WO2021232157A1

WO2021232157A1 - Sustained release pharmaceutical composition

Info

Publication number: WO2021232157A1
Application number: PCT/CA2021/050682
Authority: WO
Inventors: Molly Shoichet; Michael J. Cooke; Christopher Mclaughlin; Jessica NGAI
Original assignee: The Governing Council Of The University Of Toronto
Priority date: 2020-05-19
Filing date: 2021-05-19
Publication date: 2021-11-25

Abstract

A swellable, injectable, pharmaceutical composition comprising an active ingredient that is positively charged at biological pH and a hyaluronan-methylcellulose hydrogel. The active ingredient has a sustained release from the hydrogel as a result of an electrostatic interaction between a positive charge of the active ingredient and negatively charged carboxylate ions of the hyaluronan.

Description

SUSTAINED RELEASE PHARMACEUTICAL COMPOSITION

This application claims priority to United States patent application number 63/027,043 filed May 19, 2020.

TECHNICAL FIELD [0001] This invention relates to sustained release pharmaceutical compositions.

BACKGROUND OF THE ART

[0002] Delivery of local anesthetics presents issues of fast clearance from the injection site and dispersion throughout the body, resulting in short-term pain relief and potentially toxic systemic effects. Continuous delivery systems are invasive and carry risk of infection while systemic opioids have many side effects, including addiction and respiratory depression. There remains a need for effective sustained release pharmaceutical compositions.

BRIEF SUMMARY

[0003] In one embodiment, there is provided a swellable, sustained release pharmaceutical composition comprising: a hydrogel comprising a gel polymer matrix comprising 3 to 10 wt% hyaluronan or a derivative thereof and 1 to 9.99 wt% methylcellulose or a derivative thereof based on the weight of the hydrogel, wherein the amount of hyaluronan or derivative thereof is greater than the amount of methylcellulose or derivative thereof, and an active ingredient, wherein at a physiological pH the active ingredient or a portion thereof is positively charged. In one embodiment, the ratio of hyaluronan:methylcellulose is between 1.01:1 and 3:1. [0004] In certain embodiments, the composition is injectable.

[0005] In certain embodiments, the active ingredient is an amide active ingredient.

[0006] In certain embodiments, the active ingredient is lidocaine, bupivacaine, ropivacaine and/or a derivative or pharmaceutically acceptable salt thereof.

[0007] In certain embodiments, the active ingredient is ropivacaine and/or a pharmaceutically acceptable salt thereof.

[0008] In certain embodiments, the active ingredient is hydrophobic in some portion. [0009] In certain embodiments, less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the active ingredient is released from the pharmaceutical composition within 12 hours of administration.

[0010] In certain embodiments, less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the active ingredient is released from the pharmaceutical composition within 24 hours of administration.

[0011] In certain embodiments, less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the active ingredient is released from the pharmaceutical composition within 48 hours of administration.

[0012] Suitably, the hyaluronan or derivative thereof has a molecular weight between 100,000 g/mol and 3,000,000 g/mol and the methylcellulose or derivative thereof has a molecular weight between 10,000 g/mol and 500,000 g/mol

[0013] Also provided is a dosage form comprising between 1 ml_ and 100 ml_ of the pharmaceutical composition.

[0014] In certain embodiments, the dosage form comprises between 100 mg and 2000 mg of the active ingredient (more specifically between 350 mg and 2000 mg or between 750 mg and 1500 mg).

[0015] In certain embodiments, there is a method of treating or preventing pain comprising administering a therapeutically effective amount of the pharmaceutical composition or dosage form to a subject in need thereof.

[0016] In certain embodiments, there is a method of aiding wound healing comprising administering a therapeutically effective amount of the pharmaceutical composition or dosage form to a subject in need thereof.

[0017] In certain embodiments, there is a method of partially or fully sealing biological tissue comprising administering a therapeutically effective amount of the pharmaceutical composition or dosage form to a subject in need thereof.

[0018] Also provided is use and use in the manufacture of medicaments of compositions disclosed herein for the treatment or prevention of pain; for aiding wound healing; and/or for partially or fully sealing biological tissue. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1A shows release profiles of bupivacaine hydrochloride from different formulations of HAMC, shown as cumulative amount detected (%) (* p<0.05, ** p<0.01, *** p<0.001).

[0020] Figure 1 B shows release profiles of bupivacaine hydrochloride from 7:3 HAMC (7 wt% HA, 3 wt% MC) prepared in buffers of different pH, shown as cumulative amounted detected (%) (* p<0.05).

[0021] Figure 1C shows swelling ratio of 7:3 HAMC (7 wt% HA, 3 wt% MC) with and without bupivacaine (* p<0.05).

[0022] Figure 2A shows the duration of spinal nerve block after administration of bupivacaine hydrochloride solution (1.56 mg/kg) or HAMC-bupivacaine hydrochloride (1.56 mg/kg, 5 mg/kg, 10 mg/kg, 25 mg/kg, 34 mg/kg) using a von Frey assay (*** p<0.001).

[0023] Figure 2B shows the duration of spinal nerve block after administration of bupivacaine hydrochloride solution (1.56 mg/kg) or HAMC-bupivacaine hydrochloride (1.56 mg/kg, 5 mg/kg, 10 mg/kg, 25 mg/kg, 34 mg/kg) using a cold plantar assay (* p<0.05, ** p<0.01, *** p<0.001).

