WO2017185030A1

WO2017185030A1 - Caspofungin formulation with low impurities

Info

Publication number: WO2017185030A1
Application number: PCT/US2017/028938
Authority: WO
Inventors: John Jiang; Arunya Usayapant; Keith Kwok; Joel Peterson; Jagdish LANDE; Debajit SIRCAR
Original assignee: Fresenius Kabi Usa, Llc
Priority date: 2016-04-22
Filing date: 2017-04-21
Publication date: 2017-10-26

Abstract

Disclosed is a stable pharmaceutical composition comprising caspofungin or a pharmaceutically acceptable salt thereof, arginine, and hydrochloric acid, wherein the pharmaceutical composition further comprises at least one compound selected from the group consisting of (a) a combination of compound J1 and compound J2, (b) compound dimer 1, (c) compound K and (d) any combination thereof. Also disclosed is a method for preparing a stable pharmaceutical composition comprising caspofungin.

Description

CASPOFUNGIN FORMULATION WITH LOW IMPURITIES

BACKGROUND OF THE INVENTION

[0001] A variety of fungal infections can occur in patients due to pathogenic Candida or Aspergillus fungus species. Examples of such fungal infections include candidemia, candidiasis (including esophageal infections, abdominal infections, pleural space infections and peritoneal infections), and invasive aspergillosis.

[0002] Echinocandins are antifungal agents that fungal infections by inhibition of the enzyme that forms -(l,3)-D-glucan, an essential component of the fungal outer cell wall. Because -(l,3)-D-glucan does not occur naturally in the cell walls of mammals, the action of echinocandins is unlikely to be harmful to the cells of an infected patient. In view of this mechanism of action, echinocandins have not experienced wide resistance by target fungi.

[0003] Caspofungin acetate was the first echinocandin to be approved in the United States for use as an antifungal agent. The full name for caspofungin acetate is reported as 1- [(4R,55)-5-[(2-aminoethyl)amino]-N²-(10,12-dimethyl-l-oxotetradecyl)-4-hydroxy-L- ornithine]-5-[(3R)-3-hydroxy-L-ornithine] pneumocandin Bo diacetate (salt), and a representative chemical structure of caspofungin acetate is shown in FIG. 1.

[0004] An approved treatment regimen for adults involves an initial administration of 70 milligrams (mg) caspofungin acetate, followed by daily administration of 50 mg caspofungin acetate, where each administration is performed through intravenous infusion over 1 hour.

[0005] Caspofungin acetate is typically supplied as a lyophilized solid, which is then reconstituted before intravenous administration to a patient. In one example, a formulation of caspofungin acetate that is commercially available is CANCIDAS™ for Injection (Merck & Co, Inc. of Whitehouse Station, New Jersey). CANCIDAS™ is supplied as a lyophilized powder in vials containing either 54.6 mg or 75.6 mg of caspofungin acetate, in combination with sucrose and mannitol, and including acetic acid and sodium hydroxide as pH modifiers.

[0006] CANCIDAS™ is reconstituted for administration by combining the lyophilized powder with 10.8 milliliters (mL) of a reconstitution liquid (such as 0.9% sodium chloride), to provide a solution having a caspofungin acetate concentration of either 7 milligrams per milliliter (mg/mL) or 5 mg/mL. This reconstituted liquid typically is diluted with an infusion liquid prior to administration.

[0007] Commercially available formulations are known to exhibit a lack of stability at ambient temperatures. When stored at room temperature (about 25 °C), degradation products of caspofungin acetate include various dimers of caspofungin as well as other substances. Current protocols require caspofungin acetate to be stored at about -70 °C (±10 °C), and require the lyophilized CANCIDAS™ formulation to be stored at temperatures of about 2 °C to about 8 °C.

[0008] The reconstituted liquid formed from the CANCIDAS™ formulation must be diluted within one hour, and then either administered within 24 hours or stored at 2-8 °C for up to 48 hours. These temperature and time constraints present potential difficulties for effective administration of caspofungin acetate, both in medical professional settings and in outpatient settings. See, e.g., Tsiouris, Maria et al. "Stability and compatibility of reconstituted caspofungin in select elastomeric infusion devices", Int 'l J. ofPharm.

Compounding (2010), 14(5), 436-439.

[0009] Various alternative formulations have been reported in an apparent attempt to improve the stability of caspofungin. The use of the acetate salt form of caspofungin was an initial effort at improving the stability of caspofungin, as earlier versions of caspofungin formulations used the tartrate salt instead. The formulation of CANCIDAS™ containing caspofungin in its acetate salt form has improved stability relative to formulations of caspofungin tartrate. See U.S. Patent No. 5,952,300, col. 2, lines 22-56; col. 8, lines 11-67.

[0010] In another example, the pH modifiers acetic acid and sodium hydroxide were eliminated from the CANCIDAS™ formulation, and the resulting formulations had improved stability at ambient temperatures. Lyophilized formulations were more stable with respect to caspofungin at 25 °C for 12 weeks, and reconstituted solutions of the formulations were more stable with respect to the caspofungin at 25 °C for 2 days. See U.S. Patent Application Publication No. 2009/0170753, paragraphs [0080], [0204] - [0207], [0214] - [0217].

