and 3-methoxy-4-hydroxybenzoic acid, and a solvent; the solvent is selected from acetonitrile, dichloromethane, ethyl acetate, ethanol, acetone or a mixture thereof. In certain embodiments, VX-950, the co-crystal former, and the solvent together can form a crystalline form (i.e., form a co-crystal). The co-crystal may be a solvate due to the presence of the solvent. In certain other embodiments, the solvent is a mixture of acetonitrile and dichloromethane (e.g., about a 1: 1 volume ratio). In certain embodiments, the molar ratio of VX-950 to 4-aminosalicylic acid ranges from about 5: 1 to about 1: 5 (e.g., about 1: 1). In certain embodiments, the molar ratio of VX-950 to solvent ranges from about 1: 0.05 to about 1: 1 (e.g., about 1: 0.34 or about 1: 0.5).

These compositions may be used to treat diseases involving or associated with HCV, and the like. Accordingly, pharmaceutical compositions each comprising VX-950 and the above-identified co-crystal former in a suitable molar ratio are also within the scope of the present invention. The pharmaceutical composition may optionally comprise a solvent (e.g., acetonitrile, dichloromethane, ethyl acetate, ethanol, or acetone) to form a solvate. In addition, the pharmaceutical composition may further comprise a pharmaceutically acceptable diluent, solvent, excipient, carrier or solubilizer.

Methods of preparing the co-crystals described above are also within the scope of the invention. The method may comprise the steps of: (a) providing VX-950, (b) providing a co-crystal former selected from the group consisting of 2, 4-dihydroxybenzoic acid, 2, 5-dihydroxybenzoic acid, and 3-methoxy-4-hydroxybenzoic acid (optionally in a solvent, e.g., ethanol, acetonitrile, dichloromethane, or mixtures thereof, so as to form a solvate), (c) milling, heating, co-subliming, co-melting, or contacting in solution VX-950 and the co-crystal former under crystallization conditions to form a solid phase co-crystal, and (d) optionally isolating the co-crystal formed in step (c).

Methods of adjusting the chemical or physical properties of interest of the co-crystals described above are also within the scope of the invention. The method may comprise the steps of: (a) measuring a chemical or physical property of interest of VX-950 and a co-crystal former selected from the group consisting of 2, 4-dihydroxybenzoic acid, 2, 5-dihydroxybenzoic acid, and 3-methoxy-4-hydroxybenzoic acid, (b) determining the mole fraction of VX-950 and the co-crystal former that would result in the desired modulation of the chemical or physical property of interest, and (c) preparing the co-crystal using the mole fraction determined in step (b).

The compositions and co-crystals of the invention may be used to treat diseases related to or associated with HCV. Thus, a method of treating such diseases is also within the scope of the invention, comprising administering to a subject in need thereof a therapeutically effective amount of a co-crystal of the invention or a composition of the invention.

The compositions and co-crystals of the present invention may also be used as seeds to prepare other co-crystals comprising an active ingredient, which may be the same as or different from VX-950, and a co-crystal former, which may be the same as or different from the group consisting of 2, 4-dihydroxybenzoic acid, 2, 5-dihydroxybenzoic acid, and 3-methoxy-4-hydroxybenzoic acid. For example, a small amount of the co-crystal of the present invention may be placed in a solution containing the desired active ingredient and the co-crystal former, and the mixture allowed to stand to form additional co-crystals with and grow from the existing co-crystal.

Furthermore, the compositions and co-crystals of the present invention may be used as research tools. For example, they can be used to study the pharmacological properties (e.g., bioavailability, metabolism, and efficacy) of VX-950 in different forms and conditions, or to develop different VX-950 formulations for best delivery and absorption.

Brief Description of Drawings

FIG. 1 shows the XRPD of a co-crystal of VX-950 and 2, 5-dihydroxybenzoic acid.

FIG. 2 shows a TGA spectrum of a co-crystal of VX-950 and 2, 5-dihydroxybenzoic acid.

FIG. 3 shows a DSC spectrum of co-crystal of VX-950 and 2, 5-dihydroxybenzoic acid.

FIG. 4 shows the XRPD of a co-crystal of VX-950 and 3-methoxy-4-hydroxybenzoic acid.

FIG. 5 shows a TGA spectrum of a co-crystal of VX-950 and 3-methoxy-4-hydroxybenzoic acid.

FIG. 6 shows a DSC spectrum of co-crystal of VX-950 and 3-methoxy-4-hydroxybenzoic acid.

FIG. 7 shows the XRPD of a co-crystal of VX-950 and 2, 4-dihydroxybenzoic acid.

FIG. 8 shows a TGA spectrum of a co-crystal of VX-950 and 2, 4-dihydroxybenzoic acid.

FIG. 9 shows a DSC spectrum of co-crystal of VX-950 and 2, 4-dihydroxybenzoic acid.

