Method for producing peptide salts

Abstract

The present disclosure relates to methods for preparing peptide salts, particularly pharmaceutically acceptable peptide salts, such as salts of atosiban (e.g., atosiban pamoate). The disclosure further extends to the preparation of particles of such salts, such as by a spray drying process.

Description

Technical Field

[0001] This disclosure relates to methods for preparing peptide salts, particularly pharmaceutically acceptable peptide salts (such as salts of atosiban). This disclosure further extends to the preparation of particles of such salts, such as by spray drying processes. This disclosure further relates to peptide salts prepared by these methods and particles of such salts. Background Technology

[0002] Atosiban is a synthetic peptide that acts as an inhibitor of oxytocin and vasopressin. Various therapeutic uses of atorvastatin are described in the literature, including as an anti-labor agent (e.g., delaying preterm birth and / or preventing preterm labor). Atosiban contains the following structure:

[0003]

[0004] Atosiban contains a free base and can form acid addition salts. For example, atosiban can form an acid addition salt with acetic acid to provide atosiban acetate. Atosiban acetate is marketed under the trade name... Sold for use in delaying premature birth.

[0005] Dihydroxynaphthyl acid is an organic acid that can be used to form acid addition salts. Its chemical name is 4,4'-methylene-bis(3-hydroxy-2-naphthoic acid). Its structure is shown below:

[0006]

[0007] CN114249800A (Shenzhen Shenyin Pharmaceutical Co., Ltd.) describes a method for preparing the peptide dihydroxynaphthyl salt. This method involves using a chromatographic column with a reversed-phase polymer packing as the stationary phase. The column must be equilibrated before loading the peptide solution, and then an aqueous solution of the dihydroxynaphthyl salt and an organic solvent are loaded onto the column in a specific ratio to allow ion exchange on the column. CN114249800A describes its chromatographic method as providing better control over impurities, ions, and / or solvent residues in the resulting salt. Summary of the Invention

[0008] This disclosure is based on the discovery that atosiban dihydroxynaphthyl acid salt can be prepared by precipitation. Surprisingly, the inventors of this invention have determined that atosiban dihydroxynaphthyl acid produced by the disclosed precipitation method provides a compound with high purity without requiring complex processing and / or purification steps. Such a method is particularly suitable for scaling up manufacturing.

[0009] Therefore, according to the first aspect, a method for preparing atosiban salt of dihydroxynaphthyl acid is provided, the method comprising:

[0010] (i) Contacting and / or reacting the first atosiban salt with the inorganic dihydroxynaphthyl salt.

[0011] Atosiban dihydroxynaphthyl acid forms a precipitate.

[0012] The first atosiban salt can be any suitable atosiban salt that is soluble in a solvent insoluble in atosiban dihydroxynaphthyl acid (e.g., under the temperature and pressure conditions under which the reaction occurs). The first atosiban salt is not atosiban dihydroxynaphthyl acid. In some instances, the first atosiban salt can be a salt with a solubility of at least 10 mg / mL in aqueous solution at 25 °C. As representative examples, the first atosiban salt can be selected from atosiban acetate, atosiban hydrochloride, and atosiban trifluoroacetate, etc.

[0013] For example, the method may include contacting a first atosiban salt (e.g., atosiban acetate) and an inorganic dihydroxynaphthyl salt under conditions that allow them to react. For example, the first atosiban salt (e.g., atosiban acetate) and the inorganic dihydroxynaphthyl salt may contact and / or react in a reaction mixture (e.g., a suspension or solution).

[0014] Specifically, under the conditions disclosed herein, an ion exchange reaction can occur when a first atosiban salt (e.g., atosiban acetate) is contacted and / or reacted with an inorganic bis(hydroxynaphthyl)ate in a reaction mixture (e.g., a solution), subsequently precipitating atosiban bis(hydroxynaphthyl)ate. Surprisingly, the inventors have determined that this precipitation method can provide atosiban bis(hydroxynaphthyl)ate salt product with high purity. Therefore, this method for providing atosiban bis(hydroxynaphthyl)ate offers improved prepareability while maintaining a good level of purity.

[0015] Suitable solvents for forming the reaction mixture (e.g., solution) used in the methods described herein include aqueous solutions. In some instances, the solvent may be water, such as deionized water. As another example, suitable solvents may include aqueous buffer solutions such as sodium phosphate buffer, phosphate-buffered saline (PBS) buffer, Tris buffer (tris(hydroxymethyl)aminomethane buffer), and HEPES (2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid) buffer, etc.

[0016] In some instances, the method may include:

[0017] (i) A solution of a first atosiban salt (e.g., atosiban acetate); and

[0018] (ii) Contacting a solution of the first atosiban salt (e.g., atosiban acetate) with an inorganic dihydroxynaphthyl salt;

[0019] Atosiban dihydroxynaphthyl acid forms a precipitate.

[0020] In some instances, the method further includes preparing a solution of the first atosiban salt, such as a solution of atosiban acetate. For example, a solution of the first atosiban salt can be prepared by adding and / or dissolving the first atosiban salt in any suitable solvent. Suitable solvents for use in preparing the solution of the first atosiban salt include any of those solvents described herein (e.g., aqueous solutions, such as water (e.g., deionized water)). As another example, suitable solvents may include aqueous buffer solutions such as sodium phosphate buffer, phosphate-buffered saline (PBS) buffer, Tris buffer (tris(hydroxymethyl)aminomethane buffer), and HEPES (2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid) buffer, etc.

[0021] The concentration of the first atosiban salt (e.g., atosiban acetate) in the solution can be between about 0.05% w / v and about 50% w / v, about 0.1% w / v and about 20% w / v, about 0.5% w / v and about 10% w / v, or about 1% w / v and about 5% w / v. In some instances, the concentration of the first atosiban salt (e.g., atosiban acetate) in the solution can be between about 1.5% w / v and 4% w / v. In yet another instance, the concentration of the first atosiban salt (e.g., atosiban acetate) in the solution can be about 2.5% w / v or about 3.3% w / v. The concentration mentioned can refer to the concentration of the first atosiban salt (e.g., atosiban acetate) in the initial solution and / or the concentration of the first atosiban salt (atosiban acetate) in the solution after the addition of a metal dihydroxynaphthyl salt (e.g., if the volume increases due to the addition of an inorganic dihydroxynaphthyl salt in solution form).