[0024] Figure 3A shows the duration of sciatic nerve block after administration of bupivacaine hydrochloride solution (10 mg/kg) or HAMC-bupivacaine hydrochloride (8 mg/kg, 32 mg/kg, 64 mg/kg) using a von Frey assay (** p<0.01).

[0025] Figure 3B shows the duration of sciatic nerve block after administration of bupivacaine hydrochloride solution (10 mg/kg) or HAMC-bupivacaine hydrochloride (8 mg/kg, 32 mg/kg, 64 mg/kg) using a pinch test (* p<0.05).

[0026] Figure 3C shows the duration of sciatic nerve block after administration of bupivacaine hydrochloride solution (10 mg/kg) or HAMC-bupivacaine hydrochloride (8 mg/kg, 32 mg/kg, 64 mg/kg) using a cold plantar assay (** p<0.01).

[0027] Figures 4A to 4E show representative H&E stained sections of sciatic muscle after administration of (a) saline, (b) HAMC-bupivacaine hydrochloride (8 mg/kg), (c) HAMC- bupivacaine hydrochloride (32 mg/kg), (d) HAMC-bupivacaine hydrochloride (64 mg/kg), or (e) bupivacaine hydrochloride solution (10 mg/kg).

[0028] Figures 4F to 4J show representative H&E stained sections of sciatic nerve after administration of (f) saline, (g) HAMC-bupivacaine hydrochloride (8 mg/kg), (h) HAMC- bupivacaine hydrochloride (32 mg/kg), (i) HAMC-bupivacaine hydrochloride (64 mg/kg), or (j) bupivacaine hydrochloride solution (10 mg/kg).

DETAILED DESCRIPTION

[0029] Management of post-operative pain remains suboptimal as current treatment strategies do not sufficiently address patient needs. The current standard of care is to inject local anesthetics either directly into the incision or around a nerve to provide either local analgesia or a regional nerve block. Commonly administered analgesics include local anesthetics, which are voltage-gated sodium channel blockers and serve to block action potentials, preventing neuronal transmission of the painful stimulus. However, local anesthetics are typically delivered as a liquid bolus injection, leading to fast clearance from the injection site and rapid dispersion throughout the body. Fast clearance of the drug into systemic circulation results in the potential for systemic toxicity. Indeed, the maximum recommended doses of local anesthetics are limited not by efficacy, but by systemic side effects such as cardiorespiratory failure and neurotoxicity.

[0030] Fast clearance from the site of administration results in short-lived pain relief, necessitating continuous infusion and/or the subsequent use of systemic analgesics, such as opioids, to adequately alleviate pain. Continuous delivery systems, such as pumps, have a long length of duration but require the insertion of a catheter, which is invasive and carries the risk of infection. Systemic opioids are effective at providing pain relief, but are plagued by many adverse side effects, including addiction, respiratory depression, hyperalgesia, and nausea, constipation and vomiting.

[0031] Continuous infusion of anesthetics through an implanted catheter with a control pump is the most common method used clinically to achieve nerve block. However, this strategy involves risks associated with local infection. To overcome this limitation, local anesthetics have been formulated for slow release to achieve extended nerve block, for example as polymeric microspheres, surgically implantable pellets, microcrystals, liposomes, lipospheres, or lipid- protein-sugar particles. Notwithstanding the extended nerve block achieved, adverse effects such as neurotoxicity, myotoxicity, and local inflammation often limit their clinical use. For example, a liposomal formulation of bupivacaine, which is used clinically, causes prolonged inflammation [McAlvin, J.B., et al., Multivesicular liposomal bupivacaine at the sciatic nerve. Biomaterials, 2014. 35(15): p. 4557-64] [0032] In one embodiment, there is provided a swellable, sustained release pharmaceutical composition comprising methylcellulose or a derivative thereof, hyaluronan or a derivative thereof, and an active ingredient, wherein at a physiological pH there is an electrostatic interaction between a positive charge of the active ingredient and negatively charged carboxylate ions of the hyaluronan or derivative thereof. In one embodiment, the composition is a swellable composition for the sustained, localized release of ropivacaine. In one embodiment, the composition is injectable.

[0033] In one embodiment, “sustained release” refers to a release that is prolonged compared to a non-sustained release formulation and, in particular, in the case of pharmaceutical compositions described herein, release of the same dosage of drug when administered in solution. The PK profile of a sustained release formulation will show a reduced Cmax with an AUC similar or greater than the non-sustained release formulation.

[0034] As used herein, “electrostatic” refers to an intermolecular interaction among two or more molecules, each of which has a positive or negative charge. Examples include, but are not limited to: ionic interactions, interactions between an ion and a polar molecule, interactions between partial charges of polar molecules, hydrogen bonds and induced partial charges of polarizable molecules.