Detailed Description

Methods for preparing and characterizing co-crystals are well documented in the literature. See, e.g., Trask et al, chem. Commun., 2004, 890-; and o.almarsson and m.j.zawortko, chem.commun., 2004, 1889-. In general these methods are also suitable for preparing and characterizing the co-crystals of the invention.

Furthermore, the following specific methods can be used to identify co-crystal formers suitable for the preparation of co-crystals, in particular co-crystals of the invention.

The initial identification or screening of possible VX-950 co-crystal formers can be performed in a 96-well plate on a milligram scale. Visual comparison of XRPD results to known crystalline VX-950 diffraction patterns can be used as a screen for new crystalline forms and/or altered lattice sizes that can indicate incorporation of co-crystal formers into the crystals. Oxalic acid, 4-aminosalicylic acid, salicylic acid, and the like have been identified by this initial screen as potential candidates for co-crystal formation with VX-950.

The results of the initial screening can be used to model the identification of other VX-950 co-crystal formers. For example, 4-aminosalicylic acid can be used as a lead molecule for identifying other possible VX-950 co-crystal formers through molecular modeling due to better physical and chemical properties. Specifically, a model of 4-ASA can be built using the Quanta software package (Accelrys Inc., San Diego, Calif.) and complexed with the structure of a single molecule of VX-950 obtained by single crystal x-ray diffraction. The 4-ASA molecule can be manually placed at different positions around VX-950 to form the maximum number of hydrogen bonds between the two molecules. The site complex of the 4-ASA molecule is energy-minimizing, while the VX-950 molecule remains immobilized. The adapted-based Newton-Raphson method provided by Quanta can be used for energy minimization using default settings and distance dependent dielectrics. AutoNom software (MDL Information Systems, GmbH) can be used to convert the names of compounds in the FDA's EAFUS (everything added to food, US) and GRAS (generally regarded as safe) lists to 2D structures in SMILES format to generate a structure database. The database can then be searched for new co-crystal formers of pharmacophores suitable for identification using 4-ASA. The acceptable pharmacophore has a local energy minimum similar to that of VX-950 and 4-ASA.

DSC can also be used to screen co-crystal formers. In screening by DSC, a physical mixture of a co-crystal former showing evidence of solid phase interaction (i.e., formation of eutectic melt) with VX-950 is more likely to form a co-crystal during DSC. To detect the interaction between VX-950 and the co-crystal former, the components can be mixed in a 1: 1 molar ratio and subjected to DSC temperature ramping to raise the temperature from room temperature to, for example, 300 ℃ in 10 ℃ increments. A mixture is selected that exhibits a new thermal event (i.e., an endotherm) that differs in temperature from the endotherm of the pure component. In addition to the thermal conversion of one of the original components, when a new thermal conversion is observed, the molar ratio between VX-950 and the co-crystal former can then be adjusted so that only the new thermal conversion is produced. The observed transition temperature can be plotted as a function of composition to generate a phase diagram for the binary mixture. The composition that produces new thermally converted VX-950 and co-crystal former on DSC can then be scaled up to produce larger amounts (e.g., grams) as described above.

Mixtures of VX-950 and co-crystal formers with novel thermal conversions can be produced in large quantities (i.e., scaled up), for example by using ball milling, solvent-evaporation, melting with and without solvents, slurry conversion, mixing, sublimation, or modeling. Some such methods are described in detail below. The products so prepared can be analyzed or characterized by known methods such as XRPD, TGA and DSC, and their solubility and stability in aqueous media can also be measured by methods known in the art.

Ball mill: equimolar amounts of VX-950 and the co-crystal former are mixed with a suitable solvent. The mixture is then milled using a ball milling apparatus such as Retsch MM200(glen mills inc., Clifton, NJ) at a frequency of 15Hz for 3 hours. The mixture was then placed in a milling chamber made of sintered corundum. After milling, the material was placed in a screw-cap scintillation vial (uncapped) and dried under vacuum at room temperature. The resulting mixture was characterized by XRPD and DSC analysis.

Melting in a reaction block: equimolar amounts of VX-950 and co-crystal former were mixed, with or without solvent. The mixture is then placed in a reaction block, for example Model RR98072 from Radleys discovery technologies (Essex, UK), closed and heated to a temperature for a new thermal transition identified by DSC. The mixture was then held at the transition temperature for a period of time before opening the reaction block and the resulting mixture was cooled at ambient conditions.

Evaporation of the solvent: VX-950 and possibly the co-crystal former are dissolved in a volatile solvent or solvent mixture, respectively (e.g., 50: 50 toluene: acetonitrile). Dissolution can be assisted by stirring and ultrasound until a clear solution is obtained. The VX-950 solution was then mixed with the eutectic former solution in the desired molar ratio in a screw cap scintillation vial. Placing the uncapped vial under reduced pressureThe solvent is evaporated to dryness, which usually takes several days. A solid (crystalline) material was obtained and analyzed.