[0011] In a further example, the sucrose and mannitol in the CANCIDAS™ formulation were replaced with a non-reducing sugar such as trehalose, and the resulting formulations had improved stability at ambient temperatures. Lyophilized formulations containing trehalose more were stable with respect to caspofungin at 30 °C for up to 72 weeks, and at 40 °C for up to 24 weeks. See U.S. Patent Application Publication No. 2010/0137197, paragraphs [0114] - [0119].

[0012] Another caspofungin formulation containing arginine was more stable than the conventional mannitol and sucrose composition under various storage conditions. See U.S. Patent Application Publication No. 2014/0142032 Al, paragraphs [0027] - [0030]. [0013] There remains a need for caspofungin formulations that can be stored as lyophilized solids without the need for control of the surrounding temperature below ambient conditions. For example, it is desirable for a lyophilized formulation of caspofungin to be stable at temperatures of about 25 °C or higher for a period of from about 6 months to about 2 years. Preferably such stabilized formulations would be convenient to prepare, store, reconstitute and administer.

BRIEF SUMMARY OF THE INVENTION

[0014] The invention provides a stable pharmaceutical composition comprising a compound of formula (I :

or a pharmaceutically acceptable salt thereof, arginine, and hydrochloric acid, wherein the pharmaceutical composition further comprises a compound selected from the group consisting of :

(a) a combination of compound Jl and compound J2,

(b) compound dimer 1,

(c) compound K, and

(d) combinations thereof.

[0015] The invention further provides a stable pharmaceutical composition comprising a compound of formula (II):

arginine, and hydrochloric acid, wherein the pharmaceutical composition further comprises a compound selected from the group consisting of :

(a) a combination of compound Jl and compound J2,

(b) compound dimer 1,

(c) compound K, and

(d) combinations thereof.

[0016] The invention also provides a method for preparing a stable pharmaceutical composition comprisin mpound of formula (II):

arginine, and hydrochloric acid,

comprising the steps of:

(i) dissolving arginine in water,

(ii) adding hydrochloric acid to achieve a pH of between 6.0 and 6.1,

(iii) dissolving caspofungin acetate in the solution of arginine and hydrochloric acid, (iv) adjusting the pH of the solution obtained in (iii) to between 6.2 and 6.8 using hydrochloric acid and/or sodium hydroxide,

(v) optionally, filtering the solution obtained in step (iv), and

(vi) lyophilizing the solution of step (iv) or step (v) to provide the stable pharmaceutical composition,

which is characterized by the presence of a compound selected from the group consisting of:

(a) a combination of compound Jl and compound J2,

(b) compound dimer 1,

(c) compound K, and

(d) combinations thereof.

BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWING(S)

[0017] Figure 1 depicts the structure of caspofungin acetate process impurity compound A.

[0018] Figure 2 depicts the structure of caspofungin acetate process impurity compound F.

[0019] Figure 3 depicts the structure of caspofungin acetate process impurity compound Co.

[0020] Figure 4 depicts the structure of caspofungin acetate process impurity compound Pneumocandin Bo.

[0021] Figure 5 depicts the structure of caspofungin acetate process impurity compound dimer 2.

[0022] Figure 6 shows the LCMS/MS spectrum of compound Jl .

[0023] Figure 7 shows the LCMS/MS spectrum of compound J2.

[0024] Figure 8 shows the LCMS/MS spectrum of compound K.

[0025] Figure 9 shows a typical HPLC chromatogram of a pharmaceutical composition spiked with 0.10% of compound Bo, in accordance with an embodiment of the invention.

[0026] Figure 10 shows a typical HPLC chromatogram of a pharmaceutical composition in accordance with an embodiment of the invention. DETAILED DESCRIPTION OF THE INVENTION

[0027] The pharmaceutical composition of the invention includes caspofungin in free base form or as pharmaceutically acceptable salt. The compound of formula (I) represents caspofungin in its free base form and is generally referred to as caspofungin.

[0028] In some embodiments, the pharmaceutically acceptable salt is an acid addition salt of an organic acid. In a preferred embodiment, the pharmaceutically acceptable salt is an acetate salt. Caspofungin acetate is a salt of one equivalent of caspofungin and two equivalents of acetic acid, providing an empirical formula of C52H88N10O15 · 2C2H4O2, and a formula weight of 1,213.42 grams per mole.

[0029] The chemical structure shown above for formula (II) represents caspofungin acetate in its associated form. Caspofungin acetate may dissociate from one or both of its equivalents of acetic acid in the presence of a solvent; however, the salt still includes both the cyclic caspofungin and the acetic acid species. Thus, the term "caspofungin acetate" includes both an associated combination of caspofungin and acetic acid, as well as caspofungin and acetic acid in a dissociated state, such as may occur when the salt is combined with a solvent.

