WO2012027797A1

WO2012027797A1 - Anti-inflammatory compostion

Info

Publication number: WO2012027797A1
Application number: PCT/AU2011/001136
Authority: WO
Inventors: David Charles James
Original assignee: David Charles James
Priority date: 2010-09-02
Filing date: 2011-09-02
Publication date: 2012-03-08

Abstract

The invention relates to a sterile aqueous composition for parenteral administration to a mammal. The composition comprises an effective amount of pentosan polysulfate (PPS) and hyaluronic acid (HA), or salts, modified forms or mixtures thereof. The composition has a volume of less than 30 ml and a pH of less than 6.0, and includes at least one chelating agent. A method for providing the aqueous composition is also provided.

Description

ANTI-INFLAMMATORY COMPOSTION

BACKGROUND OF THE INVENTION

The present invention relates to a sterile aqueous anti-inflammatory composition containing pentosan polysulfate (PPS) and hyaluronic acid suitable for parenteral administration to a mammal in need thereof. The invention further relates to a method for providing the composition.

FIELD OF THE INVENTION

Pentosan polysulfate is a low molecular weight polymeric xylose known as xylan typically substituted at approximately every tenth xylose unit with an iduronic acid derivative, and is heavily sulfated. Xylan is a ubiquitous component of plant cell walls and is commonly purified from bark of the Beechwood tree, treated with a sulfating agent such as chlorosulfonic acid or sulfuryl trichloride acid, before typically being further treated with sodium hydroxide to yield PPS, although other salts are known such as pentosan polysulfate calcium. PPS generally has a weight average molecular weight (Mw) of 3.0- 10.2 kDa (and a number average molecular weight (Mn) of 2.5 kDa-8.0 IcDa) and more generally, a Mw of 4.0 kDa-7.5 kDa depending on the mode of preparation and/or the source of the xylan used in its preparation.

Pentosan polysulfate is more effective at stimulating the biosynthesis of glycosoaminoglycan (CAG) structures of the interstitial fluid than other similar polymers, and has been used for many years to treat joint injuries and conditions such as arthritis in animals. Other polymers which have been used for the treatment of such joint conditions include GAGs such as hyaluronic acid and sodium chondroitin sulfate.

Hyaluronic acid (HA) is another GAG and is also known as hylauron. It is found in the synovial fluid in joints where it plays a significant role as a lubricant and is a component of articular cartilage of joints. HA has also been reported to moderate the inflammatory response. The therapeutic advantages of PPS and HA for the treatment of noninfectious joint inflammation are well recognised by veterinarians.

Amino sugars such as glucosamine, N-Acetyl-D-glucosamine and galactosamine are building blocks of GAGs, and are commonly included in nutritional supplements or administered to animals for the maintenance and repair of joint connective tissues and synthesis of GAGs. The mechanism of action in non-infectious based joint inflammation is poorly understood. However, it is believed they stimulate the synthesis of sulfated GAGs and HA.

However, PPS has poor bioavailability when administered by ingestion, and as a result both PPS and HA are often administered to animals by intravenous injection. To do this, a veterinarian can administer the PPS and HA either as two separate injections one after the other, or as a single injection drawn from separate PPS and HA preparations immediately prior to use. This has been necessary due to the incompatibility of PPS and HA in the one composition.

Hence, when combined and injected together, the volume of the resulting PPS and

HA mixture is relatively large. For a large animal such as a horse or camel this can be SO ml or more. Nevertheless, the injection can only be administered at a normal rate. Animals need to be restrained when given injections and any delay in administering the injection risks injury to the animal and the veterinarian and/or animal handler(s). An anima] such as horse can, for instance, rear or attempt to pull away resulting in potentially significant damage being inflicted to the vein or surrounding tissue. Alternatively, or as well, the required dosage of the actives may only be partially administered. However, an animal may only be able to be restrained for 10 to 20 seconds for the injection to be made.

A veterinary or therapeutic composition for injection should be sterile. As such, drawing PPA and HA from different vials into the one syringe for a single injection is undesirable. Moreover, both PPA and HA are degraded by heat sterilization, and their different characteristics makes any single form of sterilization process unsuitable for both when provided together in a small volume. Further to this, their incompatibility has inherently required that they be administered in larger volumes that are unsuitable for intravenous injection as described above, or as solid form oral preparations as described in Australian Patent No. 747595.