As described above, the co-crystals of the present invention can be analyzed by methods known in the art for characterizing solid or crystalline materials. Examples of characterization methods include thermogravimetric analysis (TGA), Differential Scanning Calorimetry (DSC), X-ray powder diffraction (XRPD), solubility analysis, dynamic vapor adsorption, infrared off-gas analysis, and suspension stability. TGA can be used to study the presence of residual solvent in the co-crystal samples, as well as to identify the temperature at which various co-crystal samples decompose. DSC can be used to find the thermal transitions that occur in the co-crystal samples as a function of temperature and to determine the melting point of various co-crystal samples. XRPD can be used for structural characterization of the co-crystal. Solubility analysis can be performed to reflect changes in the physical state of various eutectic samples. Suspension stability analysis can be used to determine the chemical stability of the co-crystal sample in a solvent. Some such methods are described in great detail.

X-ray powder diffraction (XRPD): XRPD can be used to characterize the physical form of a material by recording the changes in the original and monitored spectra of the material over time. XRPD patterns can be obtained in reflectance mode at room temperature, for example, by using a Bruker D8 Discover diffractometer (Bruker AXS, Madison, WI, USA) equipped with a sealed tube source and a Hi-Star area detector. A copper target X-ray tube (Siemens) may be operated, for example at 40kV and 35 mA. A graphite monochromator and 0.5mm collimator available from Bruker can be used to produce a parallel monochromatic beam (CuKa,). The distance between the sample and the detector is about 30 cm. The sample can be placed on a Si zero background wafer (e.g., from The Gem Dugout, State College, PA) and then placed in The center of The XYZ stage. Data, version 4.1.16(Bruker AXS, Madison, WI, USA) may be acquired using GADDS software for Windows NT. Two frames were recorded, each at 2 different 2 theta angles 8 deg. and 26 deg. 2-theta illumination times of 120 seconds. During the irradiation the sample isVibrating in the X and Y directions with an amplitude of 1 mm. The data were then integrated over a range of 3 ° to 41 ° 2-theta, step size 0.02 ° 2-theta, and combined to form a continuous map. The instrument was calibrated using a corundum plate (NIST standard 1976).

Differential Scanning Calorimetry (DSC): DSC can be used to detect the thermal transition that occurs in a sample as a function of temperature and to determine the melting point of a crystalline material. DSC analysis was performed, for example, using an indium-calibrated MDSC Q100 differential scanning calorimeter (TA Instruments, NewCastle, DE). Samples can be prepared in aluminum pans with a single pin hole binder, with sample amounts of, for example, about 2 mg. Each run was initially equilibrated to 25 ℃, and then ramped to 300 ℃ at a 10 ℃/minute ramp. VX-950 degraded under melting, with degradation starting at about 240 ℃. Data can be obtained through Thermal Advantage Q series^TMThe software was collected and analyzed by Universal analysis software (TA Instruments, New Castle, DE).

Thermogravimetric analysis (TGA): TGA can be used to study the presence of residual solvent in a sample and to identify the temperature at which decomposition of the sample occurs. For example, a Model Q500 thermogravimetric analyzer (TAInstructions, New Castle, DE) can be used for TGA measurements. The sample may weigh about 3-8mg and be heated at a rate of about 10 deg.c/minute to a final temperature, such as 300 deg.c. Data may be obtained, for example, by Thermal Advantage Q Series^TMThe software was collected and analyzed by Universal analysis software (TA Instruments, New Castle, DE).

Fourier transform infrared (FT-IR) spectroscopy: FT-IR can be used to study hydrogen bonding in mixtures of VX-950 and co-crystal former in varying molar ratios. For example, a Nexus 670 spectrometer (Thermo Electron Corp.; Madison, Wis.) can be used from 4000-625cm^-1Infrared transmission spectra were obtained from KBr particles.

Solubility determination: solubility can be expressed as VX-950 equivalents. Solubility can be measured to reflect changes in the physical state of the material, and progress toward the goal of increasing the solubility of VX-950 can be monitoredAnd (6) unfolding. Specifically, an aliquot of the material was placed in an aqueous medium with a target solubility of 10 mg/mL. At the stated time points, aliquots of the supernatant were removed, filtered through a 0.45 micron filter (e.g., Millex; Millipore, Billerica, Mass.) and analyzed using HPLC (e.g., Agilent 1100; Palo Alto, Calif.). The sample may be detected using a detector, for example set at 270nm and a flow rate of 1mL/minThe phenyl column 150mmx4.6mm, 3.5 μm particle size (P/N186001144) (Waters Corp., Milford, MA) was run isocratically. The mobile phase contained potassium phosphate buffer (10mM, pH 7.0) and methanol in a ratio of 60: 40 (v/v). The concentration of VX-950 can be determined by comparing the chromatographic peak area to a standard curve generated using standards of known concentration.