[0030] The pharmaceutical composition according to the present invention is stable. The terms "stable" and "stability" are meant to encompass any characteristic of the

pharmaceutical composition which may be affected by storage of the composition at a predetermined temperature for a predetermined time period including, without limitation, potency, total impurities, individual impurities, process related impurities, degradation products, water content, appearance, sterility, and color. An example of a "stable" composition in the context of the present invention is a composition which does not demonstrate an increase beyond a specified amount of one or more degradation compounds following defined storage conditions. Preferred maximum amounts of degradation compounds are described in detail hereinbelow. In some embodiments, the predetermined temperature is about 5 °C, about 25 °C, about 40 °C, or about 60 °C. In other embodiments, the predetermined temperature is 5±2 °C, 25±2 °C, 40±2 °C, or 60±2 °C. The predetermined time period is about one week, about two weeks, about three weeks, about four weeks, about 2 months, about 4 months, about 6 months, about 9 months, about 12 months, about 18 months, about 24 months, about 30 months, about 36 months, or at least about these time periods. [0031] The stability may be affected by storage of the composition at a predetermined temperature at a predetermined relative humidity for a predetermined time period including, without limitation, potency, total impurities, individual impurities, process related impurities, degradation products, water content, appearance, sterility, and color. In some embodiments, the predetermined relative humidity is ambient relative humidity. In other embodiments, the predetermined relative humidity is 40±5% relative humidity, e.g., 45±5% relative humidity, 50±5% relative humidity, 55±5% relative humidity, 60±5% relative humidity, 65±5% relative humidity, 70±5% relative humidity, 75±5% relative humidity, or 80±5% relative humidity.

[0032] In certain embodiments, the pharmaceutical composition is stable for at least 12 months at about 5 °C or 5±2 °C, at ambient relative humidity. In certain embodiments, the pharmaceutical composition is stable for at least 12 months at about 25 °C or 25±2 °C at 60±5% relative humidity. In other embodiments, the pharmaceutical composition is stable for at least 24 months at about 5 °C or 5±2 °C, at ambient relative humidity. In other embodiments, the pharmaceutical composition is stable for at least 24 months at about 25 °C or 25±2 °C at 60±5% relative humidity. In yet other embodiments, the pharmaceutical composition is stable for at least 3 months at about 40 °C or 40±2 °C at 75±5% relative humidity. In still yet other embodiments, the pharmaceutical composition is stable for at least 6 months at about 40 °C or 40±2 °C at 75±5% relative humidity.

[0033] Methods for determining the stability of a pharmaceutical composition of the invention with respect to a given parameter are well-known to those of skill in the art. For example, individual impurities or total impurities, can be assessed by high-performance liquid chromatography (HPLC), thin layer chromatography (TLC), and/or liquid

chromatography-mass spectrometry (LC-MS). Unless otherwise indicated to the contrary, a percentage amount of a compound or an impurity reported herein is relative to the amount of the caspofungin present in the composition as determined by an HPLC method, such as a peak area percent HPLC method, using the following equations.

Individual impurities:

% Impurity = (Ai/(A_Caspofun_gin +∑Ai)) x 100

Total impurities:

%Total Impurities = (∑Ai/(A_Caspofungin +∑Ai)) x 100

where Ai is peak area of each impurity, Acaspofungin is peak area response of caspofungin, and ∑Ai is the sum of peak area responses of all measured impurities [0034] In an embodiment, the mass ratio of arginine to the compound of formula (I) is from about 1 :5 to about 5: 1. In a preferred embodiment, the mass ratio of arginine to the compound of formula (I) is from about 1 : 1 to about 3 : 1, or about 2: 1. In another

embodiment, the mass ratio of arginine to the acetate salt of the compound of formula (I) (i.e., the compound of formula (II) or caspofungin acetate), is about 1 : 1 to about 3 : 1. In a preferred embodiment, the mass ratio of arginine to the acetate salt of the compound of formula (II), (i.e., the compound of formula (II) or caspofungin acetate) is about 2: 1. As used herein, the term "mass ratio" of two substances means the mass of one substance (SI) relative to the mass of the other substance (S2), where both masses have identical units, expressed as S1 :S2

[0035] In an embodiment, an aqueous solution prepared by dissolving the pharmaceutical composition of the invention in water or a pharmaceutically acceptable carrier liquid, e.g., sodium chloride injection, has a pH of about 5 to about 7. In another embodiment, an aqueous solution prepared by dissolving the pharmaceutical composition of the invention in water or a pharmaceutically acceptable carrier liquid has a pH of about 5.5 to about 7.5. In a preferred embodiment, an aqueous solution prepared by dissolving the pharmaceutical composition of the invention in water or a pharmaceutically acceptable carrier liquid has a pH of about 6.2 to about 6.8. In a more preferred embodiment, an aqueous solution prepared by dissolving the pharmaceutical composition of the invention in water or a pharmaceutically acceptable carrier liquid has a pH of about 6.5.

[0036] The pharmaceutical composition further comprises, in addition to the compound of formula (I) or a pharmaceutically acceptable salt thereof and arginine, one of the following compounds: (a) a combination of compound Jl and compound J2, (b) compound dimer 1, (c) compound K, or (d) combinations thereof. Compounds Jl and J2, dimer 1, and compound K are impurities resulting from synthesis of caspofungin, manufacture of the pharmaceutical composition comprising caspofungin, and/or degradation of caspofungin during storage of the pharmaceutical composition.

