Composition for treating osteoarthropathy

Abstract

The present invention relates to a composition for intra-articular use in the treatment of osteoarthropathy (OA) comprising cross-linked hyaluronic acid or a salt or derivative thereof, clodronate or an equivalent salt thereof, and an acetate buffer.

Description

Technical Field

[0001] This invention relates to a composition, preferably for treating osteoarthritis (OA) by intra-articular application, including osteoarthritis, the composition comprising cross-linked hyaluronic acid or its salt or derivative, clodronic acid or an equal salt thereof, and acetate buffer. Background Technology

[0002] Osteoarthritis (OA) is a progressive, chronic disease that particularly affects joints most susceptible to mechanical stress, such as the hip and knee. When the condition develops, the entire joint is affected by a series of degradation and repair processes that ultimately alter the joint's anatomy and function, affecting all joint components, such as cartilage, subchondral bone, and synovial tissue. Therefore, arthritis is a result of a complex interplay between systemic factors (such as advanced age or obesity) and local factors (such as trauma or overuse), which are modulated by numerous predisposing factors and may combine with infectious and inflammatory events of various etiologies.

[0003] In such complex events, over time, people have attempted therapeutic interventions by addressing and counteracting individual causal processes, resorting to a variety of different treatment methods that are largely oriented towards managing inflammatory events and the resulting pain. These methods primarily use analgesics, local or systemic anti-inflammatory drugs, or intra-articular injections of various medications such as bisphosphonates, hyaluronic acid, anthraquinones, and chondroitin sulfate.

[0004] In order to obtain a more effective composition, combinations of the above-mentioned active ingredients were also tried.

[0005] For example, patent EP1976538, owned by the same applicant, describes a combination of linear hyaluronic acid and sodium clodronate, which has achieved excellent results in relieving pain symptoms in patients with osteoarthritis (OA). Furthermore, due to the synergistic effect of the two active ingredients, this combination allows for a reduction in the effective dosage of hyaluronic acid in the final formulation, thereby reducing its adverse effects (such as stimulating the production of inflammatory cytokines), and a reduction in the dosage of clodronate, thereby reducing adverse reactions primarily due to its acidity, thus improving overall patient compliance.

[0006] After decades of clinical use of hyaluronic acid, it is now known that the effect of linear hyaluronic acid is far less than that of cross-linked hyaluronic acid. The characteristic of cross-linked hyaluronic acid is that it stays in the joint for a longer time than linear hyaluronic acid.

[0007] Scientific research shows that cross-linked hyaluronic acid has a residence time of approximately 1-4 weeks, while linear hyaluronic acid only lasts about 2-3 days (Conrozier, Joint Bone Spine 83, 2016, 1-2). Literature reports that this is due to the unique chemical structure of cross-linked hyaluronic acid, making it less susceptible to degradation by naturally occurring hyaluronidase within the joint cavity.

[0008] It is also known that the combined use of cross-linked hyaluronic acid with clodronic acid or its salts (e.g., sodium salt) has therapeutic effects.

[0009] For example, Palmieri, Drug Design, Development and Therapy 2013:7 7-12 reported positive results in treating knee osteoarthritis pain symptoms using two active ingredients; in this study, the authors mixed commercial drug solutions containing individual active ingredients prior to use to obtain a temporary solution of their combination.

[0010] However, drugs prepared in this way (i.e., ad hoc formulations) must be used quickly because the two active ingredients are not very stable once they come into contact with each other, and the drugs tend to degrade rapidly.

[0011] Therefore, even today, the primary goal of OA treatment is to manage and alleviate the pain symptoms that always accompany the pathology, which severely impact the quality of life of affected patients. This goal can also be achieved through the most minimally invasive treatment methods possible.

[0012] Therefore, the object of the present invention is to solve the above-mentioned problems by providing a new treatment alternative for treating OA, particularly for treating OA pain symptoms, which is effective, safe and has satisfactory patient compliance. Summary of the Invention

[0013] The inventors of this invention have unexpectedly discovered that, by using a combination of cross-linked hyaluronic acid or its salts or derivatives, clodronate or its equivalent salts, and acetate buffer, it is possible to obtain the aforementioned effective treatment alternative.

[0014] In fact, adding acetate buffer to a composition containing cross-linked hyaluronic acid or its salts or derivatives, and clodronic acid or an equal amount of its salt, makes the composition surprisingly stable, with excellent patient compliance, while being effective and safe for patients.