High temperature stage (hotstage) microscopy: microscope images can be acquired, for example, using an Olympus BX51 confocal microscope with polarized film, SLMPlan 50x infinity corrected objective, C-5050 digital camera, Instec high temperature stage with variable temperature controller. The experimental procedure consisted of a linear heating ramp between different temperature steps, where the samples were allowed to equilibrate for several minutes. Digital images were collected manually throughout the ramp to capture any transformations that occurred.

An effective amount of a co-crystal or composition of the invention, each comprising VX-950 and a co-crystal former (e.g., 4-hydroxybenzoic acid, 4-aminosalicylic acid (and acetonitrile), phenylalanine, threonine, adipic acid, butylene acetate, praline, methyl 4-hydroxybenzoate, anthranilic acid, d-biotin, or tartaric acid) can be used to treat a disease involving or associated with HCV. An effective amount is the amount necessary to confer a therapeutic effect on a subject being treated, e.g., a patient. An effective amount of VX-950 and the co-crystal of the co-crystal former is about 0.1mg/kg to about 150mg/kg (e.g., about 1mg/kg to about 60 mg/kg). One skilled in the art will recognize that effective dosages will also vary with the route of administration, the use of excipients, and the possibility of co-use with other therapeutic treatments, including the use of other therapeutic agents and/or treatments.

The co-crystal or pharmaceutical composition of the invention can be administered to a subject (e.g., a cell, tissue, or patient (including animal or human)) in need thereof, e.g., orally, intravenously, or parenterally, by any method that allows for the delivery of the compound VX-950. For example, they can be administered by pills, tablets, capsules, aerosols, suppositories, liquid preparations for ingestion or injection or as eye drops or ear drops, dietary supplements and topical preparations.

The pharmaceutical compositions may contain diluents, solvents, excipients and carriers such as water, ringer's solution, isotonic saline, 5% dextrose and isotonic sodium chloride solution. In another embodiment, the pharmaceutical composition may further comprise a solubilizing agent, such as a cyclodextrin. Other examples of suitable diluents, solvents, excipients, carriers and solubilizers can be found, for example, in the united states pharmacopeia 23/national formulary 18, Rockville, MD, u.s.pharmaceutical Convention, inc. (1995); ansel HC, Popovich NG, Allen Jr LV pharmaceutical DosageForms and Drug Delivery Systems, Baltimore MD, Williams & Wilkins, (1995); gennaro ar, Remingtons: the Science and Practice of Pharmacy, Easton PA, Mack Publishing Co., (1995); wade, a., Weller, p.j., Handbook of Pharmaceutical excipients, 2 nd edition, Washington DC, American Pharmaceutical Association (1994); baner GS, Rhodes CT.Modern pharmaceuticals, 3 rd edition, New York, Marcel Dekker, Inc. (1995); ranade VV, Hollinger MA, Drug Delivery Systems, Boca Raton, CRCPress, (1996).

The pharmaceutical compositions may also include an isotonic saline, 5% dextrose, or other well-known pharmaceutically acceptable excipient aqueous solution of the co-crystal. Solubilizers such as cyclodextrins, or other solubilizers well known to those skilled in the art, can be used as pharmaceutical excipients for the delivery of the therapeutic compound VX-950. With respect to the route of administration, the co-crystal or pharmaceutical composition may be administered orally, intranasally, transdermally, intradermally, vaginally, intraotically, intraocularly, buccally, rectally, transmucosally, or by inhalation or intravenous administration. The composition may be delivered intravenously via a balloon catheter. The compositions can be administered to an animal (e.g., a mammal such as a human, a non-human primate, a horse, a dog, a cow, a pig, a sheep, a goat, a cat, a mouse, a rat, a guinea pig, a rabbit, a hamster, a gerbil, a ferret, a lizard, a reptile, or a bird).

The co-crystals or pharmaceutical compositions of the invention may also be delivered by implantation (e.g., surgery) using an implantable device. Examples of implantable devices include, but are not limited to, stents, delivery pumps, vascular filters, and implantable controlled release compositions. Any implantable device can be used to deliver compound VX-950 as an active ingredient in a co-crystal or pharmaceutical composition of the invention, provided that 1) the device, compound VX-950, and any pharmaceutical composition containing the compound are biocompatible, and 2) the device can deliver or release an effective amount of the compound to impart a therapeutic effect to a patient being treated.

Delivery of therapeutic agents through stents, delivery pumps (e.g., micro-osmotic pumps), and other implantable devices is known in the art. See, e.g., "Recent Developments in coatedsteps", Hofma et al, published in Current international pathology Reports, 2001, 3: 28-36, the entire contents of which include the references cited therein. Additional descriptions of implantable devices such as stents may be found in U.S. patent nos. 6,569,195 and 6,322,847, and PCT international publication nos. WO 2004/044405, WO 2004/018228, WO 2003/229390, WO 2003/228346, WO 2003/225450, WO 2003/216699, WO 03/0204168, WO 2008/098255a2, WO 2008/027872a2, WO 2008/027871 a2, WO 2007/140320 a2, WO 2006/124823 a2, WO 2007/128969a2, WO 2007/030478 a2, and WO 2005/120393 a2, each of which (and other publications cited herein) is incorporated herein in its entirety.