[0037] As used herein, the compound Jl has the structure:

[0038]

[0039] As used herein, compound dimer 1 has the structure:

(dimer 1)

[0041] The combination of compound Jl and compound J2 may typically be present in the composition according to the invention in an amount of not more than 0.4%, for example, in an amount of from about 0.01%> to about 0.4%, preferably in an amount of from about 0.01%) to about 0.25%), and more preferably in an amount from about 0.1%> to about 0.2%, as measured after storage of the composition at a predetermined temperature for a

predetermined time period, e.g., at about 25 °C for at least about 24 months. [0042] The compound dimer 1 may typically be present in the composition according to the invention in an amount of not more than 0.5%, for example, in an amount from about 0.01%) to about 0.5%), preferably in an amount from about 0.01%> to about 0.25%, and more preferably in an amount from about 0.1%> to about 0.2%, as measured after storage of the composition at a predetermined temperature for a predetermined time period, e.g., at about 25 °C for at least about 24 months, or at about 40 °C for at least about 3 months.

[0043] The compound K may typically be present in the composition according to the invention in an amount of not more than 0.4%, for example, in an amount from about 0.01% to about 0.4%), preferably in an amount from about 0.01%> to about 0.25%, and more preferably in an amount from about 0.1%> to about 0.25%, as measured after storage of the composition at a predetermined temperature for a predetermined time period, e.g., at about 25 °C for at least about 24 months, or at about 40 °C for at least about 3 months.

[0044] In another embodiment, the stable pharmaceutical composition of the invention comprises the compound of formula (I) or (II), arginine, and one or more compounds selected from the group consisting of :

(a) about 0.01% to about 0.4% of a combination of compound Jl and compound

J2,

(b) about 0.01%) to about 0.5% of compound dimer 1,

(c) about 0.01%) to about 0.4% of compound K, and

(d) combinations thereof,

as measured after storage of the composition at a predetermined temperature for a

predetermined time period, e.g., at about 25 °C for at least about 24 months, or at about 40 °C for at least about 3 months.

[0045] In other embodiments, the stable pharmaceutical composition further comprises, in addition to the compound of formula (I) or (II) and arginine, the following combinations of compounds:

(1) about 0.01%) to about 0.4% of a combination of compound Jl and compound J2 and about 0.01% to about 0.5% of compound dimer 1;

(2) about 0.01%) to about 0.4% of a combination of compound Jl and compound J2 and about 0.01% to about 0.4% of compound K;

(3) about 0.01%) to about 0.5% of compound dimer 1 and about 0.01% to about 0.4% of compound K; or (4) about 0.01% to about 0.4% of a combination of compound Jl and compound J2, about 0.01%) to about 0.5% of compound dimer 1, and about 0.01% to about 0.4% of compound K,

[0046] In further embodiments, the stable pharmaceutical composition comprises, in addition to the compound of formula (I) or (II) and arginine, the following combinations of compounds:

(1) about 0.1%) to about 0.2% of a combination of compound Jl and compound J2 and about 0.1%) to about 0.2% of compound dimer 1;

(2) about 0.1%) to about 0.2% of a combination of compound Jl and compound J2 and about 0.1%) to about 0.25% of compound K;

(3) about 0.1%) to about 0.2% of compound dimer 1 and about 0.1% to about 0.25% of compound K; or

(4) about 0.1%) to about 0.2% of a combination of compound Jl and compound J2, about 0.1%) to about 0.2% of compound dimer 1, and about 0.1% to about 0.25% of compound K,

[0047] The pharmaceutical composition of the invention may contain additional impurities which result from degradation during storage of the composition and/or derived from the caspofungin production process. Representative additional impurities include compounds A, B1+B2, F, Co, Pneumocandin Bo, and dimer 2, which have been described in the art (see, e.g., U.S. Patent Application Publications 2009/0170753, 2009/0291996, 2009/0324635, 2010/0256074, 2012/0101030 and 2014/0221274). The structures of compounds A, F, Co, Pneumocandin Bo, and dimer 2 are depicted in Figures 1-5,

respectively.

[0048] The pharmaceutical composition of the invention may be lyophilized or powder filled. Preferably, the pharmaceutical composition is prepared by lyophilization.

[0049] The invention also provides a method for preparing a stable pharmaceutical composition comprising a compound of formula (II):

arginine, and hydrochloric acid,

comprising the steps of :

(i) dissolving arginine in water;

(ii) adding hydrochloric acid to achieve a pH of between 6.0 and 6.1;

(iii) dissolving caspofungin acetate in the solution of arginine and hydrochloric acid;

(iv) adjusting the pH of the solution obtained in (iii) to between 6.2 and 6.8 using hydrochloric acid and/or sodium hydroxide;

(v) optionally, filtering the solution obtained in step (iv); and

(vi) lyophilizing the solution of step (iv) or step (v) to provide the stable

pharmaceutical composition.

[0050] Preferably the method results in a stable pharmaceutical composition which comprises at least one compound selected from the group consisting of: (a) a combination of compound Jl and compound J2; (b) compound dimer 1, (c) compound K; and combinations thereof, most preferably wherein the amount of compounds Jl and J2 together does not exceed 0.4%, the amount of compound dimer 1 does not exceed 0.5%, and/or the amount of compound K does not exceed 0.4%.