SUMMARY OF THE INVENTION

Broadly stated, the invention relates to the provision of a sterile aqueous composition of pentosan polysulfate (PPS) and hyaluronic acid in a small volume suitable for parenteral administration. The combination of these two actives together in the one composition is problematic due to the propensity for PPS and hyaluronic acid to form a gel like precipitate that once formed essentially cannot be easily re-dissolved back into solution, if at all.

Moreover, it is essential that a composition for parenteral administration be provided in a sterile form. However, neither PPS nor hyaluronic acid can be radio- sterilized (e.g., by gamma irradiation) or chemically sterilized (e.g., via ethylene oxide treatment) in solution without degrading the polymeric structure of the compounds.

Further, hyaluronic is not suitable for filter sterilization at concentrations and molecular weights desirable for therapeutic activity as the hyaluronic acid is fractionated by the sterilizing filter. That is, smaller hyaluronic acid chains pass through the filter but larger ones will not. Undesirably, this results in blinding (or blocking) of the filter.

Although hyaluronic acid can be heat sterilized such as by autoclaving, the application of heat results in denaturation of the tertiary structure and/or de-polymerization of the molecule. Similarly to hyaluronic acid, PPS is also degraded by beat causing desulfation to xylan rendering the molecule less soluble and thereby, making heat sterilization processes inappropriate for solutions of PPS.

The problem of providing PPS in combination with hyaluronic acid in small aqueous solution volumes is yet further exacerbated when a third active is included in the composition due to the different molecular weights, thermal stabilities and chemical characteristics of the different actives.

Despite the significant physical incompatibilities associated with the provision of PPS and hyaluronic acid together in a small volume of aqueous solution, the inventor has nevertheless been successful in providing such compositions. In particular, the inventor has recognized that although hyaluronic acid depolymerizes when subjected to heat sterilization, a larger molecular weight starting hyaluronic acid can be heat treated to provide a hyaluronic acid break down product having a molecular weight suitable for therapeutic administration. Further, the inventor has found that PPS and the hyaluronic acid can be stably maintained in solution without the formation of a gel-like precipitate with the use of chelating agent(s) and an acidic solution pH. Without being limited by theory, it is thought that PPS and hyaluronic acid are cross-linked under basic conditions by divalent metal ions such as Ca²⁺ and Mg²⁺ introduced by preparations of the PPS and/or hyaluronic acid feedstocks. However, under low pH conditions hyaluronic acid is degraded or modified as to render HA unstable in the combination. Hence, the provision of a composition embodied by the invention is the result of the combination of a number of separate findings and observations together in a controlled formulation.

In particular, in an aspect of the invention there is provided a sterile aqueous composition for parenteral administration to a mammal, comprising an effective amount of pentosan polysulfate (PPS) and hyaluronic acid, the composition having a volume of less than 30 ml and a pH of less than 6.0, and including at least one chelating agent.

Typically, the composition comprises the combination of sterilized hyaluronic acid with sterilized pentosan polysulfate.

In another aspect of the invention there is provided a method for providing a sterile aqueous composition for parenteral administration to a mammal, comprising:

filter sterilizing a solution of pentosan polysulfate (PPS);

providing a sterilized solution of hyaluronic acid (HA); and

mixing the sterilized solution of pentosan polysulfate and sterilized solution of hyaluronic acid together to provide a mixture containing at least one chelating agent and an effective amount of the PPS and hyaluronic acid in a volume of 30 ml or less, the mixture having a pH of 6.0 or less.

Typically, a solution of hyaluronic acid is heat sterilized to provide the sterilized solution of hyaluronic acid. Most typically, a method embodied by the invention comprises providing the hyaluronic acid in an initial molecular weight range, the hyaluronic acid molecular weight being reduced by the heat sterilization to a molecular weight in a range for administration to the mammal.

Typically, the composition further comprises an effective amount of at least one amino sugar, or a polymer, salt, isomer or a modified form thereof. The polymer may, for example, be an oligomer (e.g., a dimer, trimer etc.) An oligomer will generally comprise between 2 to 20 monomers of the amino sugar, more typically between 2 to 10 monomers and most usually, 2 to 6 sugar monomers. Most typically, the amino sugar is a monosaccharide such as N- Acetyl-glucosamine.