Examples of the preparation and characterization of the co-crystals of the present invention are described below for illustration only and not to be construed as limiting in any way.

Example 1 preparation by ball milling

VX-950 and an equimolar amount of the co-crystal former (e.g., 2, 4-dihydroxybenzoic acid or 3-methoxy-4-hydroxybenzoic acid) are mixed with a solvent (e.g., butanone or ethyl acetate). The components can then be ground using a Wig-L-Bug instrument, such as Retsch MM200(GlenMills Inc, Clifton, NJ), at a frequency of 15Hz for 10 minutes. After milling, the starting material is dried, for example in a vacuum oven at 75 ℃ for 2 hours, to obtain the co-crystals of the invention.

Example 2 preparation by melt Process

VX-950 and an equimolar amount of co-crystal former (e.g., 2, 4-dihydroxybenzoic acid or 3-methoxy-4-hydroxybenzoic acid) are mixed, for example, by vortexing for 5 minutes, with or without a solvent. The mixture is then placed in a reaction block (e.g., RR98072 from Radley Discovery Technologies), the lid is closed and heated to an endothermic heat. The mixture is held at the endothermic temperature for 30 minutes, then the lid is opened and the resulting mixture is cooled at ambient conditions and the solvent (if used) is removed to give the co-crystals of the invention.

Example 3 preparation by solvent-Evaporation

2, 4-Dihydroxybenzoic acid: 200mg VX-950 and 80mg 2, 4-dihydroxybenzoic acid (Sigma Chemicals Co., St. Louis, Mo., USA) were charged into a 20mL glass vial. Then 100. mu.L of dichloromethane and 100. mu.L of acetonitrile were added to the vial. The vial containing the mixture was capped and the mixture was stirred at 600rpm using a magnetic stir bar at room temperature for 16 hours. The crystalline solid was isolated and the liquid on its surface was removed by filter paper to give a co-crystal of VX-950 and 2, 4-dihydroxybenzoic acid.

3-methoxy-4-hydroxybenzoic acid: 200mg VX-950 and 80mg 3-methoxy-4-hydroxybenzoic acid (Sigma Chemicals Co., St. Louis, Mo., USA) were charged into a 20mL glass vial. Then 100. mu.L of methylene chloride and 100. mu.L of ethyl acetate were added to the vialA nitrile. The vial containing the mixture was capped and the mixture was stirred at 600rpm using a magnetic stir bar for 72 hours at room temperature. The crystalline solid was isolated and the liquid on its surface was removed by filter paper to give a co-crystal of VX-950 and 3-methoxy-4-hydroxybenzoic acid.

2, 5-Dihydroxybenzoic acid: 200mg VX-950 and 80mg 2, 5-dihydroxybenzoic acid (Sigma Chemicals Co., St. Louis, Mo., USA) were charged into a 20mL glass vial. Then 100. mu.L of dichloromethane and 100. mu.L of acetonitrile were added to the vial. The vial containing the mixture was capped and the mixture was stirred at 600rpm using a magnetic stir bar at room temperature for 16 hours. The crystalline solid was isolated and the liquid on its surface was removed by filter paper to give a co-crystal of VX-950 and 2, 5-dihydroxybenzoic acid.

Example 4: diffraction of single crystals

Single crystal diffraction of the eutectic was performed on a Bruker APEX II CCD diffractometer using CuK α radiation at 100K by using a single crystal picked from the mother liquor and fixed on glass fibers. The crystals were cooled to 100K in a nitrogen flow system inA vibration picture is taken around the angular ω -axis. APEX software was used to index, integrate and measure (scale) data. The structure is parsed and refined using the SHELX-TL software package.

EXAMPLE 5 thermogravimetric analysis (TGA)

TGA analysis of each sample was performed using a Model Q500 thermogravimetric analyzer (TA Instruments, New Castle, DE, USA) which was used to control ThermalAdvantage Q Series^TMSoftware, version 2.2.0.248, Thermal advance release4.2.1(TA Instruments-Water LLC), has the following components: qadv.exe version 2.2build 248.0; RhDII.dII version 2.2build 248.0; rhbase. dii version 2.2build 248.0; rhcomm.dii version 2.2build 248.0; taliense.dii version 2.2build 248.0; and tga.dii version 2.2build 248.0. In addition, use is made ofThe Analysis software of (2) is Universal Analysis2000 software for Windows 2000/XP, version 4.1D build 4.1.0.16(TA Instruments).