[0022] Inorganic bis(hydroxynaphthol) salts that can be used in the disclosed methods may include metal bis(hydroxynaphthol) salts. Examples of metal bis(hydroxynaphthol) salts that can be used in the methods disclosed herein include alkali metal bis(hydroxynaphthol) salts and alkaline earth metal bis(hydroxynaphthol) salts. Representative examples include, but are not limited to, sodium bis(hydroxynaphthol) salt and potassium bis(hydroxynaphthol) salt. In a particular example of this disclosure, the metal bis(hydroxynaphthol) salt may be sodium bis(hydroxynaphthol) salt.

[0023] The step of contacting a solution of a first atosiban salt (e.g., atosiban acetate) with an inorganic (e.g., metal) bis(hydroxynaphthyl) salt may include adding the inorganic bis(hydroxynaphthyl) salt to the solution of the first atosiban salt. The inorganic bis(hydroxynaphthyl) salt may be added in the form of a solution or a suspension. For example, a solution of the inorganic bis(hydroxynaphthyl) salt can be prepared by adding and / or dissolving the inorganic bis(hydroxynaphthyl) salt in any suitable solvent.

[0024] Suitable solvents for use in preparing solutions of inorganic dihydroxynaphthyl salts include aqueous solutions. In some instances, the solvent may be water, such as deionized water. As another example, suitable solvents may include aqueous buffer solutions such as sodium phosphate buffer, phosphate-buffered saline (PBS) buffer, Tris buffer (tris(hydroxymethyl)aminomethane buffer), and HEPES (2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid) buffer, etc.

[0025] The concentration of inorganic dihydroxynaphthol in the solution can be between about 0.01% w / v and about 50% w / v, between about 0.05% w / v and about 20% w / v, between about 0.1% w / v and about 10% w / v, or between about 0.25% w / v and about 5% w / v. In some examples, the concentration of inorganic dihydroxynaphthol in the solution can be between about 0.5% w / v and 4% w / v. In yet another example, the concentration of inorganic dihydroxynaphthol in the solution can be about 2.3% w / v or about 0.6% w / v.

[0026] The concentration mentioned can refer to the concentration of inorganic dihydroxynaphthol in the initial solution and / or the concentration of inorganic dihydroxynaphthol in the final solution after the addition of the first atosiban salt.

[0027] The molar ratio of the first atosiban salt (e.g., atosiban acetate) used in the reaction, in contact with and / or in the reaction, to the inorganic dihydroxynaphthyl salt can be any value between about 10:1 and about 1:10, such as between about 5:1 and about 1:5, or between 3:1 and 1:3. In some instances, the molar ratio of the first atosiban salt used in the reaction, in contact with and / or in the reaction, to the inorganic dihydroxynaphthyl salt can be any value between about 2.5:1 and about 1:1, such as between about 2.1:1 and about 1.1:1.

[0028] In some instances, a solution of inorganic dihydroxynaphthyl salt (e.g., an aqueous solution of sodium dihydroxynaphthyl salt) may be added in batches and / or gradually to a solution of the first atosiban salt. As another instance, a solution of inorganic dihydroxynaphthyl salt may be added dropwise to a solution of the first atosiban salt.

[0029] During the addition of inorganic dihydroxynaphthyl salt (which may optionally be added in solution form as described above), the resulting reaction mixture may be stirred (e.g., continuously stirred) to help promote and / or drive the ion exchange reaction.

[0030] This method can be carried out under any conditions suitable for promoting and / or driving ion exchange and / or precipitation of atosiban dihydroxynaphthyl acid from the reaction mixture (e.g., solution). The method can be carried out under ambient conditions. For example, the method can be carried out at room temperature and / or atmospheric pressure.

[0031] In some cases, the method can be performed at temperatures above about 0°C, above about 10°C, or above about 15°C. In some cases, the method can be performed at temperatures between about 0°C and about 100°C. For example, the method can be performed at temperatures between about 10°C and about 50°C, or between about 15°C and about 35°C. In some cases, the method can be performed at about room temperature (e.g., at temperatures between about 18°C ​​and about 27°C, or between about 20°C and about 25°C).

[0032] In some cases, this method can be performed at atmospheric pressure. For example, at pressures within the range of 101.325 kPa or one atmosphere.

[0033] A suitable solvent for the freeze-drying step can be any solvent in which the solubility of atosiban dihydroxynaphthyl acid is at least 20 mg / mL (e.g., at 25°C). Preferably, the melting point of the solvent used for the freeze-drying step can also be at least -80°C. A suitable solvent for the freeze-drying step can include a mixture of acetonitrile and water. As a representative example, the solvent used for the freeze-drying step can be or contains a mixture of acetonitrile and water in a ratio between 1:10 and 10:1 (v / v) (e.g., an acetonitrile / water (1 / 2, v / v) mixture).

[0034] According to another aspect, atosiban dihydroxynaphthyl acid salt prepared according to the methods described herein is provided. In particular, using the methods disclosed herein, the inventors of the present invention have determined that specific salt forms of atosiban dihydroxynaphthyl acid can be obtained. Specifically, in some instances, the atosiban dihydroxynaphthyl acid salt provided by the methods disclosed herein can be a half-salt.

[0035] Therefore, according to another aspect of this disclosure, atosiban salt of dihydroxynaphthyl acid is provided, wherein the molar ratio of atosiban to dihydroxynaphthyl acid salt in the salt is about 2:1.

[0036] The stoichiometry of atosiban dihydroxynaphthyl salt described herein allows for a higher drug loading than the single salt form, which can help reduce the volume of the formulation when the salt is formulated for pharmaceutical use (and this in turn can help increase patient compliance in clinical practice).

[0037] In some instances, it may be desirable to provide atosiban dihydroxynaphthyl acid in granular form (e.g., as granules). For example, injectable formulations often require granules of suitable and / or uniform size. One option for providing such granules is by using spray drying technology.

[0038] Therefore, the method described herein may further include the step of spray drying atosiban dihydroxynaphthyl acid to provide atosiban dihydroxynaphthyl acid granules.