[0051] In an embodiment, the arginine is dissolved in water and then hydrochloric acid is added. Preferably, the water is pre-cooled to a temperature in the range of 10 to 12 °C, and all subsequent compounding steps also are performed at a temperature in the range of 10 to 12 °C. Any suitable amount of hydrochloric acid may be added. For example, from about 1 equivalent to about 3 equivalents of hydrochloric acid, preferably 2 equivalents of hydrochloric acid are added. Preferably, the amount of hydrochloric acid added results in the solution having a pH in the range of 6.0 to 6.1. Next, caspofungin acetate is added, and the resulting solution stirred until dissolution is complete. The pH is optionally adjusted if necessary to a final pH in the range of 5.5 to 7.5, preferably 6.2 to 6.8, or more preferably about 6.5, using hydrochloric acid and/or sodium hydroxide. The solution can then be filtered prior to lyophilization.

[0052] The liquid mixture including the solvent, caspofungin acetate, amino acid, and any other optional ingredients may be lyophilized to form a solid composition, such as by subjecting the liquid mixture to freeze-drying. Freeze-drying of the liquid mixture may include maintaining the liquid mixture in an inert atmosphere, such as nitrogen or argon. Preferably the liquid mixture is placed in glass vials prior to lyophilization, and the amount of the liquid mixture in each vial is based on the amount of caspofungin acetate intended to be present in the final solid composition in the vial.

[0053] In a typical lyophilization process, the temperature of the liquid mixture is lowered to a temperature at or below the solidification point of the liquid mixture. If the liquid mixture forms a glass when cooled, the solidification point is the glass transition temperature. If the liquid mixture forms crystals when cooled, the solidification point is the eutectic point. The solidified mixture is then dried under vacuum. Typically, the drying process includes a primary drying step in which the temperature of the solidified mixture is raised gradually while most of the water is removed from the mixture by the vacuum, and a secondary drying step in which the temperature of the solidified mixture is raised further while residual moisture is removed from the mixture by the vacuum. The temperature is kept at or below the desired storage temperature for the final solid composition. Lyophilization typically is complete within 48 hours, but may require additional time. The solid

composition resulting from the lyophilization typically is sealed for later use. Details regarding the lyophilization process may be found, for example, in Remington: The Science and Practice of Pharmacy, 20th Ed., ed. A. Gennaro, Lippincott Williams & Wilkins, 2000.

[0054] In an embodiment, in step (vi), the solution is pre-frozen and subjected to a first drying stage, and then a second drying stage, wherein the first drying stage is carried out at about -37° C for not less than about 120 h and wherein the second drying stage is carried out at about +15° C for not less than about 20 h.

[0055] In a particular embodiment, in step (vi), the lyophilization process comprises the steps of:

(i) a shelf temperature is maintained at +10° C for not less than about 0.5 h, (ii) the shelf temperature is continuously or discontinuously reduced to about -50° C over a period of not less than about 1 h,

(iii) a cold trap is turned on, and the temperature of the cold trap is reduced below

-40° C,

(iv) a vacuum is applied, and the vacuum degree is reduced to about 70 mT,

(v) the shelf temperature is continuously or discontinuously raised to about -37° C over a period of not less than about 2 h,

(vi) the shelf temperature is maintained at about -37° C for not less than about 120 h,

(vii) the shelf temperature is continuously or discontinuously raised to about +15° C over a period of not less than about 4 h, and

(viii) the shelf temperature is maintained at about +15° C for not less than about 20 h.

[0056] The lyophilized solid composition may be stored for later reconstitution and administration. Preferably, the solid composition is stored at a temperature of from 10° C to 40° C, from 15° C to 35° C, from 20° C to 30° C, or about 25° C. Preferably, the solid composition is sealed in a glass vial to protect the composition from moisture in the surrounding environment.

[0057] The invention also provides a stable pharmaceutical composition prepared by any embodiment of the method for preparing a stable pharmaceutical composition as described herein. The stable pharmaceutical composition that is obtained can be characterized, for example, by an amount of compound Jl and compound J2, compound dimer 1, compound K, or combinations thereof, following storage of the composition at a predetermined temperature at a predetermined relative humidity for a predetermined time period. The amounts of compound Jl, J2, dimer 1, K or combinations thereof, methods for detection, and storage conditions for a stable pharmaceutical composition prepared by any embodiment of the method for preparing a stable pharmaceutical composition can be the same as described hereinabove for the stable pharmaceutical composition of the invention.

[0058] For example, the invention provides a stable pharmaceutical composition prepared by a method comprising

(i) dissolving arginine in water;

(ii) adding hydrochloric acid to achieve a pH of between 6.0 and 6.1;

(iv) adjusting the pH of the solution obtained in (iii) to between 6.2 and 6.8 using

hydrochloric acid and/or sodium hydroxide; (v) optionally, filtering the solution obtained in step (iv); and

wherein the stable pharmaceutical composition further comprises one or more compounds selected from the group consisting of :

(a) about 0.01% to about 0.4% of a combination of compound Jl and compound J2,

(b) about 0.01%) to about 0.5% of compound dimer 1,

(c) about 0.01%) to about 0.4% of compound K, and

(d) combinations thereof,

as measured after storage of the composition at a predetermined temperature for a predetermined time period, e.g., at about 25° C for at least about 24 months, or at about 40° C for at least about 3 months.