For all experiments, the basic procedure for performing TGA involved transferring an aliquot (approximately 3-8mg) of the sample to a platinum sample tray (tray: part number 952018.906, TA Instruments). The disks were placed on a loading platform and then automatically loaded into a Q500 thermogravimetric analyzer using control software. Differential thermograms were obtained by heating samples at 10 ℃/min in a temperature range (typically from room temperature to 400 ℃) in a dry nitrogen stream (compressed nitrogen, grade 4.8 (BOC Gases, Murray Hill, NJ, USA), with a sample purge flow rate of 90L/min and an equilibrium purge flow rate of 10L/min, respectively.

As in FIG. 2, the TGA profile of a co-crystal of VX-950 and 2, 5-dihydroxybenzoic acid (molar ratio of 1) approximately shows a sustained weight loss starting at about 210 ℃.

As in fig. 5, the TGA profile of the co-crystal of VX-950 and 3-methoxy-4-hydroxybenzoic acid (molar ratio also 1) shows a sustained weight loss starting at about 155 ℃.

As in FIG. 8, the TGA profile of the co-crystal of VX-950 and 2, 4-dihydroxybenzoic acid shows a sustained weight loss starting at about 165 ℃.

Example 6 Differential Scanning Calorimetry (DSC)

DSC analysis was performed using an MDSC Q100 differential scanning calorimeter (TA Instruments) with which to control Thermal Advantage Q Series^TMSoftware, version 2.2.0.248, Thermal Advantage Release4.2.1, has the following components: qadv.exe version 2.2build 248.0; RhDII.dII version 2.2build 248.0; rhbase. dii version 2.2build 248.0; rhcomm.dii version 2.2build 248.0; taliense.dii version 2.2build 248.0; and dsc.dii version 2.2build 248.0. In addition, the used analysis software is Universal for Windows 2000/XPAnalysis2000 software, version 4.1D build 4.1.0.16(TA Instruments). The instrument was calibrated using indium.

For all DSC analyses, an aliquot of the sample (approximately 2mg) was weighed into an aluminum sample pan (pan: part number 900786.901; lid: part number 900779.901, TAInstructions). The sample pan was equilibrated at 30 ℃ by closing it with a single pinhole flange and then loaded into a Q100 differential scanning calorimeter equipped with an autosampler. Differential thermograms were obtained by heating each sample at a rate of 50 ℃/min in a temperature range (typically from room temperature to 400 ℃) separately in a stream of dry nitrogen (compressed nitrogen, grade 4.8 (BOC Gases, Murray Hill, NJ, USA), with a sample purge flow rate of 60L/min and an equilibrium purge flow rate of 40L/min.

As in FIG. 3, the DSC thermogram shows that the co-crystal of VX-950 and 2, 5-dihydroxybenzoic acid (1: 1 molar ratio) melts at about 163.48 ℃.

As in FIG. 6, the DSC thermogram shows that the co-crystal of VX-950 and 3-methoxy-4-hydroxybenzoic acid (1: 1 molar ratio) melts at about 178.09 ℃.

As in FIG. 9, the DSC thermogram shows that the co-crystal of VX-950 and 2, 4-dihydroxybenzoic acid (1: 1 molar ratio) melts as an acetonitrile solvate at about 150.81 ℃. Example 7X-ray powder diffraction (XRPD)

XRPD patterns were obtained in reflectance mode at room temperature by using a Bruker D8 Discover diffractometer equipped with a sealed tube source and a Hi-Star area detector (Bruker AXS, Madison, WI, USA). A copper target X-ray tube (Siemens) was operated at 40kV and 35 mA. A graphite monochromator and 0.5mm collimator supplied by Bruker were used to produce parallel monochromatic beams (CuKa,). Distance between sample and detectorAbout 30cm away. The sample was placed on a Si zero background wafer (The Gem Dugout, State College, Pa.) and then placed on The center of an XYZ platform. Data was acquired using GADDS software for Windows NT, version 4.1.16(Bruker AXS, Madison, WI, USA). Two frames were recorded, each at an illumination time of 120 seconds at 2 different 2 θ angles of 8 ° and 26 °. The sample was vibrated in the X and Y directions during irradiation with an amplitude of 1 mm. The data were then integrated over a range of 3 ° to 41 ° 2- θ, step size 0.02 °, and combined to form a continuous map. The instrument was calibrated using a corundum plate (NIST standard 1976).

As shown in FIG. 1, the XRPD pattern of the co-crystal of VX-950 and 2, 5-dihydroxybenzoic acid (1: 1 molar ratio) showed peaks at approximately 8.292, 9.614, 11.675, 12.488, 12.897, 13.120, 14.649, 17.078, 17.514, 18.235, 19.241, and 20.323 degrees 2-theta.

As shown in fig. 4, the XRPD pattern of the co-crystal of VX-950 and 3-methoxy-4-hydroxybenzoic acid (1: 1 molar ratio) shows peaks at about 7.788, 8.640, 9.376, 9.917, 11.410, 11.943, 12.749, 13.166, 14.780, 16.512, 16.909, 17.734, 18.145, 18.823, 19.761, 20.674, 21.702, 22.887, 23.372, 24.042, and 24.863 degrees 2-theta.

