Patent Publication Number: US-2003225279-A1

Title: Stavudine Polymorphic Form 1 process

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS  
     [0001] This is a non-provisional application which claims the benefit of provisional applications, U.S. S. No. 60/230,261, filed Sep. 6, 2000, and U.S. S. No. 60/231,766, filed Sep. 12, 2000. 
    
    
     
       FIELD OF THE INVENTION  
       [0002] This invention relates to an improved process for obtaining Polymorphic Form I of the antiretroviral compound Stavudine, useful in the treatment of retroviral infections, particularly HIV infections.  
       BACKGROUND OF THE INVENTION  
       [0003] Stavudine, also known as d4T, is approved by the U.S. Food &amp; Drug Administration for the therapeutic treatment of patients infected with retroviruses. Stavudine chemically is 2′, 3′-didehydro-3′-deoxythymidine. The compound, a nucleoside reverse transcriptase inhibitor, and its preparation are disclosed, for example, in U.S. Pat. No. 4,978,655, issued Dec. 18, 1990. It is known that Stavudine is effective in the treatment of infections caused by retroviruses such as murine leukemia virus and human immunodeficiency virus, i.e. HIV; HTLV III/LAV virus (the AIDS virus). Stavudine has enjoyed notable commercial success since its introduction.  
       [0004] It is known that Stavudine exists in three polymorphic forms that differ in solubility, designated as Forms I, II and III, respectively. Forms I and II are anhydrous polymorphs whereas Form III is hydrated and is pseudopolymorphic with Forms I and II. Of the three forms, Form I is stable and shows no transformations to other polymorphic forms, thus demonstrating its greater thermodynamic stability relative to the other Forms.  
       [0005] The phenomenon of polymorphism, the capacity of a substance to occur in different crystalline forms in the crystalline solid state, is well known, as are its ramifications on the process of drug development. The various characteristics and properties of the polymorphic forms of a substance, e.g. shape, color, density, dissolution properties and the like, will make one or more specific polymorphic forms desirable over the others for production and/or pharmaceutical compounding. As a result, a very early step in the process of product development of a new pharmaceutical agent is the determination of whether it exists in polymorphic forms and, if so, which of such forms possesses advantages for development of the eventual commercial pharmaceutical. In the instance of Stavudine, Form I has been found to be the most thermodynamically stable form, with no tendency for solid state conversion to the other Forms. Hence, it is Polymorphic Form I of Stavudine that is offered commercially under the trademark Zerit®.  
       [0006] U.S. Pat. No. 5,608,048, issued Mar. 4, 1997, teaches a process whereby Polymorphic Form I of Stavudine is prepared in substantially pure form from a mixture containing it in combination with one or more of Polymorphic Forms II and III. This process involves dissolving the mixture under anhydrous conditions in an organic solvent to form a saturated solution at a temperature of at least about 65° C. and continuously cooling the solution with stirring until precipitation of Stavudine Polymorphic Form I is completed. A requirement of the process, however, is that the rate of cooling cannot exceed about 20° C. per hour until the temperature falls below 40° C. In a preferred embodiment, the temperature is reduced about 10° C. over 15 minutes, then held for an hour and the steps repeated until the solution temperature falls below 40° C. There are disclosed further embodiments consisting of gradients in cooling the solution of the mixture of Polymorphic Forms.  
       [0007] The solvent utilized in the process described in U.S. Pat. No. 5,608,048 is selected from the group of methanol, ethanol, n-propanol, isopropanol, acetonitrile and ethyl acetate. It is emphasized, as stated previously, that the process must be carried out under anhydrous conditions. It will be appreciated that this process suffers from a number of disadvantages, among which are strict requirements in time and temperature management and control as well as strict moisture control. In accordance with the present invention, a method has been found whereby Stavudine Polymorphic Form I can readily be produced without such strict process control requirements.  