[0039] Therefore, according to another aspect of this disclosure, a method for providing atosiban dihydroxynaphthyl acid granules is provided, the method comprising:

[0040] (i) Provide a loading solution of atosiban dihydroxynaphthyl acid; and

[0041] (ii) The loaded solution is spray-dried to form atosiban dihydroxynaphthyl acid particles.

[0042] As used herein, spray drying can refer to a process in which a liquid is formed into a dry powder (e.g., containing a compound). Spray drying processes typically involve atomizing a solution (referred to herein as a “loaded solution”) by spraying, followed by rapidly evaporating the sprayed droplets into a solid powder by hot gas.

[0043] For a spray drying process to be effective, various factors must be balanced. One such factor is determining a suitable loading solution for the compound to be spray dried. The loading solution should preferably be volatile at a temperature at which the compound in question remains stable. The compound in question should exhibit reasonable solubility in the solvent and also a good level of stability during the spray drying process. This latter point may be particularly important when the compound is to be manufactured on an industrial scale, for example, when the compound may be left in solution for a longer period of time prior to the spray drying step. For compounds intended for use in pharmaceutical formulations (such as atosiban dihydroxynaphthyl acid), the need for stability becomes even more critical, as purity is paramount. Furthermore, the use of such a loading solution in the spray drying process should itself preferably produce particles of suitable and / or uniform size with good appearance and perfusion properties (e.g., when the particles are intended for use in injectable formulations).

[0044] The inventors of this invention encountered difficulties in determining a suitable loading solution for use in the spray drying of atosiban dihydroxynaphthyl acid, specifically a loading solution that would provide at least an acceptable level of solubility while also balancing a good level of stability in the loading solution. In other words, the loading solution would be compatible with atosiban dihydroxynaphthyl acid to provide a repeatable and reliable spray drying process suitable for scaling up manufacturing.

[0045] In particular, the inventors of this invention have identified a set of loading solutions in which atosiban dihydroxynaphthyl acid exhibits good levels of solubility and stability. The loading solutions also exhibit suitable volatility for spray-dried atosiban dihydroxynaphthyl acid and / or can be used to provide particles of suitable and / or uniform size (e.g., for therapeutic purposes).

[0046] Specifically, the solubility of atosiban dihydroxynaphthyl acid in the loading solution can be at least about 15 mg / mL, at least about 20 mg / mL, or at least about 25 mg / mL. In some instances, the solubility of atosiban dihydroxynaphthyl acid in the loading solution can be between about 15 mg / mL and 100 mg / mL, between about 20 mg / mL and 75 mg / mL, or between about 20 mg / mL and 60 mg / mL. As described herein, solubility was measured at room temperature and atmospheric pressure.

[0047] Atosiban dihydroxynaphthyl acid exhibits good stability in loading solutions (e.g., when stored in loading solutions prior to the spray drying step).

[0048] As used herein, the expression “good stability level” may mean that atosiban dihydroxynaphthyl acid present in the loaded solution as described in this disclosure is substantially stable and / or exhibits very low or no degradation.

[0049] Specifically, atosiban dihydroxynaphthyl acid can be substantially stable and / or exhibit very low degradation or no degradation when stored in the disclosed loading solution for a period of at least 1 hour, at least 2 hours, at least 3 hours, or at least 4 hours. In other examples, atosiban dihydroxynaphthyl acid can be substantially stable and / or exhibit very low degradation or no degradation when stored in the disclosed loading solution for a period of at least 12 hours, at least 24 hours, at least 36 hours, or at least 48 hours. In still other examples, atosiban dihydroxynaphthyl acid can be substantially stable and / or exhibit very low degradation or no degradation when stored in the disclosed loading solution for a period of at least 1 day, at least 2 days, at least 3 days, or at least 4 days. In these examples, the loading solution can be stored at room temperature or under refrigeration. Specifically, the loading solution can be stored at room temperature (e.g., at a temperature between about 15°C and about 30°C, such as between about 20°C and about 25°C) and / or at atmospheric pressure.

[0050] As used herein, “stable” can mean that atosiban dihydroxynaphthyl acid, when stored in a loading solution, maintains at least about 98.9%, at least about 99.0%, at least about 99.1%, at least about 99.2%, at least about 99.3%, at least about 99.4%, or at least about 99.5% of its original purity level.

[0051] The loading solution can exhibit a suitable level of volatility for spray-dried atosiban dihydroxynaphthyl acid. In some instances, the loading solution may have a boiling point of less than or equal to about 150°C, less than or equal to about 125°C, or less than or equal to about 100°C.

[0052] The steps of providing a loading solution of atosiban dihydroxynaphthyl acid may include preparing a loading solution of atosiban dihydroxynaphthyl acid. For example, a loading solution of atosiban dihydroxynaphthyl acid may be prepared by adding and / or dissolving atosiban dihydroxynaphthyl acid in any suitable solvent as described herein.

[0053] In some instances, the loading solution may contain one or more solvents, such as two, three, or four solvents. In some cases, the loading solution may contain two solvents.

[0054] One or more solvents may be selected from alcohols, water, and acetonitrile. The alcohol may be saturated or unsaturated. Branched or straight-chain alcohols may be used. Suitable alcohols include, but are not limited to, C1-C6 or C1-C4 alcohols. Representative examples of suitable alcohols include (but are not limited to) methanol, ethanol, isopropanol, n-propanol, isobutanol, sec-butanol, tert-butanol, pentanol, and hexanol.

[0055] In some instances, the loading solution may contain alcohols (such as methanol).

[0056] Specifically, the loading solution may contain alcohol and water. As another example, the loading solution may contain methanol and water, or ethanol and water.

[0057] In other instances, the loading solution may contain acetonitrile and water.

[0058] In particular, in some instances, the loading solution may comprise a solvent system selected from: (i) an alcohol (e.g., a C1-C4 alcohol, such as methanol); (ii) an alcohol (e.g., a C1-C4 alcohol, such as methanol) and water; and (iii) acetonitrile and water. Specifically, the inventors have determined that such a solvent system can be used to provide an effective loading solution of atosiban dihydroxynaphthyl acid to provide good levels of solubility and volatility while maintaining good levels of stability of atosiban dihydroxynaphthyl acid. In yet another instance, the loading solution may comprise a solvent system selected from: (i) an alcohol (such as methanol) and water; and (ii) acetonitrile and water.