[0015] Surprisingly, the presence of acetate buffer can actually stabilize the composition of cross-linked hyaluronic acid or its salts or derivatives with clodronic acid or its equivalent salts so effectively that it is possible to overcome stability issues associated with the sterilization of pharmaceutical products necessary for the pharmaceutical manufacturing stage, which would lead to rapid degradation of the solution in the absence of acetate buffer or in the use of other buffers.

[0016] Therefore, the present invention relates to a composition comprising: - Cross-linked hyaluronic acid or its salts or derivatives - Chlordrolic acid or an equal salt thereof, and - Acetate buffer.

[0017] In this invention, when the term "acetate buffer" is used, it refers to an aqueous solution of acetic acid and an alkali metal acetate or an alkaline earth metal acetate. Examples of acetate buffers include aqueous solutions of acetic acid-sodium acetate, acetic acid-potassium acetate, acetic acid-calcium acetate, acetic acid-magnesium acetate, etc. According to the invention, a preferred acetate buffer is an aqueous solution of acetic acid-sodium acetate.

[0018] Optionally, the compositions of the present invention comprise a pharmaceutically acceptable carrier and / or a suitable excipient.

[0019] In a preferred embodiment, the composition of the present invention is in gel form.

[0020] In a more preferred embodiment, the composition of the present invention, preferably in gel form, comprises: - 1.8 mg / mL to 2.2 mg / mL, preferably 1.9 mg / mL to 2.1 mg / mL, more preferably about 2 mg / mL of sodium chlordronate or an equivalent amount of chlordronate or other salts thereof; - 16 mg / mL to 24 mg / mL, preferably 18 mg / mL to 22 mg / mL, more preferably about 20 mg / mL of cross-linked sodium hyaluronate; - Sodium acetate at concentrations of 0.6 mg / mL to 1.2 mg / mL, preferably 0.8 mg / mL to 1 mg / mL, and acetic acid at concentrations of 0.028 mg / mL to 0.052 mg / mL, and - water.

[0021] The compositions of the present invention in gel form optionally contain excipients.

[0022] In other preferred embodiments, the composition, preferably in gel form, comprises: - 1.8 mg / mL to 2.2 mg / mL, preferably 1.9 mg / mL to 2.1 mg / mL, more preferably about 2 mg / mL of sodium chlordronate or an equivalent amount of chlordronate or other salts thereof; - Cross-linked sodium hyaluronate at a concentration of 17 mg / mL to 21 mg / mL, preferably 18 mg / mL to 20 mg / mL, more preferably 19 mg / mL; - Sodium acetate from 0.6 mg / mL to 1.2 mg / mL, preferably from 0.8 mg / mL to 1 mg / mL, and acetic acid from 0.028 mg / mL to 0.052 mg / mL; - Linear sodium hyaluronate, from 0.5 mg / mL to 2 mg / mL, preferably about 1 mg / mL, and - water.

[0023] Optionally, the compositions of the present invention further comprise excipients.

[0024] In another aspect, the present invention relates to a single-dose administration form comprising 1 mL to 5 mL, preferably 2 mL, of the above-described gel-form composition.

[0025] In another aspect, the present invention relates to the compositions of the invention for use alone or in combination (i.e., as a co-therapeutic agent in a combination therapy of fixed-dose combinations or separately formulated active ingredients) for the treatment of osteoarthritis by intra-articular administration.

[0026] In this invention, when referring to "osteoarthritis", it also means "osteoarthritis".

[0027] The inventors also recognize that the compositions of the present invention make it possible to improve the symptoms of osteoarthritis.

[0028] Therefore, the present invention relates to a medical device comprising the composition and excipients of the present invention.

[0029] The medical device, and therefore the composition of the present invention, can be used to improve the symptoms of osteoarthritis, particularly those related to joint pain.

[0030] The medical device and therefore the composition of the present invention can then be used in combination therapy with known drugs for treating osteoarthritis.

[0031] In another aspect, the present invention relates to a method for preparing the above-described composition. Detailed Implementation

[0032] In this invention, the following terms are used: - "Clodronate or its equivalent salts" is intended to include all its polymorphic forms, including amorphous and crystalline forms, eutectic forms, and anhydrous, hydrated, and solvated forms of clodronate or its salts. The salts are intended to form with any pharmaceutically acceptable cation; the preferred cation is sodium.