       SUMMARY OF THE INVENTION  
       [0008] Stavudine Polymorphic Form I is produced in high yield and purity in a dry particulate form from a mixture comprising it and at least one of Polymorphic Forms II and III by a Solution-Enhanced Dispersion by Supercritical Fluids (SEDS) technique utilizing a particular solvent mixture as a vehicle. The process is carried out at constant temperature, without the requirement of constant stirring and further without the need for filtration and drying steps.  
       DETAILED DESCRIPTION OF THE INVENTION  
       [0009] Solution-Enhanced Dispersion by Supercritical Fluids is a recognized technique known under the trademark SEDS, owned by Bradford Particle Design Limited, Bradford, West Yorkshire, England. It is described, for example, in U.S. Pat. No. 5,851,453, issued Dec. 22, 1998. The process is advantageous in that it can be utilized to control the polymorphic form of a drug substance in a single processing step. This control is achieved by operating the SEDS process under varied process parameters, primarily temperature, solvent composition and rate of crystallization until optimum conditions are determined for the desired polymorphic form. Particles produced utilizing the SEDS technique are free from static charge and contain only trace amounts of residual solvent. A further advantage of the process is that the particles are formed dry, thus eliminating the need for filtration and solvent removal, the latter being of particular advantage in terms of both cost and environmental considerations.  
       [0010] In the SEDS process, a solution of the material of interest is introduced into a chamber, designated a particle formation vessel, through a specially designed nozzle under stable conditions of temperature and pressure in combination with a supercritical fluid. The nozzle is essentially a coaxial design or the equivalent that produces a mixing of the two fluids being introduced at the point where they enter the chamber. The supercritical fluid mixes with, disperses and rapidly extracts the solvent from the solution. The insolubility of the solute in the supercritical fluid-solvent mixture induces the formation of particles by an antisolvent precipitation mechanism. By manipulating the various working parameters of pressure, temperature, solution concentration and flow rates in the nozzle, it is possible to control the size, shape and morphology of the product particles formed in the vessel. The co-introduction of the solution or dispersion of the desired substance and the supercritical fluid into the particle formation vessel creates, substantially simultaneously, dispersion and extraction of the vehicle or solvent by the action of the supercritical fluid. As the dissolved material is freed of solvent and dispersed by the supercritical fluid at the same time, the process produces discrete, dry particles that are retained in the vessel.  
       [0011] As utilized herein, the term “supercritical fluid” means a fluid substantially at or above its critical pressure (Pc) and critical temperature (Tc) simultaneously. As a practical matter in the application of the process in accordance with the present invention, the pressure of the fluid is likely to be in the range 1.01 Pc-7.0 Pc, preferably substantially above the Pc of the fluid, and the temperature in the range 1.01 Tc-4.0 Tc, preferably slightly above the Tc of the fluid. Suitable chemicals that can be utilized as supercritical fluids in the process of the present invention include carbon dioxide, nitrous oxide, sulfur hexafluoride, xenon, ethylene, chlorotrifluoromethane, ethane and trifluoromethane. In is essential to the efficient operation of the process that the supercritical fluid be an antisolvent for the desired product. Particularly preferred for the present process is supercritical carbon dioxide since Stavudine is practically insoluble therein.  
       [0012] In accordance with the process of the present invention, the supercritical fluid, preferably carbon dioxide, and a solution or dispersion of the material to be produced are introduced into the particle formation vessel through a coaxially designed nozzle as described in detail in U.S. Pat. No. 5,851,453. In the instance of the present process, the supercritical fluid is co-introduced with a solution comprising a mixture of Stavudine Polymorphic Form I and at least one of Polymorphic Forms II and III as formed in the synthesis thereof described in U.S. Pat. No. 5,608,048 utilizing thymidine as the starting material. The synthetic route for the preparation of Stavudine as described in said patent does not form part of the process of the present invention and will not be discussed in detail herein. Further, although the synthetic route disclosed in said patent is preferred, the particular pathway utilized to produce a mixture of Stavudine Polymorphic Form I and at least one of Forms II and III to be treated in accordance with the process of the present invention is not critical thereto. It must be borne in mid that, while the mixture of polymorphic forms of Stavudine to be treated in accordance with the present invention is in a purified state, it typically will contain entrained solvent, for example, DMSO, toluene and the like, as well as other impurities resulting from the synthesis. It has been found that such impurities, particularly the entrained solvents, are materially removed in accordance with the subject process. Even synthetic impurities, particularly those that are non-polar and will dissolve in the supercritical fluid, are likewise removed by the process of the present invention.  