[0059] When two or more solvents are present in the loaded solution, they may be present in any suitable ratio that provides a suitable level of volatility, solubility, and / or stability for atosiban dihydroxynaphthyl acid. In some instances, when two solvents are present, they may be present in any ratio (volume of the first solvent:volume of the second solvent) of about 100:1 to about 1:100, about 10:1 to about 1:10, or about 1:5 to about 5:1.

[0060] As another example, when the loading solution contains an alcohol (e.g., a C1-C4 alcohol, such as methanol) and water, the two solvents can be present in any ratio of alcohol (e.g., C1-C4 alcohol, such as methanol) volume to water volume between about 10:1 and about 1:10. In some instances, the loading solution may contain an alcohol (e.g., C1-C4 alcohol, such as methanol) : water (v / v) in any ratio between about 1:1 and about 10:1, or about 2:1 and about 8:1, or about 3:1 and 7:1. In some instances, the loading solution may contain an alcohol (e.g., C1-C4 alcohol, such as methanol) : water (v / v) ratio of approximately 5:1.

[0061] As another representative example, when the loading solution contains acetonitrile and water, the two solvents can be present in any ratio between approximately 10:1 and approximately 1:10 (acetonitrile volume:water volume). In some instances, the loading solution may contain acetonitrile:water (v / v) in any ratio between approximately 1:1 and approximately 10:1, or between approximately 1:1 and approximately 5:1, or between approximately 2:1 and approximately 4:1. In some instances, the loading solution may contain approximately 3:1 acetonitrile:water (v / v).

[0062] As described above, the method includes the step of spray drying the disclosed loading solution to form atosiban dihydroxynaphthyl acid particles (sometimes referred to herein as spray-dried particles of atosiban dihydroxynaphthyl acid).

[0063] The spray drying step can be performed using any spray drying system known in the art.

[0064] Spray dryers typically consist of a drying chamber into which a liquid feed (e.g., a loaded solution as described above) is introduced through an inlet. This inlet is typically in the form of an atomizer. Drying gas is also introduced into the drying chamber, causing the loaded solution to evaporate, leaving solid particles (e.g., solid particles of atosiban dihydroxynaphthyl acid). These solid particles are typically separated and collected in a container using a particle separator (such as a cyclone separator). An overview of the spray drying process in the pharmaceutical industry can be found in Gaspar et al., European Pharmaceutical Review, October 28, 2014, “Spray drying in the pharmaceutical industry”. As a representative example, the spray drying step can be performed using a mini spray dryer, B-290 (Buchi, Switzerland).

[0065] In some instances, during the spray drying step, the inlet temperature (e.g., the temperature of the dry air at the inlet of the drying chamber) can be between about 50°C and about 150°C, between about 75°C and about 125°C, between about 90°C and about 110°C, or between about 95°C and about 105°C. In some instances, the inlet temperature can be about 100°C.

[0066] In some instances, during the spray drying step, the feed rate (e.g., the rate at which the loaded solution is introduced into the nebulizer before entering the drying chamber) can be between about 0.5 mL / min and about 20 mL / min, between about 1 and 10 mL / min, or between about 2 and 7 mL / min. In some instances, the feed rate can be between about 3 mL / min and about 6 mL / min. In yet another instance, the feed rate can be about 4 mL / min.

[0067] In some instances, during the spray drying step, the atomizing gas flow rate (e.g., the rate at which the atomizing gas is introduced into the atomizer) can provide a height reading on the gas flow meter between approximately 10 and approximately 100 mm, approximately 25 mm and 75 mm, or approximately 30 mm and 50 mm. In some instances, the atomizing gas flow rate can be approximately 40 mm. As used herein, “mm” is a unit of height for the reading on the gas flow meter. The height unit can be converted to an actual atomizing gas volumetric flow rate under standard temperature and pressure conditions. For example, a height of 40 mm can be matched with an actual gas volumetric flow rate of 667 L / h. Therefore, in some instances, the atomizing gas flow rate can be between approximately 283 L / h and approximately 1744 L / h, approximately 355 L / h and approximately 1374 L / h, or approximately 439 L / h and approximately 1052 L / h.

[0068] In some instances, during the spray drying step, the dry gas flow rate (e.g., the rate at which the dry gas is introduced into the drying chamber) can be approximately 1 m. 3 / h and approximately 100m 3 Between / h, approximately 10m 3 / h and approximately 75m 3 Between / h, approximately 20m 3 / h and 50m 3 Between / h. In some instances, the dry gas flow rate can be approximately 35m. 3 / h.

[0069] Atosiban dihydroxynaphthyl acid particles produced by spray drying can have a substantially spherical or near-spherical shape. The diameter of spray-dried atosiban dihydroxynaphthyl acid particles produced by the methods described herein (using a loading solution) can be between about 0.1 and about 50 μm (micrometers), such as between about 0.5 μm and about 25 μm, or between about 1 μm and 10 μm. In particular, the diameter of spray-dried atosiban dihydroxynaphthyl acid particles produced by the methods described herein (using a loading solution) can be between about 1 μm and 8 μm. The spray-dried atosiban dihydroxynaphthyl acid particles as described herein can be particularly suitable for use in injectable formulations.

[0070] Following the spray drying step, the purity of atosiban dihydroxynaphthyl acid present in the particles may be at least about 90%, at least about 95%, at least about 97%, or at least about 99% (based on the total weight of the particles).

[0071] In addition to other aspects of this disclosure, bis(hydroxynaphthyl)atosiban granules that can be obtained by the methods disclosed herein are further provided.

[0072] A loading solution is further provided, which contains atosiban dihydroxynaphthyl acid in any of the loading solutions described herein.

[0073] As a particular example, a loading solution for a spray drying process is provided, the loading solution comprising atosiban dihydroxynaphthyl acid and a solvent system selected from: (i) an alcohol (e.g., a C1-C4 alcohol, such as methanol):water (v / v) of approximately 5:1; and (ii) an acetonitrile:water (v / v) of approximately 3:1.