Typically, the amino sugar or modified form thereof is filter sterilized in the solution with the PPS, and the sterilized PPS and amino sugar is mixed with the hyaluronic acid.

Typically, the composition also comprises a buffer system for maintaining the pH of the composition in a predetermined pH range. The predetermined pH range maintains stability of the PPS and HA together in the one solution in the presence of absence of an amino sugar.

Typically, the pH of a composition embodied by the invention is in a range of from 4.5 to 5.5 and most usually, is in a pH range of from 5.0 to 5.3.

Compositions embodied by the invention are particularly suitable for prophylaxis or treatment of non-infectious inflammation in an animal and particularly, non-infectious joint inflammation.

Advantageously, by providing PPS and hyaluronic acid together in a small volume as an aqueous sterile composition with or without an amino sugar included in the composition in accordance with embodiments of the invention, the actives can be administered together by a veterinarian or other suitably trained person in the one injection, minimizing the time required to complete the injection and discomfort to the mammal. When the mammal is a horse, pony or similarly large animal, the risk of injury to the animal and/or the person injecting the formulation arising from the animal abruptly drawing away, rearing or otherwise fighting the treatment is likewise also reduced.

Throughout this specification the word "comprise", or variations such as

"comprises" or "comprising", will be understood to imply the inclusion of a stated element, integer or step, or group of elements, integers or steps, but not the exclusion of any other element, integer or step, or group of elements, integers or steps.

All publications mentioned in this specification are herein incorporated by reference. Any discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is solely for the purpose of providing a context for the present invention. It is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the Meld relevant to the present invention as it existed in Australia or elsewhere before the priority date of this application.

The features and advantages of the invention will become further apparent from the following detailed description of a number of embodiments of the invention.

DETAILED DESCRIPTION OF EXAMPLARY EMBODIMENTS OF THE INVENTION

The PPS utilized in a composition embodied by the invention can be a native PPS or a salt thereof such as sodium PPS, or a modified form of PPS. The PPS will generally have a weight average molecular weight (Mw) in a range of from about 3.0 kDa to about 10.2 kDa (and a number average molecular weight (Mn) of from about 2.8 kDa to about 8.0 kDa). Usually, the PPS will have a Mw in a range of from 3.0 kDa to 7.5 kDa and more usually from 4.0 kDa to 7.5 kDa. Most usually, the PPS will have a Mw in a range of from 4.6 kDa or 4.7 kDa to 7.5 kDa. PPS with a Mw in a range of from 3.0 kDa to 7.5 kDa and more typically, 3.0 kDa to 5 kDa is particularly suitable. Sodium PPS is particularly preferred although other salts may be used, and can for example, be obtained commercially from Scentio Biosciences, St Louis Missouri, USA.

Typically, a composition embodied by the invention will comprise the PPS in a concentration in a range of from about 25 mg/ml to about 300 mg/ml, preferably in a range of from about 50 mg/ml to about 250 mg/ml and more preferably, in a range of from about 50 mg/ml to about 125 mg/ml. Hyaluronic acid utilized in the preparation of an embodiment of the invention can be moist heat sterilized (e.g., steam autoclaved) separately and added to a solution of filter sterilized PPS. The moist heat sterilization process provides a de-polymerization breakdown product, reducing the molecular weight of the starting hyaluronic acid reagent 5 to within a predetermined range suitable for therapeutic administration to the mammal.

Hence, the hyaluronic acid starting reagent initially has a higher molecular weight than that present in compositions of the invention. Typically, the weight average molecular weight of the starting hyaluronic acid is in a range of from 1.3 MDa to 1.8 MDa. Once heat sterilized, the molecular weight (Mw) of the hyaluronic acid is normally reduced to within 10 a range from about 0.35 MDa to about 1.5 MDa, more typically to within a range of from 0.35 MDa to 0.8S MDa and most typically, to within a range of from 0.3S to 0.55 MDa. As with PPS, salts of hyaluronic acid and modified forms of hyaluronic acid may be utilized.

Typically, the HA will be present in a composition embodied by the invention in a 15 range of from about 0.1 mg/ml to about 20 mg/ml, more preferably in a range of from about 0.75 mg/ml to about 12.5 mg/ml or 0.75 mg/ml to about 7.5 mg/ml and most preferably, in a range of from about 1 mg/ml to about 5 mg/ml.

