Stable Amorphous Tigecycline

STABLE AMORPHOUS TIGECYCLINE Field of the Invention The present invention provides stable amorphous tigecycline, a process for its preparation, a pharmaceutical composition comprising it, and its use for the treatment of complicated skin and skin structure infections, complicated intra-abdominal infections, and community-acquired bacterial pneumonia. Background of the Invention Tigecycline is a tetracycline antibiotic having the structure depicted by Formula I. Formula I Tigecycline is chemically (4S,4aS,5aR,12aS)-9-[2-(tert-butylamino)acetamido]-4,7-bis(dimethy1amino)- 1,4,4a,5,5a,6,11,12a-octahydro-3,10,12,12a-tetrahydro1xy,-l l- dioxo-2-naphthacenecarboxamide. It is marketed in the United States as a lyophilized powder or cake for intravenous infusion under the brand name ~ ~ ~ a cfoirl t'h e treatment of complicated skin and skin structure infections, complicated intra-abdominal infections, and community-acquired bacterial pneumonia. Since the drug substance does not contain excipients or preservatives, the stability of the amorphous material used to prepare intravenous infusions is a key parameter. Tigecycline is known to degrade into its C4 epimer of Formula I1 through epimerization. Formula I1 Although the tigecycline epimer of Formula I1 is believed to be non-toxic, under certain conditions it may lack the anti-bacterial efficacy of tigecycline, and may, therefore, be an undesirable degradation product. Moreover, the amount of epimerization can be magnified during the synthesis of tigecycline on a large scale. Epimerization of tigecycline occurs quickly under acidic conditions and at elevated temperature. Due to the increase in epimerization with an increase in temperature, in order to reduce the rate of epimer formation, production and storage of tigecycline at low temperatures is desirable. Several methods are reported in the literature for minimizing epimer formation in tetracyclines, such as maintaining a pH of greater than about 6.0 during processing; avoiding contact with conjugates of weak acids such as forrnates, acetates, phosphates or boronates; and avoiding contact with moisture. However, formulating tetracycline analogs in non-aqueous vehicles to improve drug stability is found to be more appropriate for topical use than for parenteral use. In addition to the C4 epimer, other impurities known to form during the preparation of tigecycline include oxidation and degradation by-products. Oxidation of tigecycline may be more pronounced under basic reaction conditions and more so in large-scale operations since processing times are typically longer and the compounds are in contact with the base for a longer time. Degradation by-products may be formed during various steps of the manufacturing process, and separating tigecycline from these degradation by-products can be tedious. These disadvantages may hamper a large-scale synthesis. Processes for the preparation of amorphous tigecycline are reported in U.S. Patent No. 5,675,030; U.S. Patent Application Nos. 200710026080 and 200910275766; and PCT Publication Nos. WO 20071127292, WO 20081066935, WO 20091092680, and WO 20101084325, which are 3 incorporated herein for reference. However, due to the propensity for tigecycline to degrade, amorphous tigecycline is prepared and processed under low-oxygen and low-temperature conditions. Such processing is expensive because it requires special equipment and handling. Due to difficulties encountered in reducing the amount of C4 epimer and the formation of oxidative by-products during manufacturing of tigecycline, there exists a need in the art for a stable and consistently reproducible polymorph of tigecyline having a low level of C4 epimer impurities and favorable characteristics that meets the bioavailability requirement of an intravenous formulation. Summary of the Invention The present invention provides stable amorphous tigecycline. Stable amorphous tigecycline of the present invention is reproducible, contains low levels of C4 epimer and oxidative by-products, and is stable towards polymorphic conversion. Stable amorphous tigecycline of the present invention exhibits good flowability, filtration, and dissolution properties to meet the bioavailability requirements of an intravenous formulation. The process for the preparation of stable amorphous tigecycline of the present invention is simple and cost-effective as it does not require the use of special equipment such as a spray-dryer, and is suitable for large scale production. A first aspect of the present invention provides stable amorphous tigecycline. A second aspect of the present invention provides a process for the preparation of stable amorphous tigecycline comprising the steps of: (a) contacting tigecycline with water and a mixture of organic solvents; (b) adding an alcohol; and (c) isolating stable amorphous tigecycline. A third aspect of the present invention provides stable amorphous tigecycline having less than 0.5% of C4 epimer. A fourth aspect of the present invention provides stable amorphous tigecycline having HPLC purity greater than 99%. A fifth aspect of the present invention provides a pharmaceutical