[0074] The methods described herein can be particularly applied to the preparation of atosiban dihydroxynaphthyl acid for use in pharmaceutical compositions. Such pharmaceutical compositions may comprise atosiban dihydroxynaphthyl acid and one or more additional carrier components, such as diluents, excipients, buffers, flavoring agents, binders, surfactants, thickeners, lubricants, preservatives (including antioxidants), etc.

[0075] definition

[0076] In this disclosure, numerous terms are used, and unless the context otherwise indicates, these terms should be understood to have the meanings provided below. The nomenclature used in defining compounds (especially those described herein) is intended to conform to the rules of the International Union of Pure and Applied Chemistry (IUPAC) concerning chemical compounds, particularly the "IUPAC Chemical Terminology Compendium (Golden Book)" (see AD Jenkins et al., Pure & Appl. Chem., 68, 2287-2311 (1996)). For the avoidance of ambiguity, if the IUPAC rules contradict the definitions provided herein, the definitions herein shall prevail.

[0077] As used herein, when specifying numbers or values, the term “about” can refer to a value within ±5% of the specified value. For example, when particles are described as having a diameter of about 50 μm, this includes a range of 47.5 μm to 52.5 μm.

[0078] Furthermore, as used herein, when any range is described as being between a lower limit and an upper limit, the defined range includes the defined endpoints (and therefore includes both the lower limit and the upper limit values).

[0079] Isotope-labeled compounds

[0080] This disclosure also covers various deuterated forms of atosiban dihydroxynaphthyl acid and any other compounds described herein. Each available hydrogen atom attached to a carbon atom can be independently replaced by a deuterium atom. Those skilled in the art will know how to synthesize the deuterated forms of the compounds disclosed herein, including those mentioned above. For example, deuterated materials (such as alkyl groups) can be prepared by conventional techniques (see, for example: methyl-d3-amine, available from Aldrich Chemical Co., Milwaukee, Wisconsin, catalog number 489,689-2).

[0081] This disclosure also includes isotopically labeled compounds of this disclosure, but in fact, one or more atoms are replaced by atoms with atomic masses or mass numbers different from those most commonly found in nature. Examples of isotopes that can be incorporated into the compounds of this disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, fluorine, iodine, and chlorine, such as... 3 H, 11 C 14 C 18 F, 123 I or 125I. Other isotopes of this disclosure containing the aforementioned isotopes and / or other atoms, as well as pharmaceutically acceptable salts of said compounds, are within the scope of this disclosure. Isotope-labeled compounds of this disclosure (e.g., those doped with radioactive isotopes, such as…) 3 H or 14 Those compounds in (C) can be used for drug and / or substrate tissue distribution assays. Deuterated (i.e. 3 H) and carbon-14 (i.e. 14 C) Isotopes are particularly preferred due to their ease of preparation and detectability. 11 C and 18 F isotopes are particularly useful in PET (positron emission tomography).

[0082] It should be noted that throughout the specification, the term "comprising" is used to indicate the features specified in the embodiment of the invention as "comprising," and therefore may also include other features. However, in the context of the invention, the term "comprising" may also cover embodiments of the invention that are "substantially composed of related features" or "composed of related features." Detailed Implementation

[0083] This disclosure will now be further described with reference to the following non-limiting examples.

[0084] Figure 1 Atosiban acetate 1 H NMR spectrum.

[0085] Figure 2 Atosiban dihydroxynaphthyl acid prepared according to the exemplary method disclosed herein 1 H NMR spectrum.

[0086] Figure 3 UPLC determination of atosiban dihydroxynaphthyl acid powder produced by the exemplary method disclosed herein.

[0087] Figure 4 UPLC determination of spray-dried atosiban dihydroxynaphthyl acid particles produced by the exemplary spray drying method disclosed herein.

[0088] Figure 5 Scanning electron microscope (SEM) images of lyophilized atosiban dihydroxynaphthyl acid (left (500×), right (1000×)).

[0089] Figure 6 Scanning electron microscope (SEM) images of spray-dried atosiban dihydroxynaphthyl acid particles (left (3000×), right (5500×)).

[0090] Part A:

[0091] Chemistry - Materials and Methods

[0092] Unless otherwise stated, all chemicals are commercially available and can be used without further purification.

[0093] NMR analysis

[0094] NMR data were acquired using a Bruker Avance NEO 400MHz NMR spectrometer (Bruker, USA). Chemical shifts are reported in ppm relative to dimethyl sulfoxide (δ 2.50) as shown in the NMR spectral data. For sample preparation, small amounts (1–5 mg) of sample (atosiban acetate or atosiban dihydroxynaphthyl acetate) were dissolved in DMSO-d6 (0.6 mL) and transferred to test tubes for NMR analysis. All experiments were performed at 25 °C.

[0095] Ultra-high performance liquid chromatography (UPLC) analysis

[0096] Equipped with Waters Acquity UPLC was performed on a WatersAcquity UPLC H-class system (Waters Corporation, USA) using a BEH phenyl column (1.7 μm, 2.1 x 100 mm). The UPLC parameters are shown below:

[0097]

[0098] The mobile phase gradient of the UPLC is as follows:

[0099]

[0100] Scanning electron microscopy (SEM) analysis:

[0101] SEM in Phenom TM The microscopy was performed using a Pure transmission electron microscope (Thermo Fisher Scientific, USA). For sample preparation, spray-dried powder was directly dispersed onto a carbon binder, and excess powder was removed by blowing a stream of particle-free compressed gas through each sample, followed by gold-platinum coating under high vacuum. SEM images were recorded digitally at higher magnification. Particle size was determined by examining the micrographs using the built-in measuring tools.