In at least some embodiments, the HA may be provided in the form of a sterile powder and made to volume with a suitable diluent (e.g., deionized water or physiological 20 saline or the like) under sterile conditions for combination with the PPS (or PPS and amino sugar(s)).

The amino sugar included in a composition embodied by the invention may be selected from the group consisting of N- Acetyl-glucosamine, glucosamine,

N-Acetyl-galactosamine, glucosamine sulfate, glucosamine hydrochloride, galactosamine, 25 galactosamine sulfate, galactosamine phosphate, mannosamine, fructosamine, N-Acetyl- fructosamine, amino sugar polymers, and salts, isomers, modified forms, and mixtures of the foregoing. Typically, an isomer utilized in a composition embodied by the invention will be a D isomer (e.g., N-Acteyl-D- glucosamine (NAG)).

Typically, at least one amino sugar is included in the composition in a concentration 30 range of from 100 mg/ml to 220 mg/ml and preferably, in a range of from 120 mg/ml to about 200 mg/ml. Most preferably, the amino sugar is present in a concentration range of from 120 mg/ml to about 160 mg/ml. Monomeric NAG is particularly preferred for use in a composition of the invention although glucosamine and/or salts and isomers of either can also suitable. Likewise, polymeric forms of amino sugars can be employed, examples of which include polymeric NAG (chitin) and chitosan. Generally, the amino sugar polymer will have a weight average molecular weight in a range of from 10 kDa to 1 SO kDa.

However, like PPS, NAG is not suitable for being autoclaved as it deacetylates under the application of moist heat and releases glucosamine, turning solution brown. It is thought the browning is the result of the released amine forming coloured species. Also, in contrast to PPS and hyaluronic acid, the molecular weight of NAG and other amino sugars is much less. NAG also has poorer thermal stability in solution than both PPS and hyaluronic acid and while PPS is highly sulfated (and thereby highly charged), both NAG and hyaluronic acid are unsulfated. Indeed, hyaluronic acid is a polymeric disacchande of NAG and glucuronic acid.

Modified forms of PPS, NAG and/or the selected amino sugar(s) that may be utilized in the composition include those that have been chemically modified such as by esterification, sulfation, polysulfation, acetylation or methylation of the compound(s) and which at least essentially retain the biological and/or therapeutic activity of the native compound in the context of the present invention.

The physical differences between PPS, hyaluronic acid and NAG utilized in preferred embodiments of compositions of the invention are summarized below in Table 1.

Compound characteristics

Despite the significantly different characteristics of the compounds in solution and particularly, the different molecular weights and solution thermal stability, they can be retained stably dissolved by maintaining the pH of the solution in a controlled range and providing at least one chelating agent in the solution such as the hexadentate chelating agent ethylenediaminetetraacetic acid (EDTA). Generally, at least one physiologically acceptable antioxidant is also included in the composition. Any chelating agent for chelating polyvalent (e.g., di-valent) metal ions may be utilized. Typically, EDTA will be provided in a composition of the invention in a concentration range of from O.OS mg/ml to about I mg/ml, more preferably in a range of from 0.1 mg/ml to about 0.5 mg/ml and most preferably, in a concentration of 0.2 mg/ml. Other chelating agents that may be employed include amino acid based chelating agents, salts of chelating agents (e.g., EDTA tetra sodium or disodium salts), phosphonate chelating agents, such as ethylenediamine tetra(methy]ene phosphonic acid), carbonates which may act as softeners or sequestering agents, and other agents that remove the effect of polyvalent cations in the composition.

To maintain the pH of the composition in a range to inhibit formation of a gel by the hyaluronic acid and degradation of the PPS, hyaluronic acid and NAG or other amino sugar, a buffer system can be employed in the composition. The buffer system is formed by a mild acid and weak base. Typically, citric acid and disodium hydrogen phosphate (Na₂HP0₄) is utilized but any suitable buffer system can be used. A composition embodied by the invention may have a composition of 25 ml or less and more typically a volume of 20 ml, 18 ml or 12 ml or less. For instance, the composition may have a volume of 24, 23, 22, 21, 19, 18, 17, 16, 15, 14, 13, 11 or even 10 ml. As will be understood, generally the smaller the animal the smaller the volume of the composition. The volume of the composition is generally determined to provide a stable matrix for the delivery of a therapeutic dose of the PPS, hyaluronic acid and the amino sugar (when present).

The dosage of the composition to be administered to an animal can be determined by a veterinarian based on the type and size of the animal, the age of the animal, the severity of the inflammatory condition, and whether the composition is being administered for prophylactic or treatment purposes.

Compositions embodied by the invention can also include anti-oxidant(s) such as butylated hydoxy toluene, butylated hydroxy anisole, ascorbic acid and sodium ascorbate, preservatives such as sodium metabisulfite benzoic acid, sodium benzoate, alcohol group based preservatives, mercurial based preservatives, hydroxybenzoates, phenol based preservatives and quaternary ammonium compounds, isotonicity adjusters (e.g., sodium chloride), and de-ionised water.

Inflammatory conditions that may be treated with a composition as described herein include non-infectious inflammatory joint conditions, inflammatory conditions of connective tissue in joints, general joint inflammation and swelling, joint stiffness and conditions arising from lack or loss of joint fluids, arthritis and joint arthritic conditions. A composition embodied by the invention may also be administered for the protection, maintenance and/or repair of connective tissues of joints.

A composition embodied by the invention is typically formulated for administration by injection or infusion and is normally a veterinary pharmaceutical composition.

The mammal can, for instance, be a racing animal such as a horse, camel or greyhound, or a companion animal such as a cat or dog. However, it will be understood that the mammal can be any animal that may be treated with a composition of the invention. The stability of a composition embodied by the invention may be determined by assaying for one or more of degradation of the components of the composition, colour, clarity, pH, Specific gravity and sterility according to standard principals. A composition in accordance with the invention will generally exhibit substantially no discolouration (e.g., browning) or degradation of the PPS (as for instance may be evidence by an essentially constant level of sulfation), HA and (where applicable) amino sugar(s) in the composition over the period of its shelf-life. A composition as described herein may, for example, be assayed by high performance liquid chromatography (HPLC) and/or electrophoretic analysis (e.g., capillary electrophoresis). Generally, a composition embodied by the invention is considered stable when none of the actives in the composition (i.e., the PPS, HA and when included, the amino sugar(s), or modified form(s) thereof) deviate from their respective initial concentration in the composition by more than 10% when the composition is stored under controlled conditions (e.g., in light or dark in a sealed glass vial at

25°C/65% RH (relative humidity) and/or 40°C/75% RH.). Typically, a composition of the invention will have a shelf life when stored for at least 6 months and more typically, for at least 12 months up to 24 or 36 months or more, during which the composition remains stable and suitable for administration.

The invention is described further below by way of a number of non-limiting examples thereof. The molecular weights referred to in the following examples are all weight average molecular weights (Mw) unless expressly stated otherwise.

EXAMPLE 1 : Sterile formulation of PPS, hyaluronic acid and NAG suitable for a large animal The components of a composition embodied by the invention for parenteral administration to a horse or other large animal are set out in Table 2.

Components of composition

A powdered preparation of sodium pentosan polysulfate (PPS) and N-Acetyl-D- glucosamine was obtained commercially under the brand name Pentosan Gold™ (Nature Vet Pty Limited, Glenorie, NSW, Australia). The PPS and NAG were dissolved in deionised water with the remaining components of the composition to obtain the concentrations of PP§ and NAG shown above. The Mw of the PPS was in the range of from 4.0 kDa to 7.5 kOa.

The hyaluronic acid (HA) is prepared in 6 ml of physiological saline solution to a concentration of 10 mg/ml. Alternatively, the hyaluronic acid can be prepared to the same volume and concentration in another isotonicity agent such as phosphate buffered saline BP (PBS ^K 1.79 mg/ml disodium hydrogen phosphate; 1.36 mg/ml potassium dihydrogen phosphate; 7.02 mg/ml sodium chloride). The HA has a molecular weight in a range of from 1.5 MDa to 1.8 MDa. After being dissolved in physiological saline or PBS, the HA solution is sealed in suitable vials for sterilization. The sealed HA solution is then autoclaved utilising a moist heat (steam) sterilizer (typically purged of air by downward displacement) using a standard autoclave cycle, for example, } 21 °C for 30 minutes, to produce a sterilized HA solution. The sterile HA solution is stored in the sterile vials and transported to a sterile, aseptic blending and filling area, such as a sterile laminar flow hood. The combination PPS/NAG and excipients solution is sterilized by filtration in the same sterile/aseptic filling area by passage of the PPS /NAG solution through a plastic cartridge with a nylon membrane filter having a pore size of 0.2 urn or less(Pall Corp., Cheltenham, Victoria, Australia), although suitable filters membranes formed from other plastics can also be utilized.

Once filter sterilized, the PPS/NAG solution is mixed with the moist heat sterilized HA solution at a ratio of 1 part HA solution to 2 parts PPS/NAG solution, to which the EDTA, antioxidant are remaining components are added also with mixing. A 20 ml volume of the resulting mixture is then dispensed into sterile storage vials under sterile laminar air flow, and the vials are plugged and crimped.

EXAMPLE 2: Sterile formulation suitable for a companion animal

The components of a 12ml composition embodied by the invention for parenteral administration to a companion animal are set out in Table 3.

Components of composition

The above composition is manufactured to produce a sterile final product as per the protocol described in Example 1 except that the final volume of each aliquoted composition preparation is 12 ml rather than 20 ml.

EXAMPLE 3: Further sterile formulations These compositions are similar to those described in Examples 1 and 2 except that the HA solution concentration is increased to 2% and the final volume of the product is decreased to 15 ml for the large animal, and 11.0 ml for the small animal. The components and concentrations of these compositions are respectively shown in Table 4 and Table 5.

Components of composition

Table 5: ponents of composition

EXAMPLE 4: Sterile formulation for a large animal

A similar formulation to that described in Example 1 but which is suitable for an 18 ml dose and employs a 1% HA solution is set out in Table 6.

Table 6: Components of composition

Sodium metabisulfite is used as the anti-oxidant in Examples 1 to 4 above. The buffer system employed is the citric acid and disodium hydrogen phosphate system described in Example 1.

EXAMPLE 5: Sterile formulations with different molecular weights of hyaluronic acid (HA)

For injection intra-articularly (i.e., into the joint space) a composition embodied by the invention comprising a higher molecular weight (Mw) HA (typically 1.6 MDa or above is desirably employed to reduce the possibility of transient inflammation that can occasionally occur with the use of HA with a lower Mw. However, a composition embodied by the invention to be administered intravenously will desirably comprise HA with a Mw between about 0.3S MDa to about 1.0 MDa to minimize the risk of occlusion of small capillaries.

Stable sterile formulations prepared according to Example 1 that are suitable for a 20 ml dose but comprising approx. 1 % w/v HA having a range of different molecular weights (Mw) are described below in Tables 7 to 1 1.

Components of composition

Components of composition

Component Cone,

(mg/ml)

Sodium pentosan polysulphate 75.0

N-Acetyl-D>glucosamine 132.0

Sodium hyaluronic acid 3.0

(Mw 900 kDa)

Antioxidant 15.0

Buffer system 5.6

Components of composition

Components of composition

Table 1 1 : Components of composition

EXAMPLE 6: Sterile formulations with different amino sugars

Stable sterile compositions prepared according to Example 1 that are suitable for a 20 ml dose and comprising approx. 1% w/v HA and amino sugars other than N-Acetyl-D- glucosamine are described below in Table 12 and 13. Table 12: Components of composition

Component Cone,

(mg/ml)

Sodium pentosan polysulphatc 75.0

Galactosamine or 132.0

N-Acetyl-galactosamine

Sodium hyaluronic acid 3.0

(Mw 1800 kDa)

Antioxidant 15.0

Buffer system 5.6

Components of composition

EXAMPLE 7: Evaluation of formulation stability

An injectable composition of the invention comprising PPS, HA and N-Acetyl-D- glucosamine (NAG) was prepared essentially according to Example 1. The concentrations of the PPS, HA and NAG were assayed by HPLC with refractive index (RI) detection on Day 1 and at subsequent 6 month and 12 month time periods after storage in sealed glass vials under the conditions of 25°C/65% RH or 40°C/75% RH. The results are shown below in Table 14 to Table 17. The concentrations shown are in mg/ml and are the averages for 3 batches. The molecular weights of the PPS and HA are as described in Example 1. Table 14: PPS concentration (Theoretical cone, of PPS is 75 mg/ml. The limit for stability was set at 67.5 mg/ml to 82.5 mg/ml).

Table 15: NAG concentration (Theoretical cone, of NAG is 120 mg/ml. The limit for stability was set at 108 mg/ml to 132 mg/ml).

Table 16: Sodium HA concentration (Theoretical cone, of NaH A is 3 mg/ml. The limit for stability was set at 2.7 mg/ml to 3.3 mg/ml).

The results show the composition was stable over the 12 month period under each of the test conditions with respect to the PPS, HA and NAG concentrations of the composition.

It will be understood by persons skilled in the art that numerous variations and/or modifications may be made to the speciGc embodiments illustrated without departing from the invention as broadly described. The present embodiments arc, therefore, to be considered in all respects as not restrictive.

Claims

1. A sterile aqueous composition for parenteral administration to a mammal, comprising an effective amount of pentosan polysulfate (PPS) and hyaluronic acid (HA), or salts, modified forms or mixtures thereof, the composition having a volume of less than 30 ml and a pH of less than 6.0, and including at least one chelating agent.

2. A composition according to claim 1 being the combination of sterilized HA with sterilized PPS.

3. A composition according to claim 1 or 2 wherein the hyaluronic acid is a heat sterilization breakdown product of a larger molecular weight hyaluronic acid.

4. A composition according to any one of claims 1 to 3 further comprising an effective amount of at least one amino sugar, or a polymer, salt or modified form thereof.

5. A composition according to claim 4 wherein the amino sugar is selected from the group consisting of N-acetyl-glucosamine, glucosamine, glucosamine sulfate, glucosamine hydrochloride, galactosamine, and mixtures of the foregoing.

6. A composition according to claim 5 wherein the amino sugar is

jV-acetyl-D-glucosamine.

7. A composition according to any one of claims 1 to 6 wherein the composition has a volume of 25 ml or less.

8. A composition according to claim 7 wherein the composition has a volume of less than 20 ml or less.

9. A composition according to claim 8 wherein the composition has a volume of 18 ml or less.

10. A composition according to claim 9 wherein the composition has a volume of 12 ml or less.

1 1. A composition according to any one of claims 1 to 10 wherein the pH of the composition is in a range of from 4.5 to 5.5.

12. A composition according to claim 1 1 further comprising a buffer system for maintaining the pH of the composition in a range of from 5.0 to 5.3.

13. A method for providing a sterile aqueous composition for parenteral administration to a mammal, comprising:

filter sterilizing a solution of pentosan polysulfate (PPS);

providing a sterilizing a solution of hyaluronic acid (HA); and

14. A method according to claim 13 wherein a solution of hyaluronic acid is a heat sterilized to provide the sterilized solution of HA.

15. A method according to claim 14 comprising providing the hyaluronic acid in an initial molecular weight range, the hyaluronic acid being reduced by the heat sterilization to a molecular weight in a range for administration to the mammal.

16. A method according to any one of claims 13 to 15 wherein the composition further comprises at least one amino sugar, or a polymer, salt or modified form thereof.

17. A method according to claim 16 wherein the amino sugar, polymer, salt or modified form thereof is provided in the same said solution with the PPS and filter sterilized with the PPS, for subsequently being added to the sterilized solution of hyaluronic acid with the PPS.

18. A method according to claim 17 wherein the amino sugar is selected from the group consisting of N-acetyl-glucosamine, glucosamine, glucosamine sulfate, glucosamine hydrochloride, galactosamine, and mixtures of the foregoing.

19. A method according to claim 18 wherein the amino sugar is

N-acetyl-D-glucosamine.

20. A method according to any one of claims 13 to 19 wherein the PPS and hyaluronic acid are in a volume of 25 ml or less.

21. A method according to claim 20 wherein the PPS and hyaluronic acid are in a volume of less than 20 ml or less.

22. A method according to claim 21 wherein the PPS and hyaluronic acid are in a volume of 18 ml or less.

23. A method according to claim 22 wherein the PPS and hyaluronic acid are in a volume of 12 ml or less.

24. A method according to any one of claims 13 to 23 wherein the pH is in a range of from 4.5 to 5.5.

25. A method according to claim 24 wherein the mixture comprises a buffer system for maintaining the pH in a range of from 5.0 to 5.3.