WO2023119188A1

WO2023119188A1 - Composition of compound comprising 3-hydroxycyclopentanone moiety and method of its stabilization

Info

Publication number: WO2023119188A1
Application number: PCT/IB2022/062615
Authority: WO
Inventors: Ivar JÄRVING
Original assignee: As Kevelt
Priority date: 2021-12-22
Filing date: 2022-12-21
Publication date: 2023-06-29
Also published as: EE05864B1; EE202100026A

Abstract

The invention relates to a composition comprising a compound having a 3-hydroxycyclopentanone moiety in its structure, an organic solvent comprising an alcohol, an ester or a mixture thereof, and a salt capable of dissociating in the solvent, or its constitutive ions. The invention further relates to a kit comprising such composition, to a method of preparing the composition, and to a method of stabilizing a compound having a 3-hydroxycyclopentanone moiety in the solvent using the salt.

Description

COMPOSITION OF COMPOUND COMPRISING 3-HYDROXYCYCLOPENTANONE MOIETY AND METHOD OF ITS STABILIZATION

Field

The invention relates to the field of chemistry and medicine, in particular it relates to compositions comprising compounds having a 3-hydroxycyclopentanone moiety in their structure, such as prostaglandins, and to methods of stabilization of such compounds.

Background

Significant number of compounds, including those used as pharmaceutical agents, comprise 3- hydroxycyclopentanone ring and have the structure shown below, where -R is variable:

Such compounds, which further will be referred to as principle ingredients (or components), are typically unstable due to the tendency for elimination of a molecule of water via the following reaction:

Examples of such compounds are prostaglandins, in particular lipophilic prostaglandins of E series (e.g. Alprostadil (PGEi) and Dinoprostone (PGE₂)) and D series (e.g. PGD₂).

PGEi and PGE₂ are described in the European Pharmacopoeia as the representatives of the group of prostaglandin analogues used for the treatment of a number of physiological conditions. PGEi and PGE₂ based medicinal products are on the World Health Organization's (WHO) List of Essential Medicines, the most effective and safe medicines needed in a health system. They are administered mainly in parenteral form, while topical and intravaginal formulations are also available on the market. Because of the instability of the 3-hydroxycyclopentanone ring, PGEi and PGE₂ can degrade into prostaglandin Ai (PGAi) and prostaglandin A₂ (PGA₂), respectively, which represent the main degradation products in preparations of PGEi and PGE₂ and the main cause for the loss of quality of the preparations. Formation and accumulation of PGAi and PGA₂ lead to rapid degradation and quality loss of PGEi and PGE₂ based medicines.

Due to unstable nature of the PGEi and PGE₂, medicines containing those compounds are offered exclusively in the form of concentrates, which have to be diluted or reconstituted shortly before use by dissolving in a suitable solvent or solution (e.g. ready-to-use physiological saline solution for infusion). The available concentrates are either solid (e.g., lyophilizate preparations with dextran and hydroxypropyl-beta-cyclodextrin as stabilizers) or non-aqueous solutions (e.g. ethanol-based).

The ethanol-based solutions are easy to manufacture due to simple formulation, manufacturing process and quality control. However, and most importantly, ethanol-based formulations have an advantage of easy administration in case when used as a medicine. The solution can be injected into an infusion solution in one step and does not need reconstitution. The simplified handling results in less errors and decreased risks (e.g., of microbial contamination of an infusion solution due to complex handling). Simple formulation of ethanol-based solutions containing at least one principle ingredient and a solvent is an additional advantage. The simplicity of the formulation results in simple and cost-effective production of such medicines, when compared with manufacturing of lyophilizate form. Marketed ethanol-based concentrates of PGEi- and PGE₂- containing medicines, have shelf life of 2 years when stored at 2 to 8 °C. Concentration of the principle ingredients (PGEi and PGE₂) in formulation of medicines usually is defined in pg/ml and mg/ml scale in a wide range of concentrations (e.g. 20 pg/ml and 1000 mg/ml).

PGD₂ is an important endogenous prostaglandin that has been established recently as a compound for use as or in human medicines. Wider usage of the compound in formulation of human medicines is pending mainly due to its chemical instability in drug formulations.

On the other hand, the ethanol-based solutions containing at least one principle ingredient suffer from insufficient stability of the principle component, due to the tendency for elimination of a molecule of water from the structure of 3-hydroxycyclopentanone moiety as described earlier, which poses numerous threats to provide a high quality product. The instability of the principle ingredient is mainly temperature dependent, therefore stages of manufacturing, handling and storage in terms of temperature control, should be handled with highest care.

Consequently, chemical stability in terms of prevention or reduction of principle ingredient degradation in formulation of human medicinal products containing components with 3- hydroxycyclopentanone moiety in their chemical structure has been chosen as the target for this invention.

Summary

The inventors have aimed at identifying stabilizers to be used in concentrated and diluted solutions of compounds having a 3-hydroxycyclopentanone moiety in order to provide formulations which are stable, easy to use and can be simply and cost-efficiently prepared and analyzed with extended shelf-life. Further, the inventors have aimed at overcoming the limitations of the pharmaceutical use of PGD₂ and to open potential for exploiting health benefits of the molecule.

The objective of the invention is to identify conditions under which 3-hydroxycyclopentanone ring containing substances in a non-aqueous solution have decreased elimination of water from the ring and therefore improved stability and consequently quality (as defined by stable levels of the principle ingredients and degradation products inside solution) over their shelf life. This is achieved by using a small amount of a salt in the solution, capable of dissociating in a solvent and/or constitutive ions of the salt. The use of a salt in a non-aqueous solution of a compound having a 3-hydroxycyclopentanone moiety provides preventing water loss from the moiety.

One particular aim of the invention is to provide a composition comprising a compound having a 3-hydroxycyclopentanone moiety in an organic solvent, where the composition is stable, i.e. is characterized by no or low (minimal) level of degradation products as a result of elimination of water from the 3-hydroxycyclopentanone ring during storing, and by stable level of the principal ingredient, which results in extension of shelf life for such a product. Furthermore, such composition can be easily administered as a ready-to-use formulation by injecting into an infusion solution without the need of reconstitution, which excludes errors (or results in minimal errors) when preparing the composition for administering to a subject and involves no or minimal risks of contamination (e.g., biological (e.g. microbial), chemical) of the composition and ready-to-use formulation for administration.

Another particular target of the invention is to provide a kit comprising said composition, where the kit is characterized by easy handling and easy preparing for administration of the composition to a subject as a ready-to-use formulation and can be stored during long time period without any or significant loss of quality of the principal ingredient. Another particular object of the invention is to provide a simple and cost effective method for preparing said composition, which is easy to carry out and involves minimum number of necessary components.

Another particular object of the invention is to provide a method of stabilizing a compound having a 3-hydroxycyclopentanone moiety in an organic solvent.

The embodiments of the present invention will be described below with reciting technical effects achieved when implementing the invention. Further, upon reading the present description, a person skilled in the art will appreciate other technical effects achieved and other problems solved.

The first aspect of the present invention relates to a composition comprising a compound having a 3-hydroxycyclopentanone moiety, a solvent comprising an alcohol, an ester or a mixture thereof, and a salt capable of dissociating in the solvent, or its constitutive ions.

The composition according to the present invention provides stability of the principal ingredient, which is manifested in the stable content of the principal ingredient. The present composition comprises minimal number of necessary ingredients and, therefore, is characterized by easy and cost-efficient manufacture process and quality control. Furthermore, the composition can be easily administered as a ready-to-use formulation for use as a medicine, e.g. by injecting into an infusion solution in one step and, therefore, does not require reconstitution. Such simplified handling results in no or minimal risks of contamination as well as errors which can occur during formulating.

In one particular embodiment, the solvent further comprises water. In one more particular embodiment, the solvent comprises water in an amount of 0.05 - 10 wt. %, in particular 0.5-4 wt. %, preferably 2-4 wt. %. The water content in said ranges provides further improvement of the stability of the principal ingredient in comparison with the same formulation without such water content.

In another particular embodiment, the alcohol is selected from a monohydric alcohol, a polyhydric alcohol or a mixture thereof, and/or the ester is ethyl acetate. In one more particular embodiment, the monohydric alcohol is ethanol, and/or the polyhydric alcohol is selected from propylene glycol, 1 ,3-butanediol, and a mixture thereof.

In another particular embodiment, the salt has been added into the composition as a compound, or the salt has been produced in situ by adding its precursors into the composition.

In another particular embodiment, the salt is a salt of an alkali metal or an alkaline earth metal. In another particular embodiment, the salt is a halogen salt, in particular, a chloride salt. In another particular embodiment, the salt is sodium chloride, calcium chloride, or magnesium chloride.

In another particular embodiment, the salt is magnesium chloride, in particular, magnesium chloride hexahydrate.

In another particular embodiment, the salt stabilizes the compound having a 3- hydroxycyclopentanone moiety. The stabilization of the compound is achieved by preventing water loss from the compound having a 3-hydroxycyclopentanone moiety.

In another particular embodiment, the composition comprises the salt in a concentration of 0.05- 5000 pg/ml, preferably 0.5-1000 pg/ml, more preferably 0.5-500 pg/ml, in particular 1-30 pg/ml.

In another particular embodiment, the composition comprises the compound having a 3- hydroxycyclopentanone moiety in a concentration of 1-10000 pg/ml, preferably 5-5000 pg/ml, in particular 10-1000 pg/ml.

In another particular embodiment, the weight ratio of the compound to the salt is from 1 :5’000 to 200’000:1 , preferably from 1 :200 to 10’000:1 , more preferably 1 :50 to 2’000:1 , in particular from 1 :50 to 20:1.

In another particular embodiment, the weight (pg) to volume (ml) ratio of the salt to the solvent is from 0.05:1 to 5’000:1 , preferably from 0.5:1 to 1’000:1 , more preferably from 0.5:1 to 500:1 , in particular from 1 :1 to 30:1

In another particular embodiment, the compound having a 3-hydroxycyclopentanone moiety is a prostaglandin, in particular a prostaglandin E or prostaglandin D.

In another particular embodiment, the prostaglandin is prostaglandin Ei (PGEi, Alprostadil), prostaglandin E₂ (PGE₂, Dinoprostone), or prostaglandin D₂ (PGD₂).

In another particular embodiment, the composition is filled into an ampoule, vial, cartridge, prefilled syringe, bottle, or container.

The second aspect of the present invention relates to a kit of parts, comprising the composition of the invention, contained in an ampoule, vial, cartridge, pre-filled syringe, bottle, or container.

The stability of the compound having a 3-hydroxycyclopentanone moiety in the composition according to the present invention allows incorporating the composition into a kit and storing it in the form of a solution, which excludes the necessity of reconstitution before administering to a subject. The kit according to the second aspect provides easy handling, easy administration and long-term storage of the composition of the invention.

The third aspect of the present invention relates to a method of preparing the composition of the invention, the method comprising mixing a compound having a 3-hydroxycyclopentanone moiety, a solvent comprising an alcohol, an ester or a mixture thereof, and a salt capable of dissociating in the solvent, or the salt’s precursors.

The method according to the third aspect allows preparing a composition comprising a compound having a 3-hydroxycyclopentanone moiety in a solvent, where the composition is characterized by stable content of the compound during storage and, thus, no or minimal amount of impurities being products of degradation of the compound. The method of the third aspect allows preparing a composition comprising a compound having a 3-hydroxycyclopentanone moiety, for which stable compositions have not been prepared yet. Further, the method of the third aspect allows preparing stable compositions of a compound having a 3-hydroxycyclopentanone moiety in a solvent in a wide range of concentrations of the compound, i.e. both diluted and concentrated compositions.

The fourth aspect of the present invention relates to a method of stabilizing a compound having a 3-hydroxycyclopentanone moiety, the method comprising contacting the compound, a solvent comprising an alcohol, an ester or a mixture thereof, and a salt capable of dissociating in the solvent, or its constitutive ions.

The method of the fourth aspect provides effective stabilization of the compound having a 3- hydroxycyclopentanone moiety due to the use of a salt acting as a stabilizer for the moiety by preventing elimination of a water molecule from the moiety.

The fifth aspect of the present invention relates to a method of enhancing the chemical stability of a composition comprising a compound having a 3-hydroxycyclopentanone moiety and a solvent, the method comprising adding to the composition at least one salt capable of dissociating in the solvent, or its constitutive ions, wherein the solvent comprises an alcohol, an ester or a mixture thereof.

The sixth aspect of the present invention relates to use of a salt capable of dissociating in a solvent, or its constitutive ions, for stabilizing a compound having at least one 3- hydroxycyclopentanone moiety in the solvent, wherein the solvent comprises an alcohol, an ester or a mixture thereof.

The third to sixth aspects of the present invention can be characterized by the particular embodiments which are similar to the particular embodiments of the first aspect of the present invention.

The essence of the present invention will be illustrated by the following embodiments. Detailed description

The following description presents means and methods which can be used for implementing the present invention as well as provides examples of implementation of the present invention. It is to be noted that the singular forms and the articles “a”, “an” and “the” in the specification and the claims include plural references as well, unless clearly indicated otherwise.

Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the present disclosure belongs, unless it is clearly indicated otherwise. For purposes of the present disclosure, the following terms are used.

For characterizing certain quantitative features, the term “about” is used. This term reflects the uncertainty which is inherent to the measurement of any quantitative feature, and relates to a range which is “a quantitative feature ± a measurement error”. The measurement error can be 10%, more preferably 5%.

The terms “comprise”, “comprises”, “comprising” and “comprised of” are not intended to limit the lists of features following such terms and characterizing, e.g., elements, components, numbers, i.e. such terms do not imply exhaustive lists, while imply the presence of any other additional features characterizing, e.g., additional elements, components, numbers.

All percentages herein characterizing a content, amount, or concentration are indicated as weight percentages (wt. %), unless otherwise indicated or unless the context otherwise requires.

The term “a compound having a 3-hydroxycyclopentanone moiety” relates to a compound comprising a 3-hydroxycyclopentanone moiety in its structure, particularly the compound which is typically unstable due to the tendency to eliminate a molecule of water from the moiety. In the context of the present invention, the stability of such compound is improved due to the addition of a salt, which stabilizes the 3-hydroxycyclopentanone moiety of the compound by preventing water elimination. The general structure of such compounds can be represented by the following formula:

where R can be any substituent. Typical representatives of such compounds are certain prostaglandins, e.g. of E series (e.g. Alprostadil (PGEi) and Dinoprostone (PGE₂)) or D series (e.g. PGD₂):

Prostaglandin D₂ (PGD₂)

The terms "a salt” and “a salt component" in the context of the present invention mean compounds according to the “salt” definition by IUPAC, Compendium of Chemical Terminology, 2nd ed. (the "Gold Book") (1997), i.e. a chemical compound consisting of an ionic assembly of cations and anions (including salts/substances with melting point lower than the room or product storage temperature and substances that can dissociate under conditions within the product providing source of ions).

The terms “stability”, “stable”, “stabilize” and “stabilized” in the context of the present invention imply maintaining the concentration of a principal ingredient in a composition of the invention at a level of an initial concentration (i.e. immediately upon preparation) or slightly decreased, in comparison, in particular, with compositions without a stabilizer. In one embodiment, the concentration of a principal ingredient when stored at 5°C ± 3°C decreases by no more than 6% during a period of 36 months, and/or by no more than 10% during a period of 60 months. For example, the concentration of a principal ingredient when stored at 5°C ± 3°C decreases by no more than 5% during a period of 36 months, preferably by no more than 4% during a period of 36 months, preferably by no more than 3.5% during a period of 36 months, more preferably by no more than 3% during a period of 36 months, and most preferably by no more than 2.5% during a period of 36 months, and/or by no more than 8% during a period of 60 months, more preferably no more than 6% during a period of 60 months, even more preferably no more than 5.5% during a period of 60 months, even more preferably no more than 5.0% during a period of 60 months, and most preferably by no more than 4% during a period of 60 months.

Further provided are non-limiting embodiments of the claimed invention.

The applicant has been manufacturing PGEi and PGE₂ based concentrates, as solutions in anhydrous ethanol, for over 20 years as medicinal products with extensive use as hospital drugs. The solutions according to the invention, in comparison to the prior products, are much more stable with a longer shelf life. In particular, the rate of formation of degradation products in the solution of the invention is significantly reduced in comparison to the prior art. The extent of the effect and the way that the stabilization is achieved meets all the objects of the invention.

The formulations comprising the principle ingredients having a 3-hydroxycyclopentanone moiety (e.g. PGEi and PGE₂) along with a salt according to the present invention, have the shelf life which is significantly prolonged (e.g., from 24 months to at least over 36 months, in particular 60 months, and up to 7-8 years at 5°C ± 3°C storage conditions), which is beneficial in case of using the formulations as medicinal products. For example, the prior art formulations show the decrease of the content of the principle ingredient of 10-15% within 24 months of storage at 5°C ± 3°C, while for the formulations of the present invention, the similar decrease is, e.g., observed after storing for a period more than 60 months at the same storage conditions. The period for handling and storage at near room temperatures without the risk of formation of significant amounts of degradation products is increased (e.g. from under 3 months to 15-18 months at 25°C±2°C). The improvement in stability of the medicinal products based on the composition of the present invention has neither negative effect on the manufacturing process of the products and quality control, nor on the use or safety of the medicine for a user or patient.

It is known that the principle ingredients are very sensitive to bases, acids and oxidizing agents across wide range of concentrations. In particular, bases and acids accelerate the elimination of water from the 3-hydroxycyclopentanone moiety, e.g., from PGEi and PGE₂, to PGAi and PGA₂ forms, respectively. The degradation products are considered to be the main impurities in pharmaceutical preparations containing PGEi and PGE₂, and, therefore, the levels of the degradation products have to be strictly controlled.

In one embodiment of the invention, the composition comprises a salt capable of dissociating in the solvent, or its constitutive ions. Ions of the salt can be both monovalent and polyvalent and can be added as a neutral salt or can be formed in a solution in situ (e.g., via mixing acids and alkalines or bases or any other way, which increases ion concentrations in a solution). Without wishing to be bound by a particular theory, the present inventors believe that a salt used in the composition of the invention acts as an ion donor increasing ionic strength and as a stabilizer for the principle ingredient. In one embodiment of the invention, a salt is used in the concentration of about 0.05-5000 pg/ml, for example about 0.5-1000 pg/ml, preferably about 0.5-500 pg/ml, more preferably about 1-100 pg/ml, in particular about 1-30 pg/ml. When used in pg/ml and mg/ml scale, salts have very significant effect on stabilization of the principle ingredient in the solution, i.e. the salts prevent the ingredient from the degradation involving water elimination from the 3- hydroxycyclopentanone ring.

In one embodiment of the invention, a salt is a salt of an alkali metal or an alkaline earth metal, for example, sodium, potassium, lithium, magnesium, calcium. In one embodiment of the invention, a salt is a halogen salt, for example, a bromide salt, a chloride salt, a fluoride salt, an iodide salt. In one particular embodiment of the invention, a salt is sodium chloride, potassium chloride, lithium chloride, magnesium chloride, calcium chloride. In one more particular embodiment of the invention, a salt is magnesium chloride MgCI₂. In one most particular embodiment, a salt is magnesium chloride hexahydrate MgCI₂ x 6H₂O.

In one particular embodiment, for stabilizing an ethanolic solution comprising a compound having a 3-hydroxycyclopentanone ring in the concentration of 10-1000 pg/ml, a salt is magnesium chloride hexahydrate and it is added in the concentration of 30 pg/ml. Without wishing to be bound by a particular theory, the present inventors believe that the stability of the composition is improved due to stabilizing 3-hydroxycyclopentanone structure of the molecules via preventing water elimination from the ring of the structure.

The stability studies have shown that an improvement in stability (judged by a slow loss of the principle ingredient and/or slow accumulation of the degradation products) can be achieved essentially with all of the salts which fulfil the following conditions: a salt compound shall consist of a cation and an anion and shall be able to dissociate in a solvent to provide ions and, thus, to increase the ionic strength of the solution. The stabilization effect of a salt can be observed both by slow rate of decreasing the content of PGEi, PGE₂ and PGD₂ in the formulations of the invention as well as by a slow rate of increasing the amounts of impurities. In one embodiment of the invention, a solvent is a non-aqueous or organic solvent. In one particular embodiment, a solvent comprises an alcohol, an ester or a mixture thereof. In one more particular embodiment, the alcohol is selected from a monohydric alcohol, a polyhydric alcohol or a mixture thereof. In another more particular embodiment, the ester is ethyl acetate. In one most particular embodiment, the monohydric alcohol is ethanol, and/or the polyhydric alcohol is selected from propylene glycol, 1 ,3-butanediol, and a mixture thereof. In one embodiment of the present invention, when a composition of the invention is intended to be used as a medicinal product, a solvent is a physiologically and/or pharmaceutically acceptable solvent, which is approved for use in medical formulations for humans.

A solvent according to the present invention does not predominantly consist of water. However, small quantities or residual amounts of water up to few percentages may be present. In one particular embodiment of the present invention, the solvent of the present invention further comprises water in an amount of up to 10%, for example, 7.4%, in particular 0.5-4%, preferably 2-4%. The inventors have found that the presence of water in the solvent in an amount of up to 10% could additionally improve stability of a compound comprising 3-hydroxycyclopentanone moiety in an organic solvent in comparison with the same formulation without such water content. In one embodiment of the present invention, ethanol is ethanol 96 vol. % Ph. Eur. (95.1-96.9 vol. %), anhydrous ethanol Ph. Eur. (min 99.5 vol. %), absolute ethanol or any mixture thereof according to the definitions and requirements of the European Pharmacopoeia. Anhydrous ethanol Ph. Eur. (at least 99.5 vol. %) and absolute ethanol are understood as synonyms, often referred to as 100% ethanol or anhydrous ethanol, which in the present case are also synonymous.

In one embodiment of the invention, a compound having a 3-hydroxycyclopentanone moiety is a prostaglandin, in particular a prostaglandin E or prostaglandin D. In one particular embodiment of the invention, the compound is prostaglandin Ei (PGEi, Alprostadil), prostaglandin E₂ (PGE₂, Dinoprostone), or prostaglandin D₂ (PGD₂). PGEi, PGE₂ and PGD₂ are soluble in all the above mentioned solvents in a sufficient concentration and can be used in medicinal formulations due to their beneficial therapeutical effect and suitable toxicological profile. In one embodiment of the invention, a compound comprising a 3-hydroxycyclopentanon moiety is present in the composition in a concentration of 1-10000 pg/ml, preferably 5-5000 pg/ml, in particular 10-1000 pg/ml.

In one embodiment, the composition of the present invention is in the form of a concentrate (concentrated composition, concentrated solution), e.g., comprising the compound having a 3- hydroxycyclopentanon moiety in the concentration of 50-10000 pg/ml, or in the form of ready-to- use compositions, e.g., with the concentration of the principal ingredient of 1-50 pg/ml. In one embodiment of the invention, a composition of the invention is prepared by dissolving at least one principle ingredient in at least one solvent as described above, with addition of at least one salt or other source of ions to act as a stabilizer for the principle ingredient. Thus, in one particular embodiment, the composition of the invention comprises at least one principle ingredient in at least one organic solvent, and at least one source of ions. A source of ions can be added to the composition as a neutral salt or as its constitutive parts added separately, e.g. by mixing an acid and an alkaline or base or any other. In one embodiment, a stabilizer for the principle ingredient and, accordingly, the composition is represented by a source of ions in a solution with no or low water content.

In one embodiment of the invention, the salt is added to the solvent during the preparation process of the liquid formulation before packaging into distribution or market ready primary package (including but not being limited to ampoules, vials, cartridges, syringes, in particular pre-filled syringes, bottles, containers). The principle ingredients are characterized by a decreased rate of water elimination in the absence of oxygen, and, therefore, purging of formulation and over-space with an inert gas is desirable.

The stabilized formulation can be used in the same way as a traditional medicinal product not containing a stabilizer, e.g. by dilution with an infusion solution, by intravaginal or intracervical administration, or topically. The composition can be prepared as a concentrated injection solution for direct administration.

Furthermore, the stabilized formulation can be used as an intermediate step for distribution or storage before any other use (including but not being limited to: chemical modification of the principle compounds in chemical industry or direct/indirect administration to patients, bulk product for repackaging).

All values within the disclosure are descriptive for formulations intended for any mode of administration (including but not limiting to concentrates for solution for infusion). The formulation should not be used directly on patients or in any other application without taking into account safety and following correct procedures.

Examples

Further, the invention will be illustrated by means of some exemplary non-limiting embodiments. The demonstration of the stabilizing effect of the salts onto the tested substances was carried out by means of accelerated stability tests at the prescribed storage temperatures of 25°C and 30°C under non-controlled relative humidity as samples were packaged into type I glass ampoules.

The samples used for accelerated stability studies were prepared with combination of various substances with different solvents, with and without use of different stabilizing components. The stability study was performed for different periods of storage at accelerated storage conditions. Primary package used was 1 ml type I glass ampoule.

Produced samples for the accelerated stability studies were analyzed using high-performance liquid chromatography (HPLC) for measurement of concentration of substances as well as degradation products being a result of degradation of the substances used in formulation of the samples. All results at all time-points for all conditions are the average values from testing samples in triplicate.

Examples 1-3 demonstrate testing of compositions comprising a compound having a 3- hydroxycyclopentanone moiety under GMP conditions for human medicines, prior to manufacturing of validation batches.

Example 1

Preparation of a composition comprising anhydrous ethanol (as a solvent) and 20 pg/ml Alprostadil (PGEi) (reference solution)

Batches of the composition: E10021615, E10021715, E10021815.

Anhydrous ethanol was measured directly into a mixing vessel under inert gas atmosphere. An appropriate amount of Alprostadil was added and the composition was stirred until complete dissolving. The concentration of Alprostadil in the solution was 20 pg/ml. The prepared solution was aseptically filled (argon over-space) into 1 ml type I clear glass ampoules and flame sealed. The amounts of the components used for preparing the reference composition are presented in Table 1.

Example 2

Preparation of a composition comprising 97 vol. % ethanol (as a solvent), 20 pg/ml

Alprostadil and 30 pg/ml MgCI₂ x 6H₂O

Batch of the composition: E102K0118.

Anhydrous ethanol was measured directly into a mixing vessel under inert gas atmosphere. Appropriate amounts of MgCI₂ x 6H₂O and water for injection were added and obtained solutions were stirred to dissolve the salt. An appropriate amount of Alprostadil was added and the compositions were stirred until complete dissolving the components. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 20 pg/ml and 30 pg/ml, respectively. The prepared solution was aseptically filled (argon over-space) into 1 ml type I clear glass ampoules and flame sealed. The amounts of the components used for preparing the composition of Example 2 are presented in Table 2.

Example 3

Preparation of a composition comprising 97 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 100 pg/ml MgCI₂ x 6H₂O

Batch of the composition: Batch E102K0218.

Anhydrous ethanol was measured directly into a mixing vessel under inert gas atmosphere. Appropriate amounts of MgCI₂ x 6H₂O and water for injection were added and obtained solutions were stirred to dissolve the salt. An appropriate amount of Alprostadil was added and the compositions were stirred until complete dissolving. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 20 pg/ml and 100 pg/ml, respectively. The prepared solution was aseptically filled (argon over-space) into 1 ml type I clear glass ampoules and flame sealed. The amounts of the components used for preparing the composition of Example 3 are presented in Table 3.

The results of stability tests of the compositions of Examples 1-3 under accelerated conditions are presented in Tables 4 and 5. The stability tests were carried out at a temperature of 25°C ± 2°C. Table 4 demonstrates the concentration of Alprostadil (pg/ml) in the compositions of Examples 1- 3 immediately after preparation and after storing the compositions during 1 , 2, 3, 4, 6, and 10 months.

Table 5 demonstrates the content of PGAi (the product of degradation of Alprostadil) (%) in the compositions of Examples 1-3 immediately after preparation and after storing the compositions during 1 , 2, 3, 4, 6, and 10 months under accelerated conditions (25°C ± 2°C).

As can be seen from the results of the stability tests of the compositions of Examples 1-3, using a salt in a composition comprising a compound having a 3-hydroxycyclopentanone moiety results in an improved chemical stability of the compound (Alprostadil in Examples 1-3), i.e. the stable content of the compound (the decrease of only up to about 93-94% of the initial concentration) and little accumulation of the degradation product of Alprostadil (up to about 6% only) after storing under accelerated storage conditions during 10 months, compared to the reference compositions (lacking a salt) which after storing for just 6 months show the concentration of Alprostadil of only 72-77% of the initial concentration and PGAi accumulation of up to 22-25%.

Examples 4-14

Examples 4-14 were carried out in order to determine an optimal water content for two different salts under lab conditions.

Example 4

Preparation of the composition comprising anhydrous ethanol (as a solvent), 20 pg/ml Alprostadil and 1000 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of solvent. The two stock solutions were mixed in 1 :1 ratio and the prepared composition was filled into glass ampoules and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 20 pg/ml and 1000 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 4 are presented in Table 6.

Example 5

Preparation of the composition comprising 98 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 1000 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of anhydrous ethanol, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of 96 vol. % ethanol. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 20 pg/ml and 1000 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 5 are presented in Table 7.

Example 6

Preparation of the composition comprising 98 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 100 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of anhydrous ethanol, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of 96 vol. % ethanol. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 20 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 6 are presented in Table 8.

Example 1

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 1000 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 20 pg/ml and 1000 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 7 are presented in Table 9.

Example 8

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 100 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 20 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 8 are presented in Table 10.

Example 9

Preparation of the composition comprising anhydrous ethanol (as a solvent), 20 pg/ml Alprostadil and 1000 pg/ml CaCI₂ x 2H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of CaCI₂ x 2H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and CaCI₂ x 2H₂O in the solution were 20 pg/ml and 1000 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 9 are presented in Table 11.

Example 10

Preparation of the composition comprising 98 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 1000 pg/ml CaCI₂ x 2H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of anhydrous ethanol, and an appropriate amount of CaCI₂ x 2H₂O was dissolved in 14 volume of 96 vol. % ethanol. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and CaCI₂ x 2H₂O in the solution were 20 pg/ml and 1000 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 10 are presented in Table 12.

Example 11

Preparation of the composition comprising 98 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 100 pg/ml CaCI₂ x 2H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of anhydrous ethanol, and an appropriate amount of CaCI₂ x 2H₂O was dissolved in 14 volume of 96 vol. % ethanol. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and CaCI₂ x 2H₂O in the solution were 20 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 11 are presented in Table 13.

Example 12

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 1000 pg/ml CaCI₂ x 2H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of CaCI₂ x 2H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and CaCI₂ x 2H₂O in the solution were 20 pg/ml and 1000 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 12 are presented in Table 14.

Example 13

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 100 pg/ml CaCI₂ x 2H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of CaCI₂ x 2H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and CaCI₂ x 2H₂O in the solution were 20 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 13 are presented in Table 15.

Example 14

Preparation of the composition comprising anhydrous ethanol (as a solvent) and 20 pg/ml Alprostadil (reference solution)

An appropriate amount of Alprostadil was dissolved into an appropriate volume of the solvent. The solution was filled into glass ampoules and flame sealed. The concentration of Alprostadil in the solution was 20 pg/ml. The amounts of the components used for preparing the composition of Example 14 are presented in Table 16.

The results of stability tests of the compositions of Examples 4-14 under accelerated conditions are presented in Table 17. The stability tests were carried out at a temperature of 30°C ±1 °C.

Table 17 demonstrates the content of PGAi (the degradation product of Alprostadil) (%) in the compositions of Examples 4-14 after storing the compositions during 8 and 20 days under accelerated conditions.

The results of the stability tests of the compositions of Examples 4-14 as presented in Table 17 demonstrate that using a salt in a composition comprising a compound having a 3- hydroxycyclopentanone moiety results in an improved chemical stability of the compound (Alprostadil in Examples 4-14) during storing for 8 and 20 days under accelerated conditions, i.e. little accumulation of the product of degradation of Alprostadil (PGAi) of up to 4.7% only, compared to the reference composition (lacking a salt) which after storing for 20 days shows the PGAi accumulation of up to 7.5%.

At the same time, the results of Table 17 show that the PGAi accumulation in the compositions of Alprostadil in ethanol with small amounts of water (up to 4% in Examples 5-8 and 10-11 and 13) is generally lower than in the similar compositions lacking water (i.e. in anhydrous ethanol: Examples 9 and 14). Thus, the results of Table 17 demonstrate that introducing small amounts of water into a composition of a compound having a 3-hydroxycyclopentanone moiety in an organic solvent additionally improves chemical stability of the compound compared to the composition of the compound in an anhydrous solvent.

Further, the data of Table 17 illustrate that two different salts provide effective stabilization of the compound having a 3-hydroxycyclopentanone moiety in an organic solvent.

Examples 15-17

Examples 15-17 demonstrate the use of one more salt (NaCI) as a stabilizer for a compound having a 3-hydroxycyclopentanone moiety in an organic solvent.

Example 15

Preparation of the composition comprising 96 vol. % ethanol (as a solvent) and 20 pg/ml

Alprostadil (reference solution)

An appropriate amount of Alprostadil was dissolved in an appropriate volume of the solvent. The solution was filled into 1 ml type I clear glass ampoules (nitrogen over-space) and flame sealed.

The concentration of Alprostadil in the solution was 20 pg/ml. The amounts of the components used for preparing the composition of Example 15 are presented in Table 18.

Example 16

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 100 pg/ml MgCh x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into 1 ml glass ampoules (nitrogen over-space) and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 20 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 16 are presented in Table 19.

Example 17

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 20 pg/ml Alprostadil and 100 pg/ml NaCI

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of NaCI was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules (nitrogen over-space) and flame sealed. The concentrations of Alprostadil and NaCI in the solution were 20 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 17 are presented in Table 20.

The results of stability tests of the compositions of Examples 15-17 under accelerated conditions are presented in Table 21. The stability tests were carried out at a temperature of 30°C ±1 °C.

Table 21 demonstrates the content of PGAi (the degradation product of Alprostadil) (%) in the compositions of Examples 15-17 immediately after the preparation of the compositions and after storing them during 7 and 14 days under accelerated conditions.

The results of the stability tests of the compositions of Examples 15-17 as presented in Table 21 demonstrate that using a salt in a composition comprising a compound having a 3- hydroxycyclopentanone moiety results in an improved chemical stability of the compound Alprostadil during storing for 7 and 14 days under accelerated conditions, i.e. little accumulation of the product of degradation of Alprostadil (PGAi) of up to 12.2% only, compared to the reference composition (lacking a salt) which after storing for 14 days shows the PGAi accumulation of up to 17%.

Further, the results of Table 21 show that in addition to MgCI₂ x 6H₂O and CaCI₂ x 2H₂O, one more salt, i.e. NaCI, provides efficient stabilization of Alprostadil in ethanol.

Thus, the results of presented examples 1-17 clearly demonstrate that salts of different types are able to stabilize a compound having a 3-hydroxycyclopentanone moiety in an organic solvent.

Examples 18-20

Examples 18-20 demonstrate the use of various concentrations of a salt for stabilizing a compound having a 3-hydroxycyclopentanone moiety (PGEi) in an organic solvent Example 18

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 60 pg/ml Alprostadil and 100 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules (argon over-space) and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 60 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 18 are presented in Table 22.

Example 19

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 60 pg/ml Alprostadil and 30 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 60 pg/ml and 30 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 19 are presented in Table 23.

Example 20

Preparation of the composition comprising 96 vol. % ethanol (as a solvent) and 60 pg/ml Alprostadil (reference solution)

An appropriate amount of Alprostadil was dissolved in an appropriate volume of the solvent. The solution was filled into glass ampoules and flame sealed. The concentration of Alprostadil in the solution was 60 pg/ml. The amounts of the components used for preparing the composition of Example 20 are presented in Table 24.

The results of stability tests of the compositions of Examples 18-20 under accelerated conditions are presented in Table 21. The stability tests were carried out at a temperature of 30°C ±1 °C.

Table 25 demonstrates the content of PGAi (the product of degradation of Alprostadil) (%) in the compositions of Examples 18-20 immediately after the preparation of the compositions and after storing them during 7 and 14 days under accelerated conditions.

The results of the stability tests of the compositions of Examples 18-20 as presented in Table 25 demonstrate that both concentrations of MgCI₂ x 6H₂O, i.e. low 30 pg/ml and high 100 pg/ml, provide effective stabilization of Alprostadil in an organic solvent.

Examples 21-24

Examples 21-24 demonstrate the use of various concentrations of a salt for stabilizing a concentrated solution of a compound having a 3-hydroxycyclopentanone moiety (PGEi) in an organic solvent

Example 21

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 500 pg/ml Alprostadil and 100 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules (argon over-space) and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 500 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 21 are presented in Table 26.

Example 22

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 500 pg/ml Alprostadil and 30 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 500 pg/ml and 30 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 22 are presented in Table 27.

Example 23

Preparation of the composition comprising 96 vol. % ethanol (as a solvent) and 500 pg/ml Alprostadil (reference solution)

An appropriate amount of Alprostadil was dissolved in an appropriate volume of the solvent. The solution was filled into glass ampoules and flame sealed. The concentration of Alprostadil in the solution was 500 pg/ml. The amounts of the components used for preparing the composition of Example 23 are presented in Table 28.

Example 24

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 500 pg/ml Alprostadil and 1000 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Alprostadil was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules and flame sealed. The concentrations of Alprostadil and MgCI₂ x 6H₂O in the solution were 500 pg/ml and 1000 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 24 are presented in Table 29.

The results of the stability tests of the compositions of Examples 21-24 under accelerated conditions are presented in Table 30. The stability tests were carried out at a temperature of 30°C ±1 °C.

Table 30 demonstrates the content of PGAi (the degradation product of Alprostadil) (%) in the compositions of Examples 21-24 immediately after the preparation of the compositions and after storing them during 7 and 14 days under accelerated conditions.

The results of the stability tests of the compositions of Examples 21-24 as presented in Table 30 demonstrate that various concentrations of MgCL x 6H2O, i.e. from low 30 pg/ml to high 1000 pg/ml, provide effective stabilization of the concentrated solution of Alprostadil in an organic solvent.

Examples 25-27

Examples 25-27 demonstrate the use of various concentrations of a salt for stabilizing a concentrated solution of a compound having a 3-hydroxycyclopentanone moiety (PGE2) in an organic solvent. Example 25

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 1000 pg/ml Dinoprostone (PGE₂) and 1000 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Dinoprostone was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules (nitrogen over-space) and flame sealed. The concentration of both Dinoprostone and MgCI₂ x 6H₂O in the solution were 1000 pg/ml. The amounts of the components used for preparing the composition of Example 25 are presented in Table 31.

Example 26

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 1000 pg/ml Dinoprostone and 100 pg/ml MgCI₂ x 6H₂O

An appropriate amount of Dinoprostone was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules (nitrogen over-space) and flame sealed. The concentrations of Dinoprostone and MgCI₂ x 6H₂O in the solution were 1000 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 26 are presented in Table 32.

Example 27

Preparation of the composition comprising 96 vol. % ethanol (as a solvent) and 1000 pg/ml Dinoprostone (reference solution)

An appropriate amount of Dinoprostone was dissolved in an appropriate volume of the solvent. The solution was filled into glass ampoules (nitrogen over-space) and flame sealed. The concentration of Dinoprostone in the solution was 1000 pg/ml. The amounts of the components used for preparing the composition of Example 27 are presented in Table 33.

The results of the stability tests of the compositions of Examples 25-27 under accelerated conditions are presented in Tables 34 and 35. The stability tests were carried out at a temperature of 30°C ±1 °C.

Table 34 demonstrates the concentration of Dinoprostone (pg/ml) in the compositions of Examples 25-27 immediately after preparation and after storing the compositions during 7 and 14 days under accelerated conditions.

the compositions of Examples 25-27 immediately after the preparation of the compositions and after storing them during 7 and 14 days under accelerated conditions.

The results of the stability tests of the compositions of Examples 25-27 as presented in Tables 34 and 35 demonstrate that various concentrations of MgCI₂ x 6H₂O, i.e. from low 100 pg/ml to high 1000 pg/ml, provide effective stabilization of the concentrated solution of a compound having a 3-hydroxycyclopentanone moiety, i.e. Dinoprostone (PGE₂), in an organic solvent.

Examples 28-29

Examples 28-29 demonstrate the use of a salt for stabilizing a diluted solution of a compound having a 3-hydroxycyclopentanone moiety (PGD₂) in an organic solvent.

Example 28

Preparation of the composition comprising 96 vol. % ethanol (as a solvent) and 10 pg/ml PGD₂ (reference solution)

An appropriate amount of PGD₂ was dissolved in an appropriate volume of the solvent. The solution was filled into glass ampoules (nitrogen over-space) and flame sealed. The concentration of PGD₂ in the solution was 10 pg/ml. The amounts of the components used for preparing the composition of Example 28 are presented in Table 36.

Example 29

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 10 pg/ml

PGD₂ and 100 pg/ml MgCI₂ x 6H₂O

An appropriate amount of PGD₂ was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules (nitrogen over-space) and flame sealed. The concentrations of PGD₂ and MgCI₂ x 6H₂O in the solution were 10 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 29 are presented in Table 37.

The results of the stability tests of the compositions of Examples 28-29 under accelerated conditions are presented in Table 38. The stability tests were carried out at a temperature of 30°C ±1 °C.

Unlike the results for Examples 1-27, the results for Examples 28-29 are presented as relative values (%) based on the area under the curve from the detector.

Table 38 demonstrates the concentration of PGD₂ (pg/ml) in the compositions of Examples 28- 29 immediately after preparation and after storing the compositions during 7 and 14 days under accelerated conditions.

The results of the stability tests of the compositions of Examples 28-29 as presented in Table 38 demonstrate that the use of the salt MgCI₂ x 6H2O provides effective stabilization of the diluted solution of a compound having a 3-hydroxycyclopentanone moiety, i.e. PGD₂, in an organic solvent, while the similar solution without the salt is very unstable due to the degradation of PGD₂.

Examples 30-31

Examples 30-31 demonstrate the use of a salt for stabilizing a diluted solution of a compound having a 3-hydroxycyclopentanone moiety (PGD₂) in an organic solvent.

Example 30

Preparation of the composition comprising 96 vol. % ethanol (as a solvent) and 20 pg/ml PGD₂ (reference solution)

An appropriate amount of PGD₂ was dissolved in an appropriate volume of the solvent. The solution was filled into glass ampoules (nitrogen over-space) and flame sealed. The concentration of PGD₂ in the solution was 20 pg/ml. The amounts of the components used for preparing the composition of Example 30 are presented in Table 39.

Example 31

Preparation of the composition comprising 96 vol. % ethanol (as a solvent), 16 pg/ml

PGD₂ and 100 pg/ml MgCI₂ x 6H₂O

An appropriate amount of PGD₂ was dissolved in 14 volume of the solvent, and an appropriate amount of MgCI₂ x 6H₂O was dissolved in 14 volume of the solvent. The two stock solutions were mixed in 1 :1 ratio and filled into glass ampoules (nitrogen over-space) and flame sealed. The concentrations of PGD₂ and MgCI₂ x 6H₂O in the solution were 16 pg/ml and 100 pg/ml, respectively. The amounts of the components used for preparing the composition of Example 31 are presented in Table 40.

The results of the stability tests of the compositions of Examples 30-31 under accelerated conditions are presented in Table 41 . The stability tests were carried out at a temperature of 30°C ±1 °C.

Unlike the results for Examples 1-27, the results for Examples 30-31 are presented as relative values (%) based on the area under the curve from the detector.

Table 41 demonstrates the concentration of PGD₂ (pg/ml) in the compositions of Examples SO- 31 immediately after preparation and after storing the compositions during 7 and 14 days under accelerated conditions.

The results of the stability tests of the compositions of Examples 30-31 as presented in Table 41 demonstrate that the use of the salt MgCI₂ x 6H₂O provides effective stabilization of the diluted solution of a compound having a 3-hydroxycyclopentanone moiety, i.e. PGD₂, in an organic solvent compared to the reference solution lacking the salt, which is very unstable due to the degradation of PGD₂.

Example 32

Long term stability study of the compositions of the invention

3 batches of the formulation according to the present invention in primary packaging (1 ml glass ampoules) were manufactured in accordance with the procedure described in Example 2 above. Results from the long term stability study of those (storage at 5 ± 3 °C) during 18 months of storage is presented in a table below. Table 42 demonstrates the content of PGAi (the degradation product of Alprostadil) (%) in the compositions of the invention immediately after the preparation of the compositions and after storing them during 3, 6, 9, 12 and 18 months.

The results of the stability tests of the compositions of the inventions as presented in Table 42 demonstrate that the compositions of the invention exhibit excellent long term stability at 5 ± 3 °C, i.e. show the low amounts of the degradation product of Alprostadil during storing the compositions for a long period, in particular 18 months and more.

In the disclosure, only the selected results of the tests are reported, that represent main proves and trends that are also reproducible under other tested combinations. Some of tested combinations are part of commercially important formulations used as human medicines with well documented stability profile and correlation between stabilities at different tested temperatures. Stability studies have shown that an improvement in stability (judged by a slow loss of the principle ingredient and/or slow accumulation of the degradation products) can be achieved essentially with all of the salts which fulfil the following conditions specified above, i.e. a compound shall consist of a cation and an anion and shall be able to dissociate in a solvent to provide ions and, thus, to increase the ionic strength of the solution. The stabilization effect can be observed both from slow rate of decreasing the content of the principle ingredients (PGEi, PGE2 and PGD2) in the formulations of the invention as well as from a slow rate of increasing the amounts of impurities being the degradation products of the principle ingredients.

All tested concentrations had a positive stabilizing effect on all tested principle ingredients at all concentrations, which means that beneficial stabilizing effect is likely to occur at combinations of concentrations beyond tested.

Claims

1. A composition comprising a compound having a 3-hydroxycyclopentanone moiety, a solvent comprising an alcohol, an ester or a mixture thereof, and a salt capable of dissociating in the solvent, or its constitutive ions.

2. The composition according to claim 1 , characterized in that the solvent further comprises water.

3. The composition according to claim 2, characterized in that the solvent comprises water in an amount of 0.05-10 wt. %, in particular 0.5-4 wt. %, preferably 2-4 wt. %.

4. The composition according to any one of claims 1 to 3, characterized in that the alcohol is selected from a monohydric alcohol, a polyhydric alcohol or a mixture thereof, and/or the ester is ethyl acetate.

5. The composition according to claim 4, characterized in that the monohydric alcohol is ethanol, and/or the polyhydric alcohol is selected from propylene glycol, 1 ,3-butanediol, and a mixture thereof.

6. The composition according to any one of claims 1 to 5, characterized in that the salt has been added into the composition as a compound, or the salt has been produced in situ by adding its precursors into the composition.

7. The composition according to any one of claims 1 to 6, characterized in that the salt is a salt of an alkali metal or an alkaline earth metal.

8. The composition according to any one of claims 1 to 7, characterized in that the salt is a halogen salt, in particular, a chloride salt.

9. The composition according to any one of claims 1 to 8, characterized in that the salt is sodium chloride, calcium chloride, or magnesium chloride.

10. The composition according to any one of claims 1 to 9, characterized in that the salt is magnesium chloride, in particular, magnesium chloride hexahydrate.

11. The composition according to any one of claims 1 to 10, characterized in that the salt stabilizes the compound having a 3-hydroxycyclopentanone moiety.

12. The composition according to any one of claims 1 to 11 , comprising the salt in a concentration of 0.05-5000 pg/ml, preferably 0.5-1000 pg/ml, more preferably 0.5-500 pg/ml, in particular 1-30 pg/ml.

13. The composition according to any one of claims 1 to 12, comprising the compound having a 3-hydroxycyclopentanone moiety in a concentration of 1-10000 pg/ml, preferably 5-5000 pg/ml, in particular 10-1000 pg/ml.

37

14. The composition according to any one of claims 1 to 13, characterized in that the compound having a 3-hydroxycyclopentanone moiety is a prostaglandin, in particular a prostaglandin E or prostaglandin D.

15. The composition according to claim 14, characterized in that the prostaglandin is prostaglandin Ei (PGEi, Alprostadil), prostaglandin E₂ (PGE₂, Dinoprostone), or prostaglandin D₂ (PGD₂).

16. The composition according to any one of claims 1 to 15, characterized in that it is filled into an ampoule, vial, cartridge, pre-filled syringe, bottle, or container.

17. A kit of parts, comprising the composition according to any one of claims 1 to 16, contained in an ampoule, vial, cartridge, pre-filled syringe, bottle, or container.

18. A method of preparing the composition according to any one of claims 1 to 16, comprising mixing a compound having a 3-hydroxycyclopentanone moiety, a solvent comprising an alcohol, an ester or a mixture thereof, and a salt capable of dissociating in the solvent, or the salt’s precursors.

19. A method of stabilizing a compound having a 3-hydroxycyclopentanone moiety, the method comprising contacting the compound, a solvent comprising an alcohol, an ester or a mixture thereof, and a salt capable of dissociating in the solvent, or its constitutive ions.

20. The method according to claim 18 or 19, characterized in that the solvent further comprises water.

21. The method according to claim 20, characterized in that the solvent comprises water in an amount of 0.05-10 wt. %, in particular 0.5-4 wt. %, preferably 2-4 wt. %.

22. The method according to any one of claims 18 to 21 , characterized in that the alcohol is selected from a monohydric alcohol, a polyhydric alcohol or a mixture thereof, and/or the ester is ethyl acetate.

23. The method according to any one of claims 18 to 22, characterized in that the monohydric alcohol is ethanol, and/or the polyhydric alcohol is selected from propylene glycol, 1 ,3-butanediol, and a mixture thereof.

24. The method according to any one of claims 18 to 23, characterized in that the salt is a salt of an alkali metal or an alkaline earth metal.

25. The method according to any one of claims 18 to 24, characterized in that the salt is a halogen salt, in particular, a chloride salt.

26. The method according to any one of claims 18 to 25, characterized in that the salt is sodium chloride, calcium chloride, or magnesium chloride.

38

27. The method according to any one of claims 18 to 26, characterized in that the salt is magnesium chloride, in particular, magnesium chloride hexahydrate.

28. The method according to any one of claims 18 to 27, characterized in that the weight ratio of the compound to the salt or its constitutive ions is from 1 :5’000 to 200’000:1 , preferably from 1 :200 to 10’000:1 , more preferably 1 :50 to 2’000:1 , in particular from 1 :50 to 20:1.

29. The method according to any one of claims 18 to 28, characterized in that the weight (pg) to volume (ml) ratio of the salt or its constitutive ions to the solvent is from 0.05:1 to 5’000:1 , preferably from 0.5:1 to 1’000:1 , more preferably from 0.5:1 to 500:1 , in particular from 1 :1 to 30:1.

30. The method according to any one of claims 18 to 29, characterized in that the compound having a 3-hydroxycyclopentanone moiety is a prostaglandin, in particular a prostaglandin E or prostaglandin D.

31. The method according to claim 30, characterized in that the prostaglandin is prostaglandin Ei (PGEi, Alprostadil), prostaglandin E₂ (PGE₂, Dinoprostone), or prostaglandin D₂ (PGD₂).