composition comprising stable amorphous tigecycline and one or more pharmaceutically acceptable carriers, diluents, or excipients. A sixth aspect of the present invention provides the use of stable amorphous tigecycline for the treatment of complicated skin and skin structure infections, complicated intra-abdominal infections, and community-acquired bacterial pneumonia. Brief Description of the Drawings Figure 1 : X-ray powder diffraction pattern (XRPD) of stable amorphous tigecycline. Figure 2: Differential scanning calorimetry (DSC) of stable amorphous tigecycline. Figure 3: Infra-red spectrum (IR) of stable amorphous tigecycline. Figure 4: Therrno-gravimetric analysis (TGA) of stable amorphous tigecycline. Figure 5: X-ray powder diffraction pattern (XRPD) of stable amorphous tigecycline after keeping on stability. Detailed Description of the Invention Various embodiments and variants of the present invention are described hereinafter. The term "ambient temperature" as used herein, refers to temperature in the range of about 20°C to about 35°C. The term "contacting" as used herein, refers to dissolving, slurrying, stirring, or a combination thereof. Stable amorphous tigecycline of the present invention exhibits a characteristic X-ray powder diffraction peak at about 9.72 (9.09 A) -+ 0.2" 28 and a halo maximum in the range of about 16-24 * 0.2" 20. Figure 1 provides the X-ray powder diffraction pattern of stable amorphous tigecycline of the present invention. Stable amorphous tigecycline of the present invention may also be characterized by DSC, as having broad endotherms at about 79.09"C, 157.1 1 "C and 18 1.1 O°C, and a sharp exotherm at about 224.0°C. Figure 2 provides the DSC of stable amorphous tigecycline of the present invention. Stable amorphous tigecycline of the present invention may be further characterized by IR spectrum as depicted in Figure 3 and TGA as depicted in Figure 4. Tigecycline to be used as starting material for the preparation of stable amorphous tigecycline of the present invention may be obtained by any of the processes reported in literature such as those disclosed in U.S. Patent No. 5,675,030 and PCT Publication Nos. WO 2006113043 1, WO 20061130500, WO 200611 3041 8, WO 20081066908, and WO 200811 34048, which are incorporated herein by reference. Organic solvents to be used for the preparation of stable amorphous tigecycline of the present invention may be selected from alcohols, chlorinated hydrocarbons, or mixtures thereof. Examples of alcohols may include methanol, ethanol, n-propanol, n-butanol, iso-butanol, and sec-butanol. Examples of chlorinated hydrocarbons may include dichloromethane, dichloroethane, chloroform, and carbon tetrachloride. In the preferred embodiments of the present invention, methanol and dichloromethane may be used as organic solvents. Preparation of stable amorphous tigecycline may be carried out by adding tigecycline to a reaction vessel containing de-ionized water at ambient temperature. The reaction mixture is cooled and organic solvents are added. The pH of the reaction mixture is adjusted to about 6.8 by adding concentrated hydrochloric acid:water (1 : 10) mixture. Optionally, a solution of a base such as ammonium hydroxide may be added to adjust the pH of the reaction mixture. The reaction mixture is stirred for about 5 to about 30 minutes and extracted to obtain a wet solid. Methanol is added to the wet solid at about 45°C to about 50°C. The reaction mixture is filtered, washed with methanol, and stirred at ambient temperature for about 1 to about 5 hours followed by isolation to obtain stable amorphous tigecycline. Isolation may be carried out by concentration, precipitation, cooling, filtration, centrifugation, or combinations thereof, then dried. Drying may be carried out using any suitable method such as drying under reduced pressure, air drying, or vacuum tray drying at ambient temperature to about 100°C for about 2 to about 20 hours. Stable amorphous tigecycline of the present invention has an HPLC purity greater than 99%. Stable amorphous tigecycline of the present invention contains less than 0.5% of C4 epimer. Stable amorphous tigecycline of the present invention exhibits good flowability, filtration, and dissolution properties to meet the bioavailability requirements of intravenous formulation. - 1 op,lfm b #i ; .'. * n ; b b c ~ ($"y &y * I - - ? . - - - Stability assays of amorphous tigecycline of the present invention were performed after intervals of 3 months and 6 months. The samples were packed in a triple pouch which was kept in a high-density polyethylene (HDPE) container. The triple pouch was made up of three separate pouches - the inner pouch was made-up of low-density polyethylene (LDPE) which was twisted and tied; the middle-pouch was made-up of high molecular high density polyethyene (HMHDPE), or low-density polyethylene (LDPE), or linear low-density polyethylene (LLDPE); and the outer pouch was made of polyester film, aluminum foil, or low-density polyethylene (LDPE) film. The middle and outer pouches were flushed with nitrogen gas followed by vacuumisation, filled with nitrogen gas, and then heat sealed. The