As shown in FIG. 8, the XRPD pattern of the co-crystal of VX-950 and 2, 4-dihydroxybenzoic acid (1: 1 molar ratio) showed peaks at approximately 7.581, 8.532, 9.622, 11.859, 12.920, 14.815, 17.291, 17.827, 18.905, and 20.588 degrees 2-theta.

Example 8 solubility analysis

An aliquot of the co-crystal of the invention is placed in a test tube and then the aqueous medium is added. At the stated time points, aliquots of the supernatant were removed, filtered through 0.45PTFE micron filters (Millex, LCR, Millipore) and analyzed by High Performance Liquid Chromatography (HPLC) (Agilent 1100; Palo alto, Calif., USA). The system was equipped with an automatic sampler set at 25 ℃. For sample processing, aliquots of the co-crystal can be diluted with acetonitrile at a 1: 1 v/v ratio. The sample may be tested using a set at 270nmUse of the deviceA phenyl column of 150mmx4.6mm, 3.5 μm particle size (P/N186001144) (Waters, Milford, MA, USA) was run isocratically. The mobile phase may be potassium phosphate buffer (10mM, pH 7.0) to methanol in a 60: 40(v/v) ratio. It can be run at a flow rate of 1mL/min and completed in 15 minutes.

The water solubility data can be determined by equilibrating the co-crystals with water on a shaker for 24 hours at ambient conditions, followed by centrifugation and separation of the saturated solution. Solubility in simulated gastric and intestinal fluids (fed and fasted) can be determined by adding the co-crystal to the simulated fluid under continuous stirring for 24 hours at room temperature. At selected time points, the samples were filtered and the filtrates were analyzed by HPLC.

Example 9 suspension stability

The physical stability of the cocrystals of the invention suspended in an aqueous medium can also be evaluated. Specifically, the co-crystal powder may be slurried at 25 ℃ at the indicated concentration of about 6mg/mL, for example, in (1) unbuffered deionized water and (2) a 1% (w/w) HPMC (low viscosity grade) solution. The slurry was then mixed using a magnetic stir bar and plate. Samples of the solids can be isolated by filtration, for example, at 1, 2, 6, and 24 hour intervals.

Other embodiments

It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

Claims (41)

1. Comprising a co-crystal of VX-950 and 2, 5-dihydroxybenzoic acid.

2. The co-crystal of claim 1, wherein the molar ratio of VX-950 and 2, 5-dihydroxybenzoic acid ranges from about 5: 1 to about 1: 5.

3. The co-crystal of claim 1, wherein the molar ratio of VX-950 and 2, 5-dihydroxybenzoic acid is about 1: 1.

4. The co-crystal of claim 3, having at least two of the four X-ray powder diffraction peaks at about 8.9, 9.6, 11.7, and 17.1 degrees 2-theta, each with a standard deviation of about +/-0.3 degrees 2-theta.

5. The co-crystal of claim 3 having a peak in its DSC thermogram at about 163.48 ℃ with a standard deviation of about +/-5 ℃.

6. Comprising a co-crystal of VX-950 and 2, 4-dihydroxybenzoic acid.

7. The co-crystal of claim 6, wherein the molar ratio of VX-950 and 2, 4-dihydroxybenzoic acid ranges from about 5: 1 to about 1: 5.

8. The co-crystal of claim 6, wherein the molar ratio of VX-950 and 2, 4-dihydroxybenzoic acid is about 1: 1.

9. The co-crystal of claim 8, having at least two of the four X-ray powder diffraction peaks at about 7.6, 8.5, 9.6, and 11.9 degrees 2-theta, each with a standard deviation of about +/-0.3 degrees 2-theta.

10. The co-crystal of claim 8 having a peak in its DSC thermogram at about 150.81 ℃ with a standard deviation of about +/-5 ℃.

11. Comprising a co-crystal of VX-950 and 3-methoxy-4-hydroxybenzoic acid.

12. The co-crystal of claim 11, wherein the molar ratio of VX-950 and 3-methoxy-4-hydroxybenzoic acid ranges from about 5: 1 to about 1: 5.

13. The co-crystal of claim 11, wherein the molar ratio of VX-950 and 3-methoxy-4-hydroxybenzoic acid is about 1: 1.

14. The co-crystal of claim 13, having at least two of the four X-ray powder diffraction peaks at about 7.8, 8.6, 11.4, and 11.9 degrees 2-theta, each with a standard deviation of about +/-0.3 degrees 2-theta.

15. The co-crystal of claim 13 having a peak in its DSC thermogram at about 178.09 ℃ with a standard deviation of about +/-5 ℃.