[0059] The pharmaceutical composition of the invention containing caspofungin acetate, arginine, and optionally one or more other substances may be administered to a patient by combining the composition with an aqueous carrier liquid to form an aqueous mixture, and administering the aqueous mixture into the patient by, for example, injection. Preferably, the aqueous carrier liquid is a pharmaceutically acceptable carrier liquid. Non-limiting examples of pharmaceutically acceptable carrier liquids include water and saline, such as sodium chloride injection, phosphate buffered saline (PBS), Ringers solution or lactated Ringers injection. The aqueous carrier liquid also may include fixed oils, fatty esters or polyols, particularly if the aqueous mixture for injection is a suspension. The aqueous carrier liquid also may include one or more other substances such as buffers, stabilizers, solubilizers, preservatives and antioxidants. Preferably the solid composition dissolves in the aqueous carrier liquid to form a solution.

[0060] Presently preferred aqueous carrier liquids include sodium chloride injection, such as solutions containing 0.9%, 0.45% or 0.225%) sodium chloride. Presently preferred aqueous carrier liquids include sterile water for injection. Presently preferred aqueous carrier liquids include bacteriostatic water for injection, which may include, for example, either 0.9% benzyl alcohol or a combination of methylparaben and propylparaben. Presently preferred aqueous carrier liquids include lactated Ringers injection. Preferably the aqueous carrier liquid does not include dextrose.

[0061] The amount of aqueous carrier liquid may be sufficient to provide an initial aqueous mixture containing caspofungin acetate at a concentration of 5 milligrams per milliliter (mg/mL) or 7 mg/mL. At these concentrations, it is convenient to provide a 50 mg or 70 mg dose of caspofungin acetate to a patient, such as by dispensing 10 milliliters (mL) of the aqueous mixture into another aqueous liquid to form a final aqueous mixture. While an initial aqueous mixture containing caspofungin acetate at a concentration of 5 or 7 mg/mL may be injected into a patient, the presently recommended procedure includes combining the initial aqueous mixture with another aqueous liquid to form a final aqueous mixture, which is then administered to a patient.

[0062] The amount of aqueous carrier liquid may be sufficient to provide a final aqueous mixture containing caspofungin acetate at a concentration of at most 0.5 mg/mL. For example, 10 mL of an initial aqueous mixture containing 5 or 7 mg/mL caspofungin acetate may be combined with 250 mL of an aqueous carrier liquid to provide a final aqueous mixture containing 0.19 mg/mL or 0.27 mg/mL caspofungin acetate (0.192 mg/mL = 50 mg / (250 mL + 10 mL); 0.269 mg/mL = 70 mg / (250 mL + 10 mL)). Presently preferred concentrations of caspofungin acetate in a final aqueous mixture for administration to a patient are from 0.05 to 0.5 mg/mL, from 0.1 to 0.4 mg/mL, and from 0.15 to 0.3 mg/mL. Preferred concentrations of caspofungin acetate in a final aqueous mixture for administration to a patient include 0.19 mg/mL and 0.27 mg/mL.

[0063] An aqueous mixture formed from the solid composition may be administered to provide an initial dose of 50 - 70 mg of caspofungin acetate to a patient. An aqueous mixture formed from the solid composition may be administered to provide a daily dose of 35 - 70 mg of caspofungin acetate to a patient. Doses outside of these ranges also may be

administered. Typically, the initial dose includes 70 mg caspofungin acetate, and subsequent daily doses include 50 mg/mL caspofungin acetate. Daily doses of 70 mg/mL caspofungin acetate may be advisable under certain conditions, such as an insufficient response by the fungal infection and co-administration with other drugs that induce drug clearance (such as rifampin, nevirapine, efavirenz, carbamazepine, dexamethasone and phenytoin). Daily doses below 50 mg/mL of caspofungin acetate may be advisable under certain conditions, such as for pediatric patients or patients having moderate hepatic impairment.

[0064] The invention also provides an isolated compound selected from the group consisting of compound Jl, compound J2, and compound K. Furthermore, the invention provides a composition consisting essentially of, or consisting of, a compound selected from the group consisting of compound Jl, compound J2, and compound K. Compound Jl, compound J2, and compound K could be used, for example, as standards in an impurity detection assay.

[0065] The following examples further illustrate the invention but, of course, should not be construed as in any way limiting its scope.

EXAMPLE 1

[0066] This example demonstrates a preparation of a stable pharmaceutical composition comprising caspofungin acetate and arginine, in accordance with an embodiment of the invention.

[0067] L-Arginine (USP) is added to water in a compounding tank at a temperature of 11° C and stirred until completely dissolved. 6N aqueous HCl is added and the mixture is stirred. The pH of the solution is adjusted to 6.0-6.1 with 10% aqueous HCl or 1.0 N NaOH. Caspofungin acetate is added to the solution and the solution is stirred until the caspofungin acetate is dissolved. The pH is adjusted to a target value of 6.5 with 10% aqueous HCl or 1.0 N NaOH. The solution is then filtered and filled into 10 mL vials. The amounts of

L-Arginine, caspofungin acetate, and 6N aqueous HCl for a 50 mg/vial strength of caspofungin acetate are set forth in Table 1.