[0033] - "Acetate buffer" refers to an aqueous solution of acetic acid and an alkali metal acetate or an alkaline earth metal acetate. Examples of acetate buffers include aqueous solutions of acetic acid-sodium acetate, acetic acid-potassium acetate, acetic acid-calcium acetate, acetic acid-magnesium acetate, etc. According to the present invention, a preferred acetate buffer is an aqueous solution of acetic acid-sodium acetate.

[0034] - "Cross-linked hyaluronic acid or its salts or derivatives" means, for example, hyaluronic acid cross-linked by divinyl sulfone (DVS), glutaraldehyde (GTA), 1,4-butanediol diglycidyl ether (BDDE) or other chemical cross-linking methods, or hyaluronic acid cross-linked by physical cross-linking methods (e.g., photocross-linking or thermal cross-linking), or its salts having any pharmaceutically acceptable cation (preferably sodium); or derived hyaluronic acid, such as hyaluronic acid having compounds selected from the group consisting of carbodiimide, carbonyl diimidazole, and hydrazine sulfate; said derivatives also include organic and inorganic salts of hyaluronic acid cross-linked or derived by the above methods. According to the invention, preferred cross-linked hyaluronic acid is cross-linked hyaluronic acid obtained by cross-linking with 1,4-butanediol diglycidyl ether (BDDE).

[0035] - "Linear hyaluronic acid or its salt" means hyaluronic acid that has not undergone any cross-linking process, and its salt is intended to have any pharmaceutically acceptable cation; the preferred cation is sodium.

[0036] - "Pharmaceutically acceptable carrier" means a carrier suitable for the final formulation, preferably water, more preferably water for intra-articular application.

[0037] - "Osteoarthritis (OA)" refers to degenerative and / or inflammatory processes affecting joints, including osteoarthritis.

[0038] - G', or elastic modulus, refers to the energy stored in the elastic structure of a solid (e.g., a gel) during deformation; the higher the G', the greater the stiffness of the material. Elastic modulus G' is commonly used to describe the solid component of a gel.

[0039] In the context of this invention, the modulus G' is preferably measured at room temperature (typically 25°C) using a rheometer (Kinexus Pro type in this application), with a 40 mm plate / plate geometry and a 250 µm gap.

[0040] The sample was pre-sheared and allowed to stand for a period of time to eliminate "rheological memory", and then the linear viscoelastic region was determined by amplitude scanning (e.g. at a fixed frequency of 1 Hz).

[0041] The modulus G' can then be determined from the curve obtained in the linear viscoelastic region, where stress and displacement are linearly related.

[0042] - The term "extrusion force" refers to the force required to push a product out of a syringe through the needle.

[0043] In this invention, the extrusion force is measured using an MTS Adamel Lhomargy DY30 universal testing machine (or universal testing machine).

[0044] According to the protocol used, the needle was connected to the syringe, a speed of 1 cm / min was applied to the syringe plunger, and the average extrusion force between 10 mm and 30 mm displacements was recorded.

[0045] In the context of this specification and the following claims, all numerical values ​​of indicated quantities, parameters, percentages, etc., shall in all cases be considered to be preceded by the term "about," unless otherwise stated. Furthermore, in addition to those ranges specifically indicated in the following description, all ranges of numerical values ​​include all possible combinations of maximum and minimum values, as well as all possible intermediate ranges. Preferably, the composition of the present invention for treating osteoarthritis (OA) is intended to treat the pain symptoms of osteoarthritis (OA).

[0046] Therefore, the present invention relates to a composition comprising: - Cross-linked hyaluronic acid or its salts or derivatives - Chlordrolic acid or an equal salt thereof, and - Acetate buffer.

[0047] Therefore, the compositions of the present invention comprise chlordronate or an equal salt thereof. The salt of chlordronate is preferably a salt of chlordronate and a pharmaceutically acceptable cation. Preferably, the cation is sodium.

[0048] Therefore, the composition preferably contains sodium chlordronate.

[0049] Therefore, the compositions of the present invention comprise cross-linked hyaluronic acid or its salts or derivatives. Preferably, the cross-linked hyaluronic acid is in the form of a salt, more preferably in the form of an alkali metal salt or an alkaline earth metal salt, and even more preferably in the form of a sodium salt. Therefore, the cross-linked hyaluronic acid of the present invention is preferably in the form of sodium hyaluronate.