       [0013] The solution containing a mixture comprising the polymorphic forms of Stavudine as described above and the supercritical fluid are co-introduced into the particle formation vessel such that there is instantaneous mixing of the two at the point of entry. The supercritical fluid is introduced under pressure and at a high flow rate in comparison to the solution containing the mixture of Stavudine polymorphic Forms. While not wishing to be bound by any particular theory or explanation of the phenomena taking place within the vessel, it is believed that the high velocity supercritical fluid causes the solvent of the solution to be broken up into droplets or other analogous fluid elements from which the vehicle/solvent is substantially simultaneously extracted by the supercritical fluid and dispersed, thereby resulting in the formation of discrete particles of the solid previously held in solution. Further, the high shearing action of the high velocity supercritical fluid ensures both dispersion of the vehicle/solvent and thorough mixing with the supercritical fluid thereby causing substantially immediate extraction thereof with the resultant formulation of discrete, dry particles of Stavudine Polymorphic Form I.  
       [0014] As described in U.S. Pat. No. 5,851,453, the nozzle utilized to introduce the supercritical fluid and the solution of Stavudine Polymorphic Forms into the vessel may be configured in various ways to achieve optimum mixing and dispersion. For example, an axial nozzle having three passages can be utilized to introduce a flow of the solution sandwiched between an inner and an outer flow of the supercritical fluid to achieve enhanced dispersion and, hence, greater control over, and uniformity of, the particle size of Stavudine Polymorphic Form I. Regardless of the configuration of the nozzle, at least one of the passages therein carries a flow of the solution and at least one of the passages carries a flow of the supercritical fluid.  
       [0015] The particle formation vessel is equipped with a retention means, such as a fine mesh screen, to catch and hold the particles of Stavudine Polymorphic Form I as they are formed therein. The apparatus is typically equipped at its outlet with a back-pressure regulator to maintain the particle formation vessel at the required operating pressure. The effluent from the back-pressure regulator is fed into a separator where it is decompressed to the gaseous state so that it may be recycled into the system if desired. The solvent for the solution will also separate as a liquid and may be collected and recycled, utilized in other applications or discarded. The system may be operated continuously or in a batch mode. When a sufficient amount of particles is collected in the vessel, the flow of solution is discontinued and the particles are dried by continued flushing with only pure supercritical fluid and then removed. A system may be operated with two particle formation vessels so that, while particles are being collected from one and it is being flushed and prepared to receive a renewed flow of solution, the other is producing. As those of ordinary skill in the art will appreciate, running the two vessels out of phase as described will assure continuous production.  
       [0016] The benefits of the process of forming Stavudine Polymorphic Form I in accordance with the process of the present invention are that it can be run isothermally, hence multiple depressurizing and pressurizing steps are not required, there is a considerable time saving in the eliminating of the drying and solvent removal steps and there is less likelihood of exposure of workers to reagents, particularly solvents, during the recovery step. In addition, the time-consuming and temperature control-sensitive technique previously utilized is no longer required. The same is true of the requirement in the previous process that the solution of the mixture of Polymorphic Forms of Stavudine be constantly stirred during the cooling process. The process of the present invention affords Stavudine Polymorphic Form I in higher yield than has heretofore been realized and in higher purity. The higher purity is possible since the present process removes a higher percentage of entrained solvents, including residual solvents from the synthesis. The particles of Stavudine Polymorphic Form I formed in accordance with the present process contain less than 100 ppm of entrained solvents. The particles size of Stavudine Polymorphic Form I formed in accordance with the present process is also advantageous over that previously available since the particles have an average size of from about 20 to about 40 microns whereas those from the previous manufacturing process range up to about 200 microns. Stavudine Polymorphic Form I formed by the present process is more polymorphically stable than that formed by the previous process as a result of the reduction in residual isopropyl alcohol and synthesis solvents since residual solvents have been shown to induce solid state transition upon storage.  