Table 1 (Composition)

* based upon caspofungin base

[0068] The vials were then loaded into a lyophilizer and lyophilized according to the lyophilization conditions set forth in Table 2. Table 2 (Lyophilization Conditions)

EXAMPLE 2

[0069] This example demonstrates the amounts of particular impurities present in exemplary compositions containing caspofungin and arginine following storage at 5 ±2° C (ambient humidity) or 25 ±2° C (60 ± 5% relative humidity) for up to 24 months, or at 40 ± 2° C (75 ± 5% relative humidity) for up to six months.

Detection of Compounds A, Pneumocandin Bp, B1+B2, J1+J2, K, Dimer 1, and Dimer 2

[0070] The HPLC conditions were as follows:

[0071] Column: Mac-Mod Analytical Hal peptide ES-C18 (2.7 μιη, 4.6 x 159 mm) or

Supelco Ascentis Express Peptide ES-C-18 (2.7 μιτι, 4.6 x 159 mm) HPLC column

[0072] Mobile Phase A: 100 mM ammonium acetate, pH 5.3

[0073] Mobile Phase B: Mobile Phase A : Acetonitrile (34:66)

[0074] Mobile Phase C: Mobile Phase A : Methanol (28:72)

[0075] Diluent: Methanol

[0076] Column temperature: 26.0 ± 2° C

[0077] Flow rate: l .O mL/min

[0078] Separation mode: Gradient [0079] Gradient program:

[0080] Injection volume: 10 μΐ.

[0081] Autosampler temperature: 5±3° C

[0082] Detection: UV at 225 nm

[0083] Run time: 54 min

[0084] Needle wash: 50:50 WatenAcetonitrile

[0085] The approximate relative response time (RRT) to caspofungin for each impurity was as follows:

Detection of Compounds F and Co

[0086] The HPLC conditions were as follows:

[0087] Column: SeQuant ZIC-HILIC 4.6 x 150 mm, 3.5 μιη [0088] Mobile Phase: 15 mM ammonium acetate, pH 4.0 : acetonitrile (24:76)

[0089] Diluent: 25 mM ammonium dihydrogen phosphate, pH 6 : acetonitrile (50:50)

[0090] Column temperature: 50 ± 3° C

[0091] Flow rate: 1.0 mL/min

[0092] Separation mode: Isocratic

[0093] Injection volume: 10 μΙ_,

[0094] Autosampler temperature: 5±3° C

[0095] Detection: UV at 225 nm

[0096] Run time: 56 min

[0097] Needle wash: Water

[0098] The approximate RRT for compounds F and Co was 0.80 and 1.22, respectively.

[0099] The results are set forth in Tables 3-5. Each reported value is the average (n=6 lots) percentage amount of the impurity as a function of the amount of caspofungin acetate present in the composition following storage at the indicated temperature for the indicated duration as determined by FIPLC. For comparison, the average (n=3-6 lots) percentage amount of each impurity present in vials of the commercial CANCIDAS™ for injection product containing caspofungin, mannitol, and sucrose following storage at 5° C until the labeled expiration date, which was assumed to be approximately 24 months from the manufacture date.

Table 3 (Process-related impurities)

NT = not tested

Table 4 (Degradants)

Table 5 (Accelerated, 40 ±2° C, 75 ±5% RH)

[0100] As is apparent from the results set forth in Tables 4 and 5, the amount of dimer 1 found in the inventive pharmaceutical composition following storage at 25° C at any time period was 0.13% or less, and following storage at 40° C/75% RH at any time period was 0.21% or less. The amount of dimer 1 found in CANCIDAS™ following storage at 5° C until the labeled expiration date was 0.21%.

EXAMPLE 3

[0101] This example demonstrates the structural characterization of compounds Jl, J2, and K by LCMS/MS. [0102] LCMS/MS was performed on a lyophilized composition prepared as described in Example 1 and stored at 40° C for 24 months. The HPLC parameters were essentially as described in Example 1. The HPLC instrument was interfaced with an AB Sciex 5600 Triple TOF mass spectrometer. The mass spectrometer method properties were as follows:

[0103] Duration: 54 min

[0104] Source type: ESI

[0105] Polarity: positive

[0106] Ion source gas 1 : 50.00

[0107] Ion source gas 2: 50.00

[0108] Curtain gas: 30.00

[0109] Two unknown peaks eluted from the caspofungin acetate composition with relative retention times of approximately 1.1 and one unknown peak eluted with a relative retention time of approximately 1.6 (relative to caspofungin acetate). The LCMS/MS spectrum and structure of the first compound, denoted as compound Jl, with relative retention time of approximately 1.1 is shown in Figure 6. The LCMS/MS spectrum and structure of the second compound, denoted as compound J2, with relative retention time of approximately 1.1 is shown in Figure 7. The LCMS/MS spectrum and structure of the third compound, denoted as compound K, with relative retention time of approximately 1.6 is shown in Figure 8.

[0110] All references, including publications, patent applications, and patents, cited herein are hereby incorporated by reference to the same extent as if each reference were individually and specifically indicated to be incorporated by reference and were set forth in its entirety herein.