stability tests were carried out at a temperature of about 5°C k 3°C. Table 1 summarizes the results of the stability assays carried out on amorphous tigecycline of the present invention. Table 1 : Stability Assays As is evident from the data provided in Table 1, stable amorphous tigecycline of the present invention consistently contains less than 0.5% of C4 epimer. Stable amorphous tigecycline of the present invention does not get converted into other polymorphs of tigecycline, as is evident from the X-ray powder diffraction pattern provided in Figure 5. Schedule Initial 3 months 6 months Stable amorphous tigecycline of the present invention may be administered as part of a pharmaceutical composition for the treatment of complicated skin and skin structure infections, complicated intra-abdominal infections, and community-acquired bacterial pneumonia. Accordingly, in a further aspect of the present invention, there is provided a pharmaceutical composition comprising stable amorphous tigecycline and one or more pharmaceutically acceptable carriers, diluents, or excipients, and optionally other therapeutic ingredients. Assay (by HPLC; wlw) 99.2% 99.0% 99.3% Water (w/w> 0.86% 0.85% 0.92% In the foregoing section, embodiments are described by way of example to illustrate the processes of invention. However, this is not intended in any way to limit the scope of the 7 CJ Epimer (w/w> 0.35% 0.35% 0.41% Total impurities (w/w> 0.39% 0.46% 0.62% present invention. Several variants of the example would be evident to persons ordinarily skilled in the art which are within the scope of the present invention. Example Preparation of Stable Amorphous Tigecycline To a reaction vessel flushed with nitrogen gas, de-ionized water (80 mL) was added at ambient temperature. The contents were cooled to about 10°C. Tigecycline (10 g) dissolved while stirring and then cooled to about 0°C. Methanol (20 mL) was added and the contents were stirred for about 5 minutes. Dichloromethane (700 mL) was added. The pH of the reaction mixture was adjusted to about 6.8 by adding a solution of concentrated hydrochloric acid:water (1 : 10; 2 mL) and ammonium hydroxide solution (1 : 10; 1 mL). The reaction mixture was stirred at about 5°C for about 10 minutes, extracted with dichloromethane (200 mL), filtered through sodium sulphate, and washed with dichloromethane (1 00 mL) to obtain a wet solid. Methanol (700 mL) was added to the wet solid at about 45°C to about 50°C. The solution was filtered through a hyflobed under a nitrogen atmosphere, followed by washing the hyflobed with methanol (300 mL). The filtrate was stirred at about 20°C to about 25°C for about 3 hours under a nitrogen atmosphere, filtered, washed with methanol (40 mL), dried at ambient temperature for about 30 minutes,then dried at about 60°C to about 65°C for about 12 hours to obtain stable amorphous tigecycline. Yield: 0.69% wlw Chromatographic purity: 99.53% C4 epimer: 0.35%

WE CLAIM:- 1. Stable amorphous tigecycline. 2. The stable amorphous tigecycline according to claim 1, characterized by an X-ray diffraction pattern substantially similar to Figure 1. 3. The stable amorphous tigecycline according to claim 1 characterized by a DSC substantially similar to Figure 2. 4. The stable amorphous tigecycline according to claim 1, characterized by an IR spectrum substantially similar to Figure 3. 5. The stable amorphous tigecycline according to claim 1, characterized by a TGA substantially similar to Figure 4. 6. A process for the preparation of stable amorphous tigecycline comprising the steps of: (a) contacting tigecycline with water and a mixture of organic solvents; (b) adding an alcohol; and (c) isolating stable amorphous tigecycline. 7. The process according to claim 6, wherein the mixture of organic solvents in step (a) comprises alcohols and chlorinated hydrocarbons. 8. The process according to claim 7, wherein the alcohol is selected from the group comprising methanol, ethanol, n-propanol, n-butanol, iso-butanol, and sec-butanol. 9. The process according to claim 7, wherein the chlorinated hydrocarbon is selected from the group comprising dichloromethane, dichloroethane, chloroform, and carbon tetrachloride. 10. The process according to claim 6, wherein the reaction mixture of step (a) is stirred for about 5 minutes to about 30 minutes. 11. The process according to claim 6, wherein the alcohol in step (b) is selected from the group comprising methanol, ethanol, n-propanol, n-butanol, iso-butanol, and sec-butanol. 12. The process according to claim 6, wherein the addition of the alcohol is carried out at about 45°C to about 50°C. 13. The stable amorphous tigecycline according to claim 1 having less than 0.5% of a C4 epimer of Formula 11. 14. The stable amorphous tigecycline according to claim 1 having an HPLC purity greater than 99%. 15. A pharmaceutical composition comprising stable amorphous tigecycline and one or more pharmaceutically acceptable carriers, diluents or excipients. 16. Use of stable amorphous tigecycline for the treatment of complicated skin and skin structure infections, complicated intra-abdominal infections, and community-acquired bacterial pneumonia.