[0035] The present inventors found that by increasing the concentration of HA, electrostatic interactions between HA-carboxylate (negatively charged) and an active ingredient having a positive charge and, in particular, bupivacaine HCI (positively charged) at physiological pH enabled an effective sustained release product (Figure 1A). This mechanism is demonstrated in Figure 1B. At pH 2, all of the carboxylate anions of HA are protonated to carboxylic acids while bupivacaine-HCI is still protonated. Thus at pH 2, the electrostatic interaction between polymer and active ingredient is lost because even though the active ingredient is still positively-charged, HA is now neutral. It is for this reason that the active ingredient diffuses out of the gel faster at pH 2 than at pH 7.4 (where HA is negatively charged).

[0036] Increasing the concentration of HA also increases the swelling of compositions containing HA. In certain applications, swellable pharmaceutical compositions are desirable. For example, swellable compositions may temporarily prevent adhesion between tissues or between tissues and implanted medical devices, facilitating the healing process. Pharmaceutical compositions with swelling properties can be useful for medical applications such as sealing, wound healing and drug delivery. For example, a swellable formulation that provides both pain relief and swelling could have applications for cosmetic surgery. An additional example could be following surgery where pain relief is needed in combination with prevention of tissue adhesion such as in myomectomy surgery.

[0037] However, swelling can cause the composition to become fragile or disassociate, losing its functionality or damaging surrounding tissue or biological processes. Increasing the concentration of HA in a pharmaceutical composition in order to provide sufficient negatively charged HA-carboxylate for sustained release of an active ingredient could be expected to cause excess swelling and these undesirable effects; however, the present inventors have identified compositions that are sufficiently stable to enable sustained release and controlled swelling. [0038] As used herein, “swellable” refers to a formulation that swells but does not completely disintegrate in an aqueous environment at a temperature of between 20°C and 38°C for a period of at least 24 hours. In one embodiment, for a period of at least 48 hours. In one embodiment, for a period of at least 72 hours

[0039] In various embodiments, “swellable” refers to a formulation that within 24 hours of administration increases in mass or volume by at least 50%, at least 75%, at least 100%, at least 150%, at least 200% or at least 300%.

[0040] In one embodiment, the hydrogel compositions as described herein are administered by injection, which may be via a needle or directly into a wound without use of a needle.

[0041] In one embodiment, the compositions as described herein are impregnated into or onto a carrier or substrate, such as a film, a web, or a woven or nonwoven sheet or fibres.

[0042] In one embodiment, the hydrogel composition as disclosed herein is formulated as a wound dressing.

[0043] In one embodiment, compositions as described herein are applied concurrently with a surgical procedure or in the post-operative surgical period. In one embodiment, the surgery is a dental surgery. In one embodiment, the surgery is a cosmetic surgery. In one embodiment, the surgery is a gynecological surgery.

[0044] In one embodiment, a single administered dose of a sustained release pharmaceutical composition as described herein provides localized anesthetic effects for a period of at least 12 hours, in one embodiment for at least 24 hours, in one embodiment for at least 48 hours, in one embodiment for at least 72 hours, in one embodiment for between 12 hours and 7 days, in one embodiment, for between 24 hours and 5 days.

[0045] In various embodiments, less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the local anesthetic agent is released from the pharmaceutical composition within 24 hours of administration. In various embodiments, less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the local anesthetic agent is released from the pharmaceutical composition within 48 hours of administration. In various embodiments, less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the local anesthetic agent is released from the pharmaceutical composition within 72 hours of administration.

[0046] Hyaluronic acid (or hyaluronan) (HA) and derivatives of HA may be employed. HA is a linear polysaccharide composed of repeating disaccharide units of N-acetyl-glucosamine and D-glucuronic acid. HA is degraded enzymatically by hyaluronidase, which can be produced by cells. Its polymeric chains, of lengths of 10-15 thousand disaccharides, form random coils with large spheroidal hydrated volumes of up to 400-500 nm in diameter. Reactions can occur at the carboxyl group or the hydroxyl group of HA and also at the amino group when the N-acetyl group is removed.

[0047] Pharmaceutical grade HA is available in a wide variety of molecular weights, in the range of between about 100,000 and about 3,000,000 g/mol. In one embodiment the composition comprises HA in the range of 500,000 and 2,500,000 g/mol, in one embodiment in the range of 1,000,000 and 2,000,000 g/mol, and in a preferred embodiment in the range of 1,400,000 to 1,600,000 g/mol.

[0048] Blends of unmodified HA with a gelling polymer are injectable upon an application of force to a syringe because the shear-thinning properties of HA cause the polymer chains to straighten and align themselves, permitting flow through the needle. HA then returns to its high viscosity, zero shear structure upon exiting the needle as the polymeric chains once again become entangled amongst themselves.

[0049] The other polymer component of the hydrogel is methylcellulose (MC). MC is an example of a temperature sensitive gel, or a thermally reversible gel, that gels upon an increase in temperature. When the degree of substitution of hydroxyl groups with methyl groups is between 1.4 and 1.9 per monomer unit, MC has inverse thermal gelling properties. As the temperature increases, the methyl groups of MC form hydrophobic interactions and water molecules are released from interacting with MC, thereby forming a gel. [0050] The MC may have a molecular weight in the range of between about 2,000 and about

1,000,000 g/mol. In one embodiment the composition comprises MC in the range of 10,000 and 500,000 g/mol, in one embodiment in the range of 100,000 to 400,000 g/mol, and in one embodiment in the range of 200,000 to 300,000 g/mol.