16. A co-crystal comprising VX-950, a co-crystal former and a solvent, wherein the co-crystal former is selected from the group consisting of 3-methoxy-4-hydroxybenzoic acid, 2, 4-dihydroxybenzoic acid and 2, 5-dihydroxybenzoic acid; the solvent is selected from the group consisting of dichloromethane, acetonitrile, ethyl acetate, ethanol, acetone, and mixtures thereof.

17. The co-crystal of claim 16, wherein the solvent is acetonitrile, methyl chloride, or a mixture thereof.

18. The co-crystal of claim 16, wherein the co-crystal former is 3-methoxy-4-hydroxybenzoic acid.

19. The co-crystal of claim 18, wherein the molar ratio of VX-950 and 3-methoxy-4-hydroxybenzoic acid ranges from about 5: 1 to about 1: 5.

20. The co-crystal of claim 18, wherein the molar ratio of VX-950 and 3-methoxy-4-hydroxybenzoic acid is about 1: 1.

21. The co-crystal of claim 20, having at least two of the four X-ray powder diffraction peaks at about 7.788, 8.640, 11.410, and 11.943 degrees 2-theta, each with a standard deviation of about +/-0.3 degrees 2-theta.

22. The co-crystal of claim 20, having a DSC peak in its DSC thermogram at about 178.09 ℃ with a standard deviation of about +/-5 ℃.

23. The co-crystal of claim 20, wherein the solvent is a mixture of acetonitrile and dichloromethane.

24. The co-crystal of claim 16, wherein the co-crystal former is2, 4-dihydroxybenzoic acid.

25. The co-crystal of claim 24, wherein the molar ratio of VX-950 and 2, 4-dihydroxybenzoic acid ranges from about 5: 1 to about 1: 5.

26. The co-crystal of claim 24, wherein the molar ratio of VX-950 and 2, 4-dihydroxybenzoic acid is about 1: 1.

27. The co-crystal of claim 26, having at least two of the four X-ray powder diffraction peaks at about 7.581, 8.532, 9.622, and 11.859 degrees 2-theta, each with a standard deviation of about +/-0.3 degrees 2-theta.

28. The co-crystal of claim 26, having a DSC peak in its DSC thermogram at about 150.81 ℃ with a standard deviation of about +/-5 ℃.

29. The co-crystal of claim 26, wherein the solvent is a mixture of acetonitrile and dichloromethane.

30. The co-crystal of claim 16, wherein the co-crystal former is2, 5-dihydroxybenzoic acid.

31. The co-crystal of claim 30, wherein the molar ratio of VX-950 and 2, 5-dihydroxybenzoic acid ranges from about 5: 1 to about 1: 5.

32. The co-crystal of claim 30, wherein the molar ratio of VX-950 and 2, 5-dihydroxybenzoic acid is about 1: 1.

33. The co-crystal of claim 32, having at least two of the four X-ray powder diffraction peaks at about 8.292, 9.614, 11.675, and 17.078 degrees 2-theta, each with a standard deviation of about +/-0.3 ° 2-theta.

34. The co-crystal of claim 32, having a DSC peak in its DSC thermogram at about 163.48 ℃ with a standard deviation of about +/-5 ℃.

35. The co-crystal of claim 32, wherein the solvent is a mixture of acetonitrile and dichloromethane.

36. A pharmaceutical composition comprising a VX-950 co-crystal, comprising VX-950 and a co-crystal former selected from the group consisting of 3-methoxy-4-hydroxybenzoic acid, 2, 4-dihydroxybenzoic acid, and 2, 5-dihydroxybenzoic acid.

37. The pharmaceutical composition of claim 36, wherein the molar ratio of VX-950 and the co-crystal former ranges from about 5: 1 to about 1: 5.

38. The pharmaceutical composition of claim 36, wherein the molar ratio of VX-950 to the co-crystal former is about 1: 1.

39. The pharmaceutical composition of claim 36, further comprising a pharmaceutically acceptable diluent, solvent, excipient, carrier, or solubilizing agent.

40. A method of making the co-crystal of any one of claims 1-35, comprising:

a. providing a solution of VX-950 in a solvent,

b. providing a co-crystal former selected from the group consisting of 3-methoxy-4-hydroxybenzoic acid, 2, 4-dihydroxybenzoic acid and 2, 5-dihydroxybenzoic acid,

c. grinding, heating, co-subliming, co-melting or contacting in solution VX-950 with a co-crystal former under crystallization conditions to form a solid phase co-crystal, and

d. optionally isolating the co-crystal formed in step (c).

41. A method of modulating a chemical or physical property of interest of the co-crystal of any one of claims 1-35, comprising:

a. measuring chemical or physical properties of interest of VX-950 and a co-crystal former selected from the group consisting of 3-methoxy-4-hydroxybenzoic acid, 2, 4-dihydroxybenzoic acid and 2, 5-dihydroxybenzoic acid,

b. determining the molar fraction of VX-950 and co-crystal former that results in the desired modulation of the chemical or physical property of interest, and

c. preparing a co-crystal using the mole fraction determined in step (b).