       [0017] In accordance with the process of the present invention, it has been found that a particular solvent combination for dissolution of a mixture of Stavudine Polymorphic Form I and at least one of Form II and Form III yields Form I in very high yield and purity in the SEDS process. Experiments were conducted with a wide variety of solvent combinations and process conditions and, as a result, it was determined that a mixture of isopropyl alcohol and water provided the optimum results. It has been found in accordance with the present invention that the presence of water in the solvent is essential since in the absence of water, all solvents tested as possible vehicles for the subject process produced Stavudine Polymorphic Form II to a significant degree.  
       [0018] The solution to be processed in accordance with the present invention preferably contains from about 0.1% to about 2%, most preferably about 1%, weight to volume of the mixture of Polymorphic Forms of Stavudine in a solvent mixture preferably from about 96:4 to 94:6, most preferably about 95:5, volume to volume isopropanol and water. The flow rates into the particle formation vessel are preferably a ratio of Stavudine solution to supercritical fluid of from about 0.005:1.0, most preferably about 0.02:1.0. The temperature and the pressure in the particle formation vessel are controlled during the process such that the temperature is above the Tc of the supercritical fluid and the pressure is substantially above the Pc of the supercritical fluid. Using carbon dioxide as the supercritical fluid, the temperature in the vessel is preferably from about 31.4 to 50° C., most preferably about 35° C., and the pressure is preferably from about 80 to 115 bar, most preferably about 90 bar.  
       [0019] In order to illustrate the unexpected nature of the solvent combination of the present invention in the SEDS process, starting with identical mixtures of the polymorphic forms of Stavudine, a solvent mixture of isopropyl alcohol and water containing 10% by volume water produced predominately Stavudine Polymorphic Form III; a solvent mixture containing 7.5% by volume water produced a mixture of Forms I and II; and a solvent mixture containing 2.5% by volume produced predominately Form II, all by analogous SEDS techniques. In view whereof, it is considered unexpected that the solvent combination of the invention produces pure Stavudine Polymorphic Form I. It is also considered unexpected that Form I can be produced by a process involving rapid cooling since the previous process is dependent on slow cooling under very controlled conditions with constant stirring. Further in view of the fact that U.S. Pat. No. 5,608,048 teaches that Stavudine Polymorphic Form I can only be produced under strict anhydrous conditions, it is considered unexpected that any combination of solvents containing water will even produce Form I. There is certainly no teaching in the patents discussed above that would suggest that, in the SEDS process, a solvent combination containing water would yield Stavudine Polymorphic Form I in high yield and high purity.  
       [0020] It is understood that various other embodiments and modifications in the practice of the invention will be apparent to, and can be readily made by, those of ordinary skill in the art without departing from the scope and spirit of the invention as described above. Accordingly, it is not intended that the scope of the claims appended hereto be limited to the exact description set forth above, but rather that the claims be construed as encompassing all of the features of patentable novelty that reside in the present invention, including all the features and embodiments that would be treated as equivalents thereof by those skilled in the relevant art. The invention is further described with reference to the following experimental work. 
     
    
    
     Example 1  
     [0021] The following series of experiments was conducted in an SEDS apparatus such as described in U.S. Pat. No. 5, 851,453 and consisting essentially of a particle formation vessel, an axial configured double nozzle connected to a source of test solution under pressure and a source of carbon dioxide supercritical fluid under pressure, a means for particle retention within the vessel, a back-pressure regulator and a separator for separating the solvent from the carbon dioxide which reforms as a gas under reduced pressure. Various solvents were utilized to dissolve a mixture of Polymorphic Forms I, II and III of Stavudine. The results are shown in Table I.  