[0050] The crosslinked hyaluronic acid or its salt or derivative of the present invention is hyaluronic acid that has been crosslinked with a crosslinking agent. The crosslinking agent is preferably selected from the group consisting of 1,4-butanediol diglycidyl ether (BDDE), divinyl sulfone (DVS) and glutaraldehyde (GTA); more preferably it is 1,4-butanediol diglycidyl ether (BDDE).

[0051] The compositions of the present invention comprise an acetate buffer.

[0052] According to the present invention, the acetate buffer is an aqueous solution of acetic acid and an alkali metal acetate or an alkaline earth metal acetate. Examples of acetate buffers include aqueous solutions of acetic acid-sodium acetate, acetic acid-potassium acetate, acetic acid-calcium acetate, acetic acid-magnesium acetate, etc. According to the present invention, a preferred acetate buffer is an aqueous solution of acetic acid-sodium acetate, and a more preferred aqueous solution comprising acetic acid, sodium acetate, and sodium chloride.

[0053] Surprisingly, the presence of acetate buffer can actually stabilize the combination of cross-linked hyaluronic acid or its salts or derivatives with clodronic acid or its equivalent salts so effectively that it is possible to overcome stability issues associated with the sterilization process of pharmaceutical products necessary for the pharmaceutical manufacturing stage, which would lead to the rapid degradation of active clodronic acid / clodronate in the absence of acetate buffer or in the use of other buffers, as shown in the experimental section below.

[0054] Clearly, the chemical equilibrium involved in the various substances in the composition is a complex equilibrium, involving not only the definition of the pH value that can stabilize the substances involved, but also the fact that these substances are very stable at completely different pH values.

[0055] Reasonably, there are other equilibria and reactions that are also temperature-triggered, particularly during the sterilization phase of complex compositions, which can lead to chemical reactions and degradations that depend entirely on the present chemicals and are independent of the selected pH value.

[0056] In fact, as shown in the experimental section below, buffer solutions capable of imparting the same pH value unexpectedly failed to ensure that the pharmaceutical compositions prepared throughout the entire manufacturing cycle (including the subsequent sterilization step) have the same stability properties; the entire manufacturing cycle includes not only the preparation of an initial stable solution of crosslinked hyaluronic acid or its salts or derivatives with clodronic acid or its salts (which is itself challenging due to the different stability conditions required by the two active ingredients), but also the subsequent sterilization step.

[0057] In fact, downstream of the sterilization stage, the inventors of this invention unexpectedly discovered that apparently stable compositions obtained by using different buffer solutions with similar pH values ​​experienced different stresses, which had an unexpected effect on the stability of these compositions, which only showed acceptable impurity profiles and degradation in the presence of acetate buffer.

[0058] Therefore, unexpectedly, acetate buffer plays a fundamental role in the chemical properties of these cross-linked hyaluronic acids or their salts or derivatives in combination with clodronic acid or its equivalent salts, stabilizing these substances during the pharmaceutical formulation and subsequent sterilization stages, while also ensuring sufficient G' value (i.e., elastic modulus) for use in gel form, preferably by pre-filled syringes, thus allowing for their safe and reliable manufacture and commercialization.

[0059] Preferably, the acetate buffer solution of the present invention is an aqueous solution of sodium acetate / acetic acid.

[0060] The compositions of the present invention preferably also contain linear hyaluronic acid or a salt thereof.

[0061] The compositions of the present invention preferably also contain excipients, more preferably pharmacologically acceptable excipients for intra-articular administration.

[0062] The compositions of the present invention may also contain a pharmaceutically acceptable carrier. The carrier is preferably water.

[0063] Suitable pharmaceutically acceptable excipients for intra-articular application include, for example, pH adjusters, isotonic adjusters, stabilizers, chelating agents, preservatives, and antioxidants.

[0064] Sodium chloride can be cited as an example of an excipient present in the composition.

[0065] In another aspect, the composition of the present invention is in gel form.

[0066] In a preferred embodiment, the composition, preferably in gel form, comprises: - 1.8 mg / mL to 2.2 mg / mL, preferably 1.9 mg / mL to 2.1 mg / mL, more preferably about 2 mg / mL of sodium chlordronate or an equivalent amount of chlordronate or other salts thereof; - 16 mg / mL to 24 mg / mL, preferably 18 mg / mL to 22 mg / mL, more preferably about 20 mg / mL of cross-linked sodium hyaluronate; - Sodium acetate at a concentration of 0.6 mg / mL to 1.2 mg / mL, preferably 0.8 mg / mL to 1 mg / mL, and acetic acid at a concentration of 0.028 mg / mL to 0.052 mg / mL; and - water.