[0111] The use of the terms "a" and "an" and "the" and "at least one" and similar referents in the context of describing the invention (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. The use of the term "at least one" followed by a list of one or more items (for example, "at least one of A and B") is to be construed to mean one item selected from the listed items (A or B) or any combination of two or more of the listed items (A and B), unless otherwise indicated herein or clearly

contradicted by context. The terms "comprising," "having," "including," and "containing" are to be construed as open-ended terms (i.e., meaning "including, but not limited to,") unless otherwise noted. Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. A composition which is specified by the presence of a compound in a certain range (of percentages) is to be understood as not to comprise additional or lesser amounts of that compound. The upper value of a range is understood to be the maximum amount in said composition, and the lower value of a range is understood to be the minimum amount in said composition. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "such as") provided herein, is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0112] Preferred embodiments of this invention are described herein, including the best mode known to the inventors for carrying out the invention. Variations of those preferred embodiments may become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventors expect skilled artisans to employ such variations as appropriate, and the inventors intend for the invention to be practiced otherwise than as specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

Claims

CLAIM(S):

1. A stable pharmaceutical composition comprising caspofungin or a

pharmaceutically acceptable salt thereof, arginine, and hydrochloric acid, wherein the pharmaceutical composition further comprises at least one compound selected from the group consisting of:

(a) a combination of compound Jl and compound J2,

(b) compound dimer 1,

(c) compound K, and

(d) combinations thereof.

2. The pharmaceutical composition of claim 1, wherein following storage of the pharmaceutical composition at a predetermined temperature for a predetermined time period, the pharmaceutical composition comprises:

(a) not more than 0.4% of a combination of compound Jl and compound J2,

(b) not more than 0.5% of compound dimer 1,

(c) not more than 0.4% of compound K, or

(d) combinations thereof,

as determined by an HPLC peak area percent method.

3. The pharmaceutical composition of claim 2, wherein the predetermined temperature is about 25° C and the predetermined time period is about 24 months.

4. The pharmaceutical composition of any one of claims 1-3, wherein the pharmaceutically acceptable salt is an acetate salt.

5. The pharmaceutical composition of any one of claims 1-4, wherein the composition is lyophilized.

6. The pharmaceutical composition of any one of claims 1-5, wherein the mass ratio of arginine to caspofungin is from about 1 : 1 to about 3 : 1.

7. The pharmaceutical composition of claim 6, wherein the mass ratio of arginine to caspofungin is about 2: 1.

8. The pharmaceutical composition of claim 7, wherein an aqueous solution prepared by dissolving the pharmaceutical composition in water or a pharmaceutically acceptable carrier liquid has a pH of about 6.5.

9. The pharmaceutical composition of any one of claims 1-8, wherein the composition further comprises not more than 0.4% of a combination of compound Jl and compound J2, as determined by an HPLC peak area percent method.

10. The pharmaceutical composition of any one of claims 1-8, wherein the composition further comprises not more than 0.5% of compound dimer 1, as determined by an HPLC peak area percent method.

11. The pharmaceutical composition of any one of claims 1-8, wherein the composition further comprises not more than 0.4% of compound K, as determined by an HPLC peak area percent method.

12. The pharmaceutical composition of any one of claims 1-8, wherein the composition further comprises not more than 0.2% of a combination of compound Jl and compound J2, as determined by an HPLC peak area percent method.

13. The pharmaceutical composition of any one of claims 1-8, wherein the composition further comprises not more than 0.2% of compound dimer 1, as determined by an HPLC peak area percent method.

14. The pharmaceutical composition of any one of claims 1-8, wherein the composition further comprises not more than 0.25% of compound K, as determined by an HPLC peak area percent method.

15. A method for preparing a stable pharmaceutical composition comprising caspofungin acetate, arginine, and hydrochloric acid,

comprising the steps of:

(i) dissolving arginine in water,

(ii) adding hydrochloric acid to achieve a pH of between 6.0 and 6.1,

(iii) dissolving caspofungin acetate in the solution of arginine and hydrochloric acid,

(iv) adjusting the pH of the solution obtained in (iii) to between 6.2 and 6.8 using hydrochloric acid and/or sodium hydroxide,

(v) optionally, filtering the solution obtained in step (iv), and

(vi) lyophilizing the solution of step (iv) or step (v) to provide the stable pharmaceutical composition.

16. The method of claim 15, wherein the stable pharmaceutical composition further comprises a compound selected from the group consisting of:

(a) a combination of compound Jl and compound J2,

(b) compound dimer 1,

(c) compound K, and

(d) combinations thereof.

17. The method of claim 15 or 16, wherein the mass ratio of arginine to caspofungin is from 1 : 1 to 3 : 1.

18. The method of any one of claims 15-17, wherein, in step (vi), the solution is pre-frozen and subjected to a first drying stage, and then a second drying stage, wherein the first drying stage is carried out at about -37° C for not less than about 120 h and wherein the second drying stage is carried out at about +15° C for not less than about 20 h.

19. The method of any one of claims 15-18, wherein, in step (vi), the

lyophilization process comprises the steps of:

(i) a shelf temperature is maintained at +10° C for not less than about 0.5 h,

(ii) the shelf temperature is continuously or discontinuously reduced to about -50° C over a period of not less than about 1 h,

-40° C,

(iv) a vacuum is applied, and the vacuum degree is reduced to about 70 mT,

20. A method of treating or preventing a fungal infection in a mammal in need thereof, comprising administering to the mammal the pharmaceutical composition of claim 1.

21. A stable pharmaceutical composition prepared by the method according to any one of claims 15-19.

22. An isolated compound selected from the group consisting of: 28