[0102] A.1 Preparation of atosiban dihydroxynaphthyl acid

[0103] Atosiban dihydroxynaphthyl acetate was prepared by precipitation. 5 g of atosiban acetate (94.5% free base, MW = 994.5 Da) was dissolved in 150 mL of deionized (DI) water. Separately, 1.15 g of sodium dihydroxynaphthyl acetate (Na₂Pamoate; MW = 432.34 Da, acid to base molar ratio 1.1) was dissolved in 50 mL of deionized (DI) water. The mixture was prepared by adding the atosiban acetate solution dropwise to the sodium dihydroxynaphthyl acetate solution in a 500 mL beaker at room temperature with continuous stirring using a top stirrer. A precipitate formed during continuous stirring, and stirring was continued for another 4 hours at room temperature. The colloidal precipitate was rapidly washed twice with deionized (DI) water and redissolved in an acetonitrile / water (1 / 2, v / v) mixture. Finally, the redissolved atosiban dihydroxynaphthyl acid solution was pre-frozen at -40°C for 24 hours and then placed in a lyophilizer for 72 hours to obtain atosiban dihydroxynaphthyl acid in powder form.

[0104] To confirm the successful synthesis of atosiban dihydroxynaphthyl acid, using 1 The synthesized product was analyzed by ¹H-NMR and UPLC. Equilibrium solubility studies were conducted to verify the change in the solubility of atosiban in aqueous media after hydrophobic counterion exchange.

[0105] pass 1 ¹H NMR (DMSO-d6) determined the stoichiometry of the starting material atosiban:acetate to be approximately 0.93:1, indicating that atosiban acetate is a monoacetate (see [link to original text]). Figure 1 ).

[0106] pass 1 ¹H NMR (DMSO-d6) determined the stoichiometry of atosiban:dihydroxynaphthol to be 1:0.5 (see [reference]). Figure 2 No acetate signal was detected, indicating that atosiban dihydroxynaphthate was formed as a hemihydroxynaphthate.

[0107] Figure 3 UPLC plots of atosiban acetate (top curve) and atosiban dihydroxynaphthyl acetate (bottom curve) are shown. UPLC analysis was performed according to the methods outlined in the "Materials and Methods" section, indicating that the purity of atosiban dihydroxynaphthyl acetate was close to 100%. (Note that in the UPLC plot of atosiban dihydroxynaphthyl acetate, the peak at approximately 2.6 min is the free base of atosiban, while the peak at approximately 6.6 min is the dihydroxynaphthyl acetate. In contrast, the atosiban acetate plot shows only a peak of the free base of atosiban at approximately 2.6 min, as acetic acid did not have a peak under the experimental conditions.)

[0108] A.2 Equilibrium solubility study:

[0109] The results of the equilibrium solubility study of atosiban acetate and atosiban dihydroxynaphthyl acetate in phosphate-buffered saline (PBS) solution are shown in Table 1 below.

[0110]

[0111] Table 1 shows the equilibrium solubility results of atosiban acetate and atosiban dihydroxynaphthyl acetate in phosphate-buffered saline (PBS) solution. S: solubility (mg / mL), calculated using free base.

[0112] At room temperature (approximately 25°C), the approximate solubility of atosiban acetate in PBS solution (pH = 7.4) is greater than 197 mg / mL. Atosiban acetate exhibits high solubility (>60 mg / mL), and the solution became clear after shaking at 37°C for 8 hours and remained clear for 48 hours. In contrast, dihydroxynaphthyl acid salt showed solubility in PBS solution at 37°C for 48 hours (approximately 7.8 mg / mL, average). Furthermore, a gel was observed at the bottom after shaking for 8 hours. These results indicate that the solubility of dihydroxynaphthyl acid salt in PBS is significantly lower than that of atosiban acetate.

[0113] Part B: Spray drying of atosiban dihydroxynaphthyl acid

[0114] Typically, gelation of atosiban dihydroxynaphthyl acid has been observed in certain solvent systems used in spray drying processes, indicating that they are unsuitable for use in the spray drying of atosiban dihydroxynaphthyl acid. The inventors conducted further investigations to determine suitable solvents to provide the loading solution for the spray drying step.

[0115] B1: Initial Solubility Study

[0116] Approximate solubility of atosiban dihydroxynaphthyl acid at room temperature and atmospheric pressure.

[0117] The solubility screening method based on visual inspection is used to measure approximate solubility according to the following procedure:

[0118] 1. Weigh approximately 2 mg of atosiban-dihydroxynaphthyl salt powder into each 2 mL vial;

[0119] 2. For each group, add the test solvent in the following order:

[0120] Step 1: Add 50 μl of solvent to the vial. If the salt dissolves, the solubility data is calculated as (S) > 40 mg / mL (actual S is calculated case by case using the following equation: S = actual weight / 0.05); if the salt does not dissolve, the tester proceeds to Step 2.

[0121] Step 2: Add 50 μl of solvent to the vial. If the salt dissolves, the solubility data is calculated as 40 > S > 20 mg / mL (actual S is calculated for each case); if the salt does not dissolve, the tester proceeds to Step 3.

[0122] Step 3: Add 100 μl of solvent to the vial. If the salt dissolves, the solubility data is calculated as 20 > S > 10 mg / mL (actual S is calculated for each case); if the salt does not dissolve, the tester proceeds to Step 4.

[0123] Step 4: Add 200 μl of solvent to the vial. If the salt dissolves, the solubility data is calculated as 10 > S > 5 mg / mL (actual S is calculated for each case); if the salt does not dissolve, the tester proceeds to Step 5.

[0124] Step 5: Add 200 μl of solvent to the vial. If the salt dissolves, the solubility data is calculated as 5 > S > 3.33 mg / mL (actual S is calculated on a case-by-case basis); if the salt does not dissolve, the tester proceeds to Step 6.

[0125] Step 6: Add 400 μl of solvent to the vial. If the salt dissolves, the solubility data is calculated as 3.33 > S > 2 mg / mL (actual S is calculated for each case); if the salt does not dissolve, then S < 2 mg / mL (actual S should be calculated for each case).

[0126] The results of the initial solubility study in pure volatile solvents are summarized in Table 2 below.

[0127] solvent Solubility (mg / mL) <![CDATA[H2O (water)]]> S<2.4 Acetonitrile S<2.0 EtOAc (ethyl acetate) S<2.1 DCM (dichloromethane) S<2.6 THF (Tetrahydrofuran) S<2.0 acetone S<2.1 MeOH (methanol) 23<S<46 EtOH (ethanol) S<1.9

[0128] Table 2 shows the initial solubility results of atosiban dihydroxynaphthyl acid in pure volatile solvents.