[0067] Optionally, the compositions of the present invention contain suitable excipients.

[0068] In other preferred embodiments, the composition, preferably in gel form, comprises: - 1.8 mg / mL to 2.2 mg / mL, preferably 1.9 mg / mL to 2.1 mg / mL, more preferably about 2 mg / mL of sodium chlordronate or an equivalent amount of chlordronate or other salts thereof; - 17 mg / mL to 21 mg / mL, preferably 18 mg / mL to 20 mg / mL, more preferably about 19 mg / mL of cross-linked sodium hyaluronate; - Linear sodium hyaluronate from 0.5 mg / mL to 2 mg / mL, preferably about 1 mg / mL; - Sodium acetate from 0.6 mg / mL to 1.2 mg / mL, preferably from 0.8 mg / mL to 1 mg / mL, and acetic acid from 0.028 mg / mL to 0.052 mg / mL; - water.

[0069] Optionally, the composition contains a suitable excipient.

[0070] The compositions of the present invention (preferably in the preferred embodiment containing the addition of linear sodium hyaluronate) advantageously allow a gel extrusion force in the range of 18 N to 65 N, preferably less than or equal to 30 N, more preferably about 30 N, as measured by a universal testing machine at a speed of 1 cm / min; the product is extruded from a 2 mL volume syringe with a 21G1” 1 / 2 type needle.

[0071] The resulting gel-form composition is the preferred composition for intra-articular application.

[0072] The gel-form composition can be prepared by a method comprising the following steps: a. A gel is obtained by subjecting linear hyaluronic acid to hydration and cross-linking through an aqueous solution containing a cross-linking agent; b. Neutralize the gel obtained in step (a) by adding an acid solution to obtain a neutralized gel; c. The neutralized gel from step (b) is purified by dialysis to obtain a dialyzed gel; d. Add a solution containing sodium clodronate, acetate buffer, and optionally linear hyaluronic acid to the dialyzed gel from step (c); e. Mix the components from step (d) to obtain a uniform gel.

[0073] In step (a), linear hyaluronic acid is hydrated and then crosslinked with an aqueous solution containing a crosslinking agent, preferably by adding it to a basic hydrated solution containing water and the crosslinking agent. In a preferred embodiment of the invention, the basic aqueous solution of the crosslinking agent is a basic solution of the crosslinking agent 1,4-butanediol diglycidyl ether (BDDE). The basic solution containing 1,4-butanediol diglycidyl ether (BDDE) is preferably an aqueous solution of sodium hydroxide.

[0074] In step (b), the gel obtained in step (a) is preferably dissolved in a hydrochloric acid solution diluted in a buffer solution.

[0075] In step (d), the acetate buffer solution is preferably in the form of an aqueous solution of sodium chloride, sodium acetate and acetic acid.

[0076] The gel-form compositions used in this invention can advantageously undergo further processing steps in order to be dispensed in a suitable application form.

[0077] In a preferred embodiment of the invention, the composition used in gel form undergoes a sterilization step after insertion into an application form (e.g., a syringe).

[0078] Preferably, the sterilization step is performed in an autoclave at 125°C for 250 seconds.

[0079] In a preferred embodiment, the pH of the gel-form composition is 4.8 to 5.4, preferably 5 to 5.2.

[0080] In another preferred embodiment, the elastic modulus G' of the gel-form composition at 1 Hz, measured using a plate rheometer at a 250 µm gap and 1 Hz, is greater than or equal to 50 Pa.

[0081] In other preferred embodiments, the compositions used in this invention have an osmotic pressure in the range of about 250 to about 400 mOsm / L, preferably about 270 to about 380 mOsm / L.

[0082] In other preferred embodiments, the gel-form composition of the present invention is administered as a single dose in a volume of 1 mL to 5 mL, more preferably 2 mL.

[0083] The preferred form of single-dose administration is a syringe.

[0084] The compositions of the present invention or the previously described single-dose administration forms preferably comprise about 38 mg of cross-linked sodium hyaluronate, about 2 mg of linear sodium hyaluronate, about 4 mg of clodronate sodium, about 1.8 mg of sodium acetate, about 0.08 mg of acetic acid, and about 18 mg / mL of sodium chloride.

[0085] The compositions of the present invention or the previously described single-dose administration forms are advantageously used as medicines, preferably for the treatment of osteoarthritis.