[0051] It is possible to alter the rate of resorption of the composition by changing the hydrophobicity of HA or the derivative thereof. More specifically, an altered rate of resorption may be provided by the addition of at least one functional group to the HA or the derivative thereof or the MC or the derivative thereof, selected from the group consisting of carboxylic acid, primary amine, aldehyde, hydrazide, maleimide, thiol, furan, alkyne, azide, alkene, urethane, and primary alcohol. [0052] As used herein, “bioresorbable compositions” are compositions that can be dispersed by biological processes so that the composition or a percentage thereof cannot be detected at the site of administration.

[0053] In various embodiments, bioresorbable composition refers to a composition wherein >50%, >60%, >70%, >80%, >90% or >99% of the composition cannot be detected at the site of administration at 28 days post-administration. In one embodiment, the composition cannot be detected at the site of administration at 28 days post-administration.

[0054] In various embodiments, bioresorbable composition refers to a composition wherein >50%, >60%, >70%, >80%, >90% or >99% of the composition cannot be detected at the site of administration at 14 days post-administration. In one embodiment, the composition cannot be detected at the site of administration at 14 days post-administration.

[0055] In various embodiments, bioresorbable composition refers to a composition wherein >50%, >60%, >70%, >80%, >90% or >99% of the composition cannot be detected at the site of administration at 7 days post-administration. In one embodiment, the composition cannot be detected at the site of administration at 7 days post-administration. [0056] In various embodiments, bioresorbable composition refers to a composition wherein >50%, >60%, >70%, >80%, >90% or >99% of the composition cannot be detected at the site of administration at 3 days post-administration. In one embodiment, the composition cannot be detected at the site of administration at 3 days post-administration.

[0057] In various embodiments, bioresorbable composition refers to a composition wherein >50%, >60%, >70%, >80%, >90% or >99% of the composition cannot be detected at the site of administration at 1 day post-administration. In one embodiment, the composition cannot be detected at the site of administration at 1 day post-administration.

[0058] The amount of composition at the site of administration may be detected by mass as would be determined by a person skilled in the art.

[0059] In one embodiment, the active ingredient is an amide active ingredient, examples of which include articaine, bupivacaine, cinchocaine dibucaine, etidocaine, levobupivacaine, lidocaine, mepivacaine, oxetacaine, prilocaine, ropivacaine, sameridine, tolycaine, tonicaine and trimecaine and pharmaceutically acceptable salts thereof, i.e. salts that retain the anesthetic activity of these compounds and do not impart undesired toxicological effects.

[0060] Of the active ingredients, bupivacaine and ropivacaine are of particular interest due to their potency and widespread clinical use. Both are similar structurally and functionally, differing only by a single methyl group; however, bupivacaine is a racemic mixture of R- and S- enantiomers, whereas ropivacaine is the enantiomerically pure S-enantiomer. Functionally, bupivacaine is more potent and ropivacaine is typically administered at higher doses. However, even at equipotent concentrations, bupivacaine is also more toxic than ropivacaine. As ropivacaine is less lipophilic than bupivacaine, it is less likely to penetrate large myelinated motor fibres, and it consequently has a more selective action on the pain-transmitting Ad or C nerve fibres, rather than the Ab fibres that are involved in motor function.

[0061] Ropivacaine is a promising and effective analgesic and would be of even greater value if its efficacy could be extended through sustained release. Various strategies to prolong the efficacy of ropivacaine have been investigated, including encapsulation of the drug within liposomes, microparticles or nanoparticles. While these strategies extend release and offer advantages over a liquid bolus injection, the particles do not remain localized and thus can be transported away from the surgery site, resulting in systemic side effects. [0062] High doses of bupivacaine remain a concern clinically as they have been associated with dose-dependent cardiotoxicity, neurotoxicity and myotoxicity [Lirk, P., Picardi, S., and Hollmann, M.W., Local anaesthetics: 10 essentials. Eur J Anaesthesiol, 2014. 31(11): p. 575-85; Werdehausen, R., et al. , Apoptosis induction by different local anaesthetics in a neuroblastoma cell line. British journal of anaesthesia, 2009. 103(5): p. 711-718; Mulroy, M.F., Systemic toxicity and cardiotoxicity from local anesthetics: incidence and preventive measures. Reg. Anesth. Pain Med. 2002. 27(6): p. 556-61; Takenami, T., et al., Neurotoxicity of intrathecally administered bupivacaine involves the posterior roots/posterior white matter and is milder than lidocaine in rats. Regional anesthesia and pain medicine, 2005. 30(5): p. 464-472.; Sakura, S., et al., The comparative neurotoxicity of intrathecal lidocaine and bupivacaine in rats. Anesthesia & Analgesia, 2005. 101(2): p. 541-7] By comparison, ropivacaine has been found to reduce cardiovascular and neurologic complications at equipotent concentrations. The improved safety profile of ropivacaine has been attributed to its stereoselectivity and reduced lipophilicity relative to bupivacaine. Moreover, ropivacaine is metabolized by both cytochrome P50 (CYP) 1A2 and CYP3A4, while bupivacaine is metabolized mainly by CYP3A4. This further improves the safety profile for ropivacaine as the CYP3A4 bupivacaine receptor is common to many other drugs that patients may have been prescribed. Co-administration of these drugs with ropivacaine and bupivacaine may result in adverse pharmacokinetic interactions due to competition for CYP3A4; however, as ropivacaine is also largely metabolized by CYP1A2, adverse consequences are less likely to occur.