[0129] Limited solubility of atosiban dihydroxynaphthyl acid was observed in almost all tested pure volatile solvents (except methanol). Therefore, the inventors conducted further studies on the solubility of atosiban dihydroxynaphthyl acid in various mixed solvents of water and organic solvents. The results are shown in Table 3 below.

[0130] Solvent (v / v) Solubility (mg / mL) Solvent (v / v) Solubility (mg / mL) <![CDATA[Acetonitrile:H2O (1:2)]]> S>45 <![CDATA[MeOH:H2O(1:2)]]> 2.3<S<3.8 <![CDATA[Acetonitrile:H2O (1:1)]]> S>50 <![CDATA[MeOH:H2O(1:1)]]> 21<S<42 <![CDATA[Acetonitrile:H2O (2:1)]]> S>46 <![CDATA[MeOH:H2O(2:1)]]> S>58 <![CDATA[THF:H2O(1:2)]]> S>54 <![CDATA[EtOH:H2O(1:2)]]> 22<S<43 <![CDATA[THF:H2O(1:1)]]> S>39 <![CDATA[EtOH:H2O(1:1)]]> S>50 <![CDATA[THF:H2O(2:1)]]> S>47 <![CDATA[EtOH:H2O(2:1)]]> 22<S<43 <![CDATA[Acetone:H2O(1:2)]]> S>43 <![CDATA[Acetone:H2O(1:1)]]> S>45 <![CDATA[Acetone:H2O (2:1)]]> S>41

[0131] Table 3 shows the initial solubility results of atosiban dihydroxynaphthyl acid in various mixed solvent systems.

[0132] These initial solubility studies highlight many possible solvent systems that provide good and / or acceptable solubility levels for atosiban dihydroxynaphthyl acid.

[0133] B2: Short-term stability study

[0134] The inventors then further investigated the short-term stability of atosiban dihydroxynaphthyl acid to confirm its compatibility with different solvent systems. Stability studies were conducted at 25°C and atmospheric pressure. After adding atosiban dihydroxynaphthyl acid to the solvent system, UPLC was used to determine the purity of atosiban dihydroxynaphthyl acid at different time points (0 min, 4 h, 24 h, and 4 days). The changes in the peak area ratio of the free atosiban base (atosiban purity) at different time points were summarized to evaluate the short-term stability of atosiban dihydroxynaphthyl acid. (The UPLC measurement methods used in this study are summarized in the "Materials and Methods" section.)

[0135] The results are shown in Table 4.

[0136]

[0137] Table 4 shows the results of short-term stability studies of atosiban dihydroxynaphthyl acid in different solvent systems at 0 h, 4 h, 24 h, and 4 days after its addition to the solvent system.

[0138] Short-term stability data revealed that THF and acetone may lead to the degradation of atosiban peptides under certain conditions, while acetonitrile, MeOH, and EtOH have good compatibility with atosiban-dihydroxynaphthate and are suitable for use in spray drying processes.

[0139] B3: Development of spray drying technology:

[0140] Following an initial study of suitable solvents for use in the loading solution, further parameters of the spray drying process, including solvent, inlet temperature, and feed rate, were investigated. During process development, the atomizing gas flow rate (40 mm) and the drying gas flow rate (35 m³ / s) were also studied. 3 The flow rate ( / h) remains constant. Regarding the atomized gas flow rate, it's important to note that 40mm represents the height reading on the gas flow meter. This figure can be converted to the actual atomized gas volumetric flow rate under standard temperature and pressure conditions. For example, a height of 40mm corresponds to an actual gas volumetric flow rate of 667 L / h.

[0141] Then, several properties of the spray-dried product were analyzed, including purity, content determination, and percolation.

[0142] Specifically, six batches of 300mg-scale spray drying manufacturing processes were conducted. The results are shown in Table 5 below.

[0143]

[0144] Table 5 shows the results of the study on the spray drying process of atosiban dihydroxynaphthyl acid.

[0145] In Table 5, purity, based on the peak area ratio of the free atosiban base (as determined by UPLC analysis), indicates the degree of degradation of the free atosiban base in atosiban dihydroxynaphthyl acid salt. For example, approximately 100% purity indicates that the free atosiban base in atosiban dihydroxynaphthyl acid salt has essentially not degraded, indicating good stability during processing. In Table 5, the percentage data for content determination indicates the content of the free atosiban base in the atosiban dihydroxynaphthyl acid powder. Due to the presence of dihydroxynaphthyl acid in atosiban dihydroxynaphthyl acid, the final percentage content determination for atosiban dihydroxynaphthyl acid particles is approximately 80%.

[0146] B4: Spray-dried atosiban dihydroxynaphthyl acid granules

[0147] B4.1: Method:

[0148] Spray drying was used to prepare atosiban dihydroxynaphthyl acid granules. 1.2 g of lyophilized atosiban dihydroxynaphthyl acid powder was dissolved in a MeOH / H₂O (40 mL, 5:1, v / v) mixed solvent. The solution was filtered through a 0.22 μm filter and spray-dried using a B-290 micro-spray dryer (Buch-Switzerland, Switzerland) at an inlet temperature of 100 °C and a feed rate of 4.0 mL / min. The atomizing gas flow rate and the drying gas flow rate were maintained at 40 mm and 35 m, respectively. 3 / h.

[0149] B4.2: Analysis:

[0150] Spray-dried atosiban dihydroxynaphthyl acid particles, produced according to the method outlined in B4:1, were characterized by ultra-high performance liquid chromatography (UPLC) and scanning electron microscopy (SEM) (according to the method outlined in the "Materials and Methods" section). The results showed... Figure 4 , Figure 5 and Figure 6 middle.

[0151] refer to Figure 4 UPLC analysis of atosiban acetate (top curve) and spray-dried atosiban dihydroxynaphthyl acid particles (bottom curve). UPLC analysis indicated that the purity of the spray-dried atosiban dihydroxynaphthyl acid powder was close to 100%. (Note again that in the UPLC curve of atosiban dihydroxynaphthyl acid, the peak at approximately 2.6 min is the free base of atosiban, while the peak at approximately 6.6 min is the dihydroxynaphthyl acid. In contrast, the atosiban acetate curve only shows a peak of the free base of atosiban at approximately 2.6 min, because acetic acid did not have a peak under the experimental conditions).

[0152] SEM analysis of spray-dried atosiban dihydroxynaphthyl acid particles (manufactured according to the procedure outlined in “B4.1: Method” above) showed that these particles were spherical or nearly spherical in shape, with a diameter of approximately 1 to 8 μm (see [link to article]). Figure 6 (SEM images in [reference]). Such particles are considered suitable for use in injectable formulations. In contrast, SEM analysis of lyophilized atosiban dihydroxynaphthyl acid powder showed that the particles were irregularly flaky in shape and generally larger in size (diameter ranging from 20 μm to 80 μm) (see [reference]). Figure 5 (SEM images from [the source]). These are considered less suitable for use in injectable formulations.

[0153] Although this disclosure has been described in detail by way of illustration and examples for the purpose of clarity, the descriptions and examples should not be construed as limiting the scope of this disclosure. All disclosures of patents and scientific literature cited herein are expressly incorporated herein by reference in their entirety.

Claims

1. A method for preparing atosiban dihydroxynaphthyl acid, the method comprising: (i) Contacting and / or reacting the first atosiban salt and the inorganic dihydroxynaphthyl salt in the reaction mixture. The dihydroxynaphthyl acid atosiban forms a precipitate.

2. The method according to claim 1, wherein the first atosiban salt is an atosiban salt with a solubility of at least 10 mg / mL in an aqueous solution at 25°C, and / or wherein the first atosiban salt is selected from atosiban acetate, atosiban hydrochloride, and atosiban trifluoroacetate.

3. The method according to claim 1 or 2, wherein the inorganic dihydroxynaphthate is a metal dihydroxynaphthate, optionally selected from sodium dihydroxynaphthate and potassium dihydroxynaphthate.

4. The method according to any one of the preceding claims, wherein the reaction mixture is a solution, such as an aqueous solution, optionally water or an aqueous buffer solution.

5. The method according to any one of the preceding claims, further comprising: (i) Provide a solution of the first atosiban salt; as well as (ii) Contact the solution of the first atosiban salt with the inorganic dihydroxynaphthyl salt; And optionally further include: (iii) Adding the inorganic dihydroxynaphthyl salt to the solution of the first atosiban salt, optionally further comprising adding the solution of the inorganic dihydroxynaphthyl salt to the solution of the atosiban salt.

6. The method according to any one of the preceding claims, wherein the molar ratio of the atosiban salt used, contacted and / or reacted to the inorganic dihydroxynaphthyl salt is any molar ratio between about 10:1 and about 1:10, about 5:1 and about 1:5, or about 3:1 and about 1:

3.

7. The method according to any one of the preceding claims, further comprising separating and / or removing the atosiban dihydroxynaphthyl acid precipitate; and optionally (i) Freeze-drying the atosiban dihydroxynaphthyl acid precipitate to form freeze-dried atosiban dihydroxynaphthyl acid; and / or (ii) Spray-dry atosiban dihydroxynaphthyl acid to provide spray-dried atosiban dihydroxynaphthyl acid granules.

8. A method for providing atosiban dihydroxynaphthyl acid granules, the method comprising: (i) Provide a loading solution of atosiban dihydroxynaphthyl acid; as well as (ii) The loaded solution is spray-dried to form atosiban dihydroxynaphthyl acid granules; Optionally, the solubility of the dihydroxynaphthyl atosiban in the loading solution is at least about 15 mg / mL, at least about 20 mg / mL, or at least about 25 mg / mL.

9. The method of claim 8, wherein the dihydroxynaphthyl atosiban is substantially stable and / or exhibits very low degradation or no degradation during storage in the loading solution for a period of at least 1 hour, at least 2 hours, at least 3 hours, at least 4 hours, at least 12 hours, at least 24 hours, at least 36 hours, or at least 48 hours; Optionally, the atosiban dihydroxynaphthyl acid, when stored in the loading solution, maintains a level of original purity of at least about 98.9%, at least about 99.0%, at least about 99.1%, at least about 99.2%, at least about 99.3%, at least about 99.4%, or at least about 99.5%.

10. The method according to claim 8 or 9, wherein the loading solution: (a) Having a boiling point of less than or equal to about 150°C, less than or equal to about 125°C, or less than or equal to about 100°C; and / or (b) Contains one or more solvents, optionally wherein the one or more solvents are selected from alcohols, water, and acetonitrile; and / or (c) Contains a solvent system selected from the following: (i) alcohols (e.g., C1-C4 alcohols, such as methanol); (ii) alcohols (e.g., C1-C4 alcohols, such as methanol) and water; and (iii) acetonitrile and water.

11. The method according to any one of claims 8 to 10, wherein the loading solution comprises a solvent system selected from: (i) an alcohol (e.g., C1-C4 alcohols, such as methanol):water (v / v) in a ratio of approximately 5:1; and (ii) Approximately 3:1 acetonitrile:water (v / v).

12. A loading solution for a spray drying process, the loading solution comprising atosiban dihydroxynaphthyl acid and a solvent system selected from: (i) an alcohol (e.g., C1-C4 alcohols, such as methanol):water (v / v) in a ratio of approximately 5:1; and (ii) Approximately 3:1 acetonitrile:water (v / v).

13. A spray-dried atosiban dihydroxynaphthyl acid granules, which are obtained or can be obtained by the method according to any one of claims 8 to 11, optionally wherein the diameter of the granules is between about 0.1 μm and about 50 μm (micrometers), between about 0.5 μm and about 25 μm, or between about 1 μm and about 10 μm.

14. An atosiban salt of dihydroxynaphthyl acid, wherein: (i) The molar ratio of atosiban to dihydroxynaphthyl salt in the salt is approximately 2:1; and / or (ii) The salt is obtained or is available by the method according to any one of claims 1 to 7.

15. A pharmaceutical composition comprising atosiban dihydroxynaphthyl acid obtained or available by the method according to any one of claims 1 to 7, spray-dried atosiban dihydroxynaphthyl acid granules according to claim 13, or atosiban dihydroxynaphthyl acid salt according to claim 14.

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