[0086] In another aspect, the compositions of the present invention can be used alone or in combination, i.e., as a co-treatment agent in a combination therapy of fixed-dose combinations or separately formulated active ingredients, for the treatment of osteoarthritis.

[0087] In another aspect, the present invention relates to a medical device comprising the composition of the present invention and optionally a suitable excipient.

[0088] The medical device is preferably used to improve the symptoms of osteoarthritis, wherein the improvement includes reducing joint pain.

[0089] As shown in the experimental section below, the compositions used in this invention are stable, reproducible, and can be used to prepare single-dose administration forms, such as pre-filled syringes.

[0090] The present invention will now be described by way of some embodiments, which should be understood as explaining the invention rather than limiting it.

[0091] Experimental Section Example 1. Preparation of a gel composition comprising cross-linked hyaluronic acid, sodium chlordronate and acetate buffer according to the present invention.

[0092] A gel composition was prepared according to the present invention using the following procedure: a. By adding an alkaline aqueous solution of sodium hydroxide at a pH of approximately 13 (containing 25% 1,4-butanediol diglycidyl ether (BDDE)), linear hyaluronic acid with an average molecular weight of approximately 3 MDa undergoes hydration and cross-linking until a cross-linked hyaluronic acid gel is formed.

[0093] b. The gel obtained in step (a) is then neutralized by adding hydrochloric acid solution diluted in acetate buffer; c. Then, the neutralized gel from step (b) is purified by dialysis to obtain a purified gel with a pH of approximately 4.98; d. Add an aqueous solution containing 100 mg / mL sodium chlordronate and acetate buffer (obtained by mixing 100 mg / mL acetic acid, 100 mg / mL sodium acetate and 100 mg / mL sodium chloride) to the dialyzed gel in step (c), and mix thoroughly until a homogeneous gel according to the invention is obtained.

[0094] The gel, measured at 1 Hz using a plate rheometer with a 250 µm gap, had a G' value of approximately 267.2 Pa and a pH of approximately 4.81.

[0095] The gel was then sterilized by treating it at approximately 125°C for 250 seconds.

[0096] At the end of the sterilization process, the G' value of the gel dropped to approximately 77.1 Pa, retaining only about 28.5% of its initial value.

[0097] The pH did not change significantly, and the result was approximately 4.88.

[0098] Accelerated stability tests were also conducted on the sterilized gel. Under these stress conditions of 60°C and 96% humidity, the G' value further decreased to approximately 36.3 Pa.

[0099] Example 2. Preparation of a gel composition comprising cross-linked hyaluronic acid, sodium chlordronate and citrate buffer.

[0100] A composition identical to that of Example 1 was prepared, the only difference being that citrate buffer was used instead of the acetate buffer of the present invention.

[0101] Specifically, the added citrate buffer is an aqueous solution containing 0.98 mg / mL citric acid, 3.54 mg / mL sodium citrate, and 9 mg / mL sodium chloride.

[0102] Under the same conditions, a cross-linked hyaluronic acid gel was obtained; the gel had a G' value of approximately 406.8 Pa and a pH of approximately 5.0, as measured by a plate rheometer at 1 Hz with a gap of 250 µm.

[0103] The gel was then sterilized by treating it at approximately 125°C for 250 seconds.

[0104] At the end of the sterilization process, the G' value of the gel dropped to approximately 159.2 Pa, retaining only about 39.0% of its initial value.

[0105] The pH remained unchanged at approximately 5.0.

[0106] Accelerated stability tests were also conducted on the sterilized gel. Under these stress conditions of 60°C and 96% humidity, the G' value further decreased to approximately 113.1 Pa.

[0107] Example 3. Comparison of test results obtained from the compositions of Example 1 and Example 2 according to the present invention. Impurity profile analysis of the sterilized gel.

[0108] The results of G' before and after sterilization were then compared between the two compositions of Example 1 (COMP1inv) and Example 2 (COMP2).

[0109] These results are reported in Table 1 below.

[0110]

[0111] Table 1 - G' values ​​of the gels of the compositions of Examples 1 and 2 before and after the sterilization step.

[0112] As can be seen from the results reported in the table, the composition of Example 2 containing citrate buffer showed an even better profile in terms of parameter G'.

[0113] To test the effectiveness of using two buffer solutions in the gel formulation, the impurity profiles of each gel were analyzed after the sterilization stage, as the sterilization stage is known to cause significant degradation of the active ingredients, particularly sodium clodronate. Impurity determination was performed using ion chromatography (Thermo Fisher Scientific instrument).