[0063] In one embodiment, the active ingredient is lidocaine, bupivacaine, ropivacaine or a pharmaceutically acceptable salt of any of the foregoing. In one embodiment, the active ingredient is ropivacaine.

[0064] Pharmaceutical compositions as described herein may suitably be prepared through the physical blending of HA and MC in phosphate buffered saline (PBS). After MC and HA are dispersed in PBS and allowed to dissolve, the local anesthetic, suitably in particle form, may be dispersed in HAMC. The compositions may be sterilized by autoclave, gamma sterilization, filter sterilization or steam sterilization. Compositions are suitably stored at a range of 4°C to room temperature (25°C).

[0065] The pharmaceutical compositions described herein are injectable, wherein injection may be, for example, by syringe, via a catheter or other device for delivering a liquid material across the skin such as by microjet (see e.g. United States patent 8,369,942, incorporated by reference herein in its entirety). Alternatively, the composition may be administered by injection by ejecting the material from a syringe without a needle, topically, or into an open wound in some embodiments. When administered via injection, the composition can operate as a depot injection, the composition forming a localized mass. In one embodiment the composition is administered by a single injection. The pharmaceutical compositions as described herein may be administered in a number of ways depending upon the area to be treated. Without limiting the generality of the foregoing, in a particular embodiment, the compositions are administered by subcutaneous, intradermal or intramuscular injection.

[0066] In one embodiment, the pharmaceutical composition is administrable with a 10-30 gauge needle, in one embodiment, a 20-25 gauge needle, in one embodiment, without a needle.

[0067] The pharmaceutical compositions described herein may be combined with any pharmaceutically acceptable carrier or excipient. As used herein, a “pharmaceutically acceptable carrier” or “excipient” can be a pharmaceutically acceptable solvent, suspending agent or any other pharmacologically inert vehicle selected to facilitate delivery of the pharmaceutical composition to a subject. The excipient may be liquid or solid and is selected, with the planned manner of administration in mind, so as to provide for the desired bulk, consistency, etc., when combined with the other components of the pharmaceutical composition. Examples of pharmaceutically acceptable carriers include one or more of water, saline, phosphate buffered saline, glycerol, ethanol and the like, as well as combinations thereof. Pharmaceutically acceptable carriers may further comprise minor amounts of auxiliary substances such as wetting or emulsifying agents, preservatives or buffers, which enhance the shelf life or effectiveness of the pharmacological agent.

[0068] In some embodiments, the pharmaceutically acceptable carrier is phosphate buffered saline or saline.

[0069] The pharmaceutical composition as described herein may conveniently be presented in unit dosage form of a single-use syringe that has been sterilized for injection with or without a needle.

[0070] In one embodiment, the pharmaceutical composition is 5.0 to 9.0 wt% HA and 1.0 to 5.0 wt% MC. The pharmaceutical composition is loaded into a syringe and sterilized using steam. The pharmaceutical composition volume can vary from 1 ml_ to 20 ml_; and the syringe size can vary from 1 ml_ to 20 ml_. [0071] As used herein, “therapeutically effective amount” refers to an amount effective, at dosages and for a particular period of time necessary, to achieve the desired therapeutic result. A therapeutically effective amount of the pharmacological agent may vary according to factors such as the disease state, age, sex, and weight of the individual, and the ability of the pharmacological agent to elicit a desired response in the individual. A therapeutically effective amount is also one in which any toxic or detrimental effects of the pharmacological agent are outweighed by the therapeutically beneficial effects.

[0072] As used herein “subject” refers to an animal being administered a local anesthetic, in one embodiment a mammal, in one embodiment a human patient.

[0073] As used herein “treatment” and grammatical variations thereof refers to administering a compound or composition of the present invention, in one embodiment in order to provide localized pain relief. This treatment may be to alleviate pain or the use may be prophylactic to prevent pain. In one embodiment in order to aid wound healing. In one embodiment to partially or fully seal biological tissue. The treatment may require administration of multiple doses, which may be at regular intervals.

[0074] In one embodiment, there is provided a method of treating or preventing pain comprising administering, preferably by injection, a therapeutically effective amount of a pharmaceutical composition as described herein.

[0075] In one embodiment, there is provided a method of treating or preventing pain and preventing surgical adhesions comprising administering, preferably by injection, a therapeutically effective amount of a pharmaceutical composition as described herein.

[0076] In one embodiment, there is provided a method of aiding wound healing comprising administering, preferably by injection, a therapeutically effective amount of a pharmaceutical composition as described herein.

[0077] In one embodiment, there is provided a method of partially or fully sealing biological tissue comprising administering, preferably by injection, a therapeutically effective amount of a pharmaceutical composition as described herein.

[0078] All documents referenced herein are incorporated by reference, however, it should be appreciated that any patent, publication, or other disclosure material, in whole or in part, that is incorporated by reference herein is incorporated only to the extent that the incorporated material does not conflict with definitions, statements, or other disclosure material set forth in this disclosure. As such, and to the extent necessary, the disclosure as explicitly set forth herein supersedes any conflicting material incorporated herein by reference.