                                                   TABLE I                               Sol.       Solution   CO 2     DSC           SEM       Press   Temp   Conc.       Flow   Flow   Peak   XRPD   Yield   Morphology       bar   ° C.   w/v %   Solvent   mlmin −1     Mlmin −1     ° C.   Forms   %   Description                                                                        200   35   5   MeOH   n/a   1   —   II   18   Acicular,                                           some &gt;600μ       200   35   3   IPA   n/a   1   —   I &amp; II   63   Acicular,                                           some &gt;600μ       120   35   1   Aceton   0.3   20   —   II   62   Acicular,                                           up tp 100μ       120   70   1   Aceton   0.3   18   171.5   II   47   Acicular,                                           10-20μ       250   35   1   Aceto   0.2   10   172.3   II   27   Plates                   nitrile                       up tp 100μ       80   70   1   Aceto   0.2   10   172.6   I &amp; II   50   Plates                   nitrile                       &gt;100μ       250   35   1   IPA   0.2   10   172.7   II   36   Acicular/                                           Plates                                           some &gt;300μ       80   70   10   IPA   0.2   10   167.4   II   33   Plates &gt;20μ       150   50   8   H 2 O   0.03   8           0   No product       80   90   2   IPA   0.3   15   168.6   I &amp; II   68   Plates,                                           20-30μ       80   120   20   IPA   0.3   15   —   I &amp; II   62   Plates,                                           20-30μ       85   120   1   Aceto-   0.3   15   —   I &amp; II   41   Plates,                   nitrile                       20-30μ       90   90   0.75   Ethyl   0.3   15   —   II   52   Acicular,                   Acetate                       Part. 1-3μ       80   90   1   EtOH   0.3   15   —   II   53   Fused                                           plates &gt;10μ       80   90   1   IPA   1   10   —   —   0   No product       250   35   1   IPA   1   10   —   —   0   Trace       250   40   1   t-BuOH   0.3   15   —   —   0   Abandoned       250   35   5   DMSO   0.3   15   —   II   33   Plates,                                           20-30μ       150   35   1   EtOH   0.4   9   170.8   II   68   Large Rods                  
 
     [0022] In Table I, the DSC peak was obtained by accurately weighing a sample of between 2 and 5 mg and scanning it in a pierced, crimped aluminum pan by differential scanning calorimetry (DSC7, Perkin Elmer Ltd., UK). Since the melting points of the three polymorphic forms of Stavudine are very similar, this method was not utilized to determine the polymorphic form of the product. Polymorphic form was determined by X-ray powder diffraction (XRPD) using a Siemens model D-5000 diffractometer. Test samples were ground to a fine powder, using a mortar and pestle. The random orientation of the resulting crystallites ensures that every possible reflection place was represented parallel to the specimen surface. Data was collected between 1.5°, and 40° 2┘, using CuK 4  with a step increase of 0.05° and count intervals of three seconds. The SEM observation was carried out using a Hitachi S-520 electron microscope. Small quantities of sample were affixed to aluminum SEM stubs, coated with a conducting layer of gold and examined under a range of magnifications. The resulting micrographs were used to determine and describe particle morphology and estimate particle size.  
     [0023] The results shown in Table I demonstrate that, of the range of solvents and conditions tested, none was optimum for producing Stavudine Polymorphic Form I.  
     Example 2  
     [0024] The following experimental runs were conducted utilizing mixtures of isopropanol (IPA) and water. All runs were conducted at a vessel temperature of 35° C., a flow of supercritical carbon dioxide of 9 mlmin −1  and, with the exception of the final two runs that were conducted at a solution flow rate of 0.3 mlmin −1 , all runs were at a solution flow rate of 0.2 mlmin −1 . The results are reported in Table II.  
                                           TABLE II                           Sol.   % H 2 O   DSC                       Press.   Conc.   plus   Peak   XRPD   Yield   SEM           Bar   % w/v   IPA   ° C.   Form   %   Morphology   Comments                                                                120   1   10%    168.5   III   37   Acicular &gt;500μ   Weight loss 2.94%       120   1   10%    168.7   III   43   Mix of large %   Particles different than                               small acicular   previous run                               plates to 100μ       120   1   5%   170.2   I   91   Long, well-   Strong IPA smell in                               defined needles   powder                               20-40μ       120   1   5%   168.9   I &amp; II   83   Large acicular   Longer drying time                               particles, some   than previous run.                               chunks   Product was dry       120   1   5%   169.5   I &amp; II   —   Long, well-   Strong IPA smell,                               defined needles   predominately Form I       90   1   5%   169.7   I   85   Long, well-   Dry, similar                               defined needles   morphology to starting                               20-100μ   material       90   0.5   5%   170.6   I   92   Long, well-   Reduced solution                               defined needles   conc. Similar to                               20-80μ   starting material       90   0.5   5%   170.6   I   87   Long, well-   Reduced solution                               defined needles   conc. Similar to                               up to 100μ   starting material       90   0.5   5%   169.7   I &amp; II   79   Long, well-   Raised filter and                               defined needles   utilized different                               up to 150μ   nozzle       90   1   5%   170.2   I   95   Long, well-   Original nozzle, no                               defined needles   raised filter, increased                               up to 100μ   conc.*       90   0.5   5%   169.6   I   75   Long, well-   Original nozzle, no                               defined needles   raised filter                               up to 150μ       90   1   5%   —   I   90   Long, well-   500 ml vessel used, no                               defined needles   sign of Form II                               up to 100μ       90   0.5   2.5%     —   II   78   Long   Water content reduced                               needles/flat   for effect                               bars, some &gt;200μ       90   0.5   7.5%     —   I &amp; II   69   Very long sharp   Water content                               needles, some   increased for effect                               &gt;300μ       90   1   5%   —   I   93       Increased throughput                                   in 50 ml vessel       90   1   5%   —   I   90       500 ml vessel used                  
 
     [0025] The run with the increased concentration (*) was conducted with a supercritical carbon dioxide flow rate of 10 mlmin −1 . The weight loss in the first run was determined using Thermogravimetric Analysis (TGA 7, Perkin Elmer Ltd.) by heating samples in open pans at 10° C. min −1  between 25 and 200° C. These results demonstrate preferred conditions for producing Stavudine Polymorphic Form I in accordance with the subject process utilizing mixtures of 95:5 IPA and water.  
     Example 3  
     [0026] Residual solvent analysis was performed on samples of Stavudine Polymorphic Form I prepared in accordance with the process of the present invention and commercial material that had not been processed in accordance with the present process. The analysis was performed using headspace-gas chromatography having the capacity to quantify residual isopropyl alcohol levels up to 2023 ppm using external standardization. Deionized water was utilized as the solvent as it is not detected by flame ionization detection, hence does not interfere with the analysis. Standard solutions of Stavudine Polymorphic Form I were prepared with concentrations up to 500 μgml −1 . Test samples solutions containing high Stavudine concentrations between 5 and 25 mg ml −1  were prepared and tested in sealed vials in a Varian Star 3400cx with Flame Ionization Detector, Varian, UK. The results of analysis of the headspace in each sealed vial are given in Table III below.  
                               TABLE III                                       Total ppm       Processed   Total Sample   (μg) Equivalent   Actual Total   IPA in the       (Y/N)   Wt. in Grams   to 1 ppm   (μg) Present   Sample                                                    Unprocessed   0.04302   0.04302   10.326   240       Unprocessed   0.4828   0.04828   10.806   224       Processed   0.02013   0.02013   0.992   49       Processed   0.02   0.02   1.082   54       Processed   0.02419   0.02419   2.016   83                  
 
     [0027] The benefit of the subject process in terms of residual solvent content of processed Stavudine Polymorphic Form I are clearly demonstrated by the data in Table III.