[0114] The known impurities associated with chlordrophosphonate and reported in the European Pharmacopoeia (Monograph 1777) consist of so-called impurity D and phosphate. Unexpectedly, in the analysis conducted by the inventors, previously undescribed impurities were also discovered, and thus designated Unk1, Unk2 and Unk3 (where the prefix "Unk" naturally stands for unknown).

[0115] The analytical results for the citrate-containing buffer composition of Example 2 (three replicate experiments were performed, and the average value was reported) are shown in Table 2, and the analytical results for the acetate-containing buffer composition of Example 1 according to the present invention (three replicate experiments were performed, and the average value was reported) are shown in Table 3.

[0116] Impurity D was not reported in these tables because it was below the instrument detection threshold in both cases.

[0117]

[0118] Table 2 shows the changes in chlorodrophosphonate titers and impurity profiles of the COMP2 composition after sterilization.

[0119]

[0120] Table 3 shows the changes in chlorophosphonate titer and impurity profile of the COMP1inv composition after sterilization.

[0121] As can be seen from Tables 2 and 3, the results demonstrate the reproducibility and reliability of the method.

[0122] Regarding the total degradation of chlorodrophosphonate, the results in both cases appear comparable, with the results of the Example 2 composition containing citrate buffer still slightly superior to those obtained in Example 1 using acetate buffer according to the invention. Indeed, in the latter case, approximately 1% or more of the active ingredient was degraded.

[0123] However, quite unexpectedly, the impurity spectrum that appeared was completely different.

[0124] Although the compositions of the two starting gels (except for the buffer) were in fact the same, and the pH values ​​of the two gels were almost the same before and after sterilization (approximately 5 in the case of citrate buffer and approximately 4.9 in the case of acetate buffer according to the invention), completely different individual impurity values ​​were obtained.

[0125] This demonstrates that unpredictable physicochemical equilibria and chemical reactions are established within the gel, primarily due to the role of the buffer solution present therein.

[0126] In fact, the chemicals that form the buffer, in addition to controlling the pH of the composition in the same way in both cases (considering the same pH value obtained), also affect other properties of the system, namely the stability of chlordronate in the gel and its tendency to follow kinetic degradation that is significantly very different in both cases.

[0127] Therefore, the citrate buffer and acetate buffer according to the present invention have completely different abilities to interact with hyaluronic acid and chlorodrophosphonate gels. Obviously, this difference is manifested by the different rheological properties of chlorodrophosphonate when it comes into contact with one or the other sample, resulting in such different impurity spectra.

[0128] In particular, in the case of citrate buffer (COMP2, Table 2), the percentage value of the component referred to as UNK3 is as high as 2.48%.

[0129] As is well known, in the pharmaceutical field, ICH guidelines do not recommend the use of active ingredients that may degrade, resulting in the release of unclassified or non-classifiable substances at concentrations higher than 1%.

[0130] Therefore, citrate buffer, although it appears to be the preferred buffer based on the results obtained based on the G' parameter (even under stress conditions), can never be used to formulate combinations of hyaluronic acid and chlordronate due to its unsuitable impurity profile.

[0131] On the other hand, in the case of the acetate buffer according to the invention, the concentration of impurity Unk3 detected was 0.97%, which is therefore within acceptable limits. The same applies to the other two impurities Unk1 and Unk2, both of which are below the identification threshold of 0.5%; except for phosphate, which is a known impurity that is not considered a danger to human health in any way.

[0132] Thanks to the presence of acetate buffer, the compositions of the present invention are therefore the only compositions suitable for drug development.

[0133] Example 4. Preparation of a formulation comprising linear hyaluronic acid according to the present invention.

[0134] A gel-like composition according to the invention was also prepared, comprising hyaluronic acid, sodium chlordronate, acetate buffer, and linear sodium hyaluronate.

[0135] Table 4 below details the qualitative and quantitative composition:

[0136] Table 4 - Compositions according to the invention that also contain linear hyaluronic acid in gel form.

[0137] The compositions of the present invention shown in Table 4 for intra-articular use in gel form were prepared according to the same procedure as defined in the preceding examples, except that in step (d), linear hyaluronic acid with an average molecular weight of 1.5 MDa was added to the reaction mixture at the same time as the sodium chlordronate solution was added.

[0138] This gel formulation has been shown to be completely stable over time, and even after the sterilization stage, its impurity levels and degradation of sodium clodronate are completely acceptable according to pharmacopoeia standards. Furthermore, its G' value of approximately 383 Pa makes it easy to administer from a pre-filled syringe, thus allowing the composition to be marketed in a pharmaceutical form with high patient compliance.

Claims

1. A composition comprising: - Cross-linked hyaluronic acid or its salts or derivatives - Clodronic acid or an equal salt thereof, and - Acetate buffer.

2. The composition according to claim 1, wherein the crosslinked hyaluronic acid or its salt or derivative is sodium hyaluronate crosslinked with butylene glycol diglycidyl ether (BDDE).

3. The composition according to claim 1 or 2, wherein the chlordronate or its salt is sodium chlordronate.

4. The composition according to any one of claims 1 to 3, wherein the acetate buffer is an aqueous solution of acetic acid / sodium acetate, preferably in the form of an aqueous solution of sodium chloride, sodium acetate and acetic acid.

5. The composition according to any one of claims 1 to 4, wherein the composition is in gel form.

6. The composition according to any one of claims 1 to 5, wherein the composition comprises: - 1.8 mg / mL to 2.2 mg / mL, preferably 1.9 mg / mL to 2.1 mg / mL, more preferably about 2 mg / mL of sodium chlordronate or an equivalent amount of chlordronate or other salts thereof; - 16 mg / mL to 24 mg / mL, preferably 18 mg / mL to 22 mg / mL, more preferably about 20 mg / mL of cross-linked sodium hyaluronate; - Acetate ions from 0.6 mg / mL to 1.2 mg / mL, preferably 0.8 mg / mL to 1 mg / mL, and acetic acid from 0.028 mg / mL to 0.052 mg / mL; and - water.

7. The composition according to any one of claims 1 to 5, wherein the composition comprises: - 1.8 mg / mL to 2.2 mg / mL, preferably 1.9 mg / mL to 2.1 mg / mL, more preferably about 2 mg / mL of sodium chlordronate or an equivalent amount of chlordronate or other salts thereof; - 17 mg / mL to 21 mg / mL, preferably 18 mg / mL to 20 mg / mL, more preferably about 19 mg / mL of cross-linked sodium hyaluronate; - Linear sodium hyaluronate from 0.5 mg / mL to 2 mg / mL, preferably about 1 mg / mL; - Acetate ions from 0.6 mg / mL to 1.2 mg / mL, preferably 0.8 mg / mL to 1 mg / mL, and acetic acid from 0.028 mg / mL to 0.052 mg / mL, and - water.

8. The composition according to any one of claims 1 to 7, wherein the composition has an extrusion force in the range of 18 N to 65 N, preferably less than or equal to 30 N, more preferably about 30 N, as measured by a universal testing machine (or universal tester) at a speed of 1 cm / min using a 21G1” 1 / 2 needle.

9. The composition according to any one of claims 1 to 8, wherein the pH of the composition is 4.8 to 5.4, preferably 5 to 5.

2.

10. The composition according to any one of claims 1 to 9, wherein the elastic modulus G' of the composition at 1 Hz is greater than or equal to 50 Pa, as measured using a plate rheometer at a 250 µm gap and 1 Hz.

11. The composition according to any one of claims 1 to 10, wherein the composition has an osmotic pressure in the range of 250 mOsm / L to 400 mOsm / L, preferably 270 mOsm / L to 380 mOsm / L.

12. A single-dose administration formulation comprising 1 mL to 5 mL, preferably about 2 mL, of the composition according to any one of claims 1 to 11.

13. The single-dose administration method according to claim 12, wherein the administration method is a syringe.

14. The composition according to any one of claims 1 to 11, or the single-dose administration form according to claim 12 or 13, comprising about 38 mg of cross-linked sodium hyaluronate and about 2 mg of linear sodium hyaluronate as a medicament.

15. The composition according to any one of claims 1 to 11, or the single-dose administration form according to claim 12 or 13, for the treatment of osteoarthritis.

16. The composition according to any one of claims 1 to 11, used alone or in combination, i.e. as a co-therapeutic agent in a fixed-dose combination or in a combination therapy of separately formulated active ingredients, for the treatment of osteoarthritis.

17. A medical device comprising the composition according to any one of claims 1 to 11 and optionally a suitable excipient.

18. A medical device comprising the composition according to any one of claims 1 to 11 for improving symptoms of osteoarthritis, wherein the improvement includes relief of joint pain.

Images