[0079] It will be understood that numerous modifications thereto will appear to those skilled in the art. Accordingly, the above description and accompanying drawings should be taken as illustrative of the invention and not in a limiting sense. It will further be understood that it is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departures from the present disclosure as come within known or customary practice within the art to which the invention pertains and as may be applied to the essential features herein before set forth, and as follows in the scope of the appended claims.

[0080] The embodiments of the invention described above are intended to be exemplary only. The scope of the invention is therefore intended to be limited solely by the scope of the appended claims.

EXAMPLE 1 - Sterilization of materials and preparation of HAMC gels

[0081] 1 g of HA and 1 g of MC were each separately dissolved in 500 mL of MilliQ water overnight with stirring at 4°C. HA was sterilized by autoclave and MC was sterile-filtered through a 0.2 pm pore size filter. The sterile HA and MC were snap frozen in liquid nitrogen and lyophilized. Bupivacaine-hydrochloride was sterilized by gamma radiation at 2.5 MRad. Sterile HA and MC were dissolved in artificial cerebrospinal fluid (aCSF) to which bupivacaine hydrochloride was added and then mixed using a dual asymmetric centrifugal mixer for 30 seconds.

EXAMPLE 2 - Effect of HA concentration on release of bupivacaine hydrochloride

[0082] Sterile HA and MC were dissolved in artificial cerebrospinal fluid (aCSF) to which bupivacaine hydrochloride was added and then mixed using a dual asymmetric centrifugal mixer for 30 seconds. Given that, at physiological pH, bupivacaine hydrochloride is positively charged, HA is negatively charged, and MC is neutral, to control the release of bupivacaine hydrochloride, the concentration of HA in the HAMC formulation was increased from 1.4 wt% to 7.0 wt% while maintaining MC concentration constant at 3.0 wt%.A 100 pL volume of each HAMC-bupivacaine hydrochloride formulation was aliquoted into 2 mL Eppendorf microcentrifuge tubes. To ensure a planar geometrical surface, the microcentrifuge tubes were spun for 10 seconds at 5000 rpm. Samples were placed in the 37°C incubator for 30 minutes to allow the hydrogel to gel. At time zero (t=0), 1.9 ml_ of pre-warmed (37°C) aCSF was placed on top of the hydrogel and, at specific time points, the aCSF was completely removed, stored and replaced with an equal volume of fresh aCSF. Once all the samples were collected, they were measured and quantified using a UV-Vis spectrophotometer at 210 nm. The absorbance was compared to a standard curve in order to determine the mass of released bupivacaine hydrochloride. For mass balance, at the terminal time point, 1.5 ml_ of acetonitrile was added to the hydrogel and left at 4°C overnight on a shaker to extract any remaining bupivacaine hydrochloride.

[0083] As shown in Figure 1A, a prolonged rate of bupivacaine hydrochloride release was observed with increased HA concentration, suggesting an electrostatic interaction between bupivacaine hydrochloride and HA that controls the release. At all timepoints studied, there was a significant difference in the bupivacaine hydrochloride released between 7:3 HAMC (7.0 wt% HA, 3.0 wt% MC) and 1.4:3 HAMC (1.4 wt% HA, 3.0 wt% MC).

EXAMPLE 3 - Effect of pH on the release of bupivacaine hydrochloride from HAMC

[0084] To verify that the slower release profile of bupivacaine hydrochloride observed with higher concentrations of HA was due to an electrostatic interaction between the negatively- charged carboxylate of the HA and the positively-charged drug, a separate release was performed under acidic conditions (pH 2) and compared to physiological conditions (pH 7.4). At pH 2, the HA carboxylate anion is protonated to carboxylic acid, thereby neutralizing the negative charge on HA. Both HAMC and release media were prepared with acidic aCSF. As shown in Figure 1 B, the electrostatic interaction between HA and bupivacaine HCI was lost, resulting in faster release.

EXAMPLE 4 - Swelling ratio of HAMC combined with bupivacaine

[0085] To further characterize bupivacaine-loaded HAMC, the swelling ratio was determined. The mass of each respective tube was pre-weighed, 180 mg of HAMC or drug-loaded HAMC was added to each tube and contents were spun down to the bottom. The mass of HAMC at time zero was recorded after adding and removing 1620 pL of aCSF. Fresh aCSF was replaced on top of the hydrogel until the next time point, where as much aCSF was removed and the total mass of the tube and HAMC was measured. Time points investigated were 30 minutes, 1 hour, 3 hours, 6 hours, 1 day, 2 days, 3 days, 4 days, 6 days, 8 days and 10 days. The swelling ratio (Q) produces a number that corresponds to the normalized amount of aCSF absorbed by the original mass of hydrogel: Amount of aCSF absorbed M_j-Mf

Q = Original mass of hydrogel at time zero M₀-M_t

[0086] As shown in Figure 1C, 7:3 HAMC was found to swell and the swelling ratio increased further with the addition of bupivacaine base.

EXAMPLE 5 - Spinal nerve block [0087] Female Sprague Dawley rats weighing between 250-350 g were used to assess the pain model and the effect of drug-loaded HAMC placed onto the injured site. Under isoflurane- oxygen anesthesia, an incision was made and the paraspinal muscle was retracted laterally, exposing the facet joint and interlaminar space. A partial unilateral laminotomy and medial facetectomy was carried out to reveal the left L5 spinal nerve root. [0088] Animals were treated with 50 pL bupivacaine hydrochloride-loaded 7:3 HAMC at doses of 1.56, 5, 10, 25 or 34 mg/kg. Control animals were treated with HAMC only or bupivacaine hydrochloride in solution at 1.56 mg/kg. Pain responses were monitored over 3 days using von Frey filaments and a cold plantar test. Animal were sacrificed at 7 days and the tissue at the site of gel infiltration collected. Sensory block evaluation

[0089] Sensory function of the spinal nerve was evaluated using a von Frey assay and a cold plantar assay. These behavioral tests assess fine touch, and temperature and pain, respectively, and have been used extensively [Pitcher, G.M., J. Ritchie, and J.L. Henry, Peripheral neuropathy induces cutaneous hypersensitivity in chronically spinalized rats. Pain Med, 2013. 14(7): p. 1057- 71 ; Jasmin, L, et al., The cold plate as a test of nociceptive behaviors: description and application to the study of chronic neuropathic and inflammatory pain models. Pain, 1998. 75(2-3): p. 367- 82] Von Frey filaments were used to evaluate tactile allodynia and the cold plantar assay to evaluate the response to a thermal stimulus on the plantar surface of the foot. Prior to the surgical procedure, a baseline for every animal using each assay was conducted. In the spinal nerve model, pain responses were monitored at 2, 6, 12, 18, 24, 48 and 72 hours post-operation. Von Frey assay

[0090] For the von Frey assay, the 50% withdrawal threshold for mechanical allodynia was measured. Briefly, animals were placed in individual enclosures on top of a wire mesh and allowed to acclimatize for 20 minutes. Next, von Frey filaments of increasing stiffness (6, 8, 10, 15, 26, 60, 100, 180, 300 g) were applied to the mid-plantar region of the hind paw for three seconds per application, either for a maximum of six applications or until the animal sharply withdrew the tested paw three times. If three responses were not observed upon application of the 300 g filament, a value of 300 g was recorded; this was defined as a complete block. The response from the contralateral uninjured paw was used as an internal control.

Cold plantar assay

[0091] Animals were placed in individual enclosures on top of a glass plate made from transparent plexiglass and allowed to acclimatize for 10 minutes. To make the cold probe, dry ice was crushed into a fine powder and packed into a dense pellet with a flush surface in a custom syringe. Using the plunger, the compressed ice pellet was applied against the glass, directly underneath the mid-plantar region of the hind paw. The time for the animal to sharply withdraw its paw was recorded. An interval of at least 2 minutes was given between the testing of the same paw, and a cut-off of 30 seconds was given to avoid potential tissue damage. If a response was not observed by 30 seconds, the value was recorded as 30 seconds and was defined as a complete block. Both hind paws were tested twice at every time point.

Extended bupivacaine hydrochloride release in HAMC and spinal nerve block in vivo

[0092] The in vivo efficacy of bupivacaine hydrochloride-loaded HAMC was investigated in a rat L5 spinal nerve block model. A complete block for the von Frey assay was defined as a non responder to the application of a 300 g filament, while a complete block for the cold plantar assay was defined as non-responder to 30 seconds of cold application. As shown in Figure 2A, higher doses of bupivacaine hydrochloride-loaded HAMC yielded longer complete mechanical blocks, with a 34 mg/kg dose in HAMC lasting 22.8 ± 2.7 h vs. a 1.56 mg/kg dose in HAMC lasting 2.0 ± 0.0 h. At an equivalent 1.56 mg/kg dose of bupivacaine hydrochloride in solution, the complete mechanical block was only 0.67 ± 1.0 h; however, due to the very large standard deviation, the difference between HAMC and solution delivered bupivacaine hydrochloride at 1.56 mg/kg was not significant. As shown in Figure 2B, the extended nerve block was corroborated by the cold plantar assay, where higher HAMC doses similarly led to longer complete thermal blocks, with a 34 mg/kg dose lasting 21.6 ± 3.3 h vs. a 1.56 mg/kg dose lasting 2.0 ± 0.0 h. Similarly, dose matched bupivacaine hydrochloride delivered in solution did not last as long as that delivered in HAMC; however the difference was not significant due to the large standard deviation of solution- released bupivacaine hydrochloride. The HAMC delivery system, in addition to providing sustained sensory effects, yielded a local effect as no mechanical or thermal blocks were observed on the contralateral paw.

EXAMPLE 6 - Sciatic nerve block model

[0093] Male Sprague Dawley rats weighing between 250-350 g were used in the study. Under isoflurane-oxygen anesthesia, the rats were shaved and placed in a right decubitus position. To avoid the possibility of nerve damage from the needle during injection, the right sciatic nerve was exposed using a mini-open technique at proximal thigh level. A 1 cm incision was made under the femoral head, the bicep femoris muscle was split parallel to the longitudinal of the muscle fibers to avoid injury, and the sciatic nerve was identified. A cotton swab was inserted between the muscle layers around the nerve to gently create a pocket into which the HAMC-bupivacaine hydrochloride (or other solutions) was injected using a 20 gauge needle. The following groups were compared with a 400 pL injection of: 8 mg/kg bupivacaine hydrochloride in HAMC; 32 mg/kg bupivacaine hydrochloride in HAMC; 64 mg/kg bupivacaine hydrochloride in HAMC; HAMC alone; aCSF alone; or conventional bupivacaine hydrochloride solution at 10 mg/kg.

Sensory block evaluation [0094] Sensory function of the sciatic nerve was evaluated with von Frey monofilaments, the cold plantar assay, and a noxious pinch of the foot. The pinch test was performed on the first and fifth toe to evaluate reflex to a noxious stimulus. Prior to the surgical procedure, a baseline for every animal using each assay was conducted. In the sciatic nerve model, sensation was evaluated at 0.5, 1 , 4, and 7 hours and then every 3 hours thereafter post-injection until sensation returned to baseline.

Pinch assay

[0095] Animals were placed in individual enclosures and allowed to acclimatize for 10 minutes. Using toothed forceps, the first and fifth toe of each foot were pinched. A positive response consisted of a sharp reflexive twitching of the foot. Each hind paw was tested three times, with an interval of at least 2 minutes between each trial. A complete block was defined when two out of three pinches had no responses.

Tissue harvesting and histology

[0096] After euthanasia with carbon dioxide at 7 days, the sciatic nerve and surrounding muscle tissue were collected, and samples were stained with hematoxylin and eosin. Specimens were evaluated for neurotoxicity and myotoxicity by a trained histologist, who was blind to the treatment groups.

Extended bupivacaine release in HAMC provides prolonged sciatic nerve block in vivo

[0097] Using male Sprague Dawley rats, bupivacaine hydrochloride was delivered in either saline or HAMC by injection on the sciatic nerve, which is located at the sciatic notch under the greater trochanter bone. To test efficacy, the animals were tested by: von Frey assay for mechanical stimulus, pinch test for sensory pain, and cold plantar assay for thermal sensation. Similar to the spinal nerve root block, HAMC formulations significantly extended the duration of efficacy (Figure 3 A-C)

Tissue reaction

[0098] The sciatic nerves and surrounding muscle tissue were removed 7 days post treatment and processed for histology. A trained histologist, blinded to the sample groups, evaluated the tissue reaction. As shown in Figure 4, all muscle specimens treated with bupivacaine hydrochloride-loaded HAMC were comparable to either saline or bupivacaine hydrochloride solution controls. Following treatment with saline or HAMC at 8mg/kg, muscle tissue appeared normal and there were no remarkable inflammatory or degenerative features. As shown in Figure 4A to 4E, while some degenerate muscle fibers were present following treatment with bupivacaine hydrochloride solution, and bupivacaine hydrochloride delivered in HAMC at 32 mg/kg and 64 mg/kg, these were interspersed with regenerative muscle fibres. Further, while at 64 mg/kg in HAMC, the biceps femoris and the adductor magnus muscles beside the site of injection showed some necrosis at post-operative day 4, the tissue recovered by post-operative day 28. This result was comparable to tissue treated by bupivacaine hydrochloride solution at the much lower dose of 10 mg/kg.

[0099] Sciatic nerve specimens embedded in the surrounding tissue were also evaluated by the trained histologist. As shown in Figure 4F to 4J, similarly, all nerves treated with HAMC were comparable to either vehicle or bupivacaine hydrochloride solution controls. Following treatment with bupivacaine hydrochloride solution, the peripheral nerves were unremarkable in appearance. Following treatment with saline, and HAMC at 8mg/kg, 32mg/kg and 64mg/kg, the nerves appeared unremarkable, where mild inflammation and no overt degeneration of the surrounding tissue was present.

Claims

WHAT IS CLAIMED IS:

1. A swellable, sustained release pharmaceutical composition comprising: a hydrogel comprising a gel polymer matrix comprising 3 to 10 wt% hyaluronan or a derivative thereof and 1 to 9.99 wt% methylcellulose or a derivative thereof based on the weight of the hydrogel, wherein the amount of hyaluronan or derivative thereof is greater than the amount of methylcellulose or derivative thereof, and an active ingredient, wherein at a physiological pH the active ingredient or a portion thereof is positively charged.

2. The pharmaceutical composition of claim 1, wherein the ratio of hyaluronan:methylcellulose is between 1.01:1 and 3:1.

3. The pharmaceutical composition of claim 1 or 2, wherein the composition is injectable.

4. The pharmaceutical composition of any one of claims 1 to 3, wherein the active ingredient is an amide active ingredient.

5. The pharmaceutical composition of claim 4, wherein the active ingredient is lidocaine, bupivacaine, ropivacaine and/or a derivative or pharmaceutically acceptable salt thereof.

6. The pharmaceutical composition of claim 5, wherein the active ingredient is ropivacaine and/or a pharmaceutically acceptable salt thereof.

7. The pharmaceutical composition of any one of claims 1 to 6, wherein the active ingredient is hydrophobic in some portion.

8. The pharmaceutical composition of any one of claims 1 to 7 wherein less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the active ingredient is released from the pharmaceutical composition within 12 hours of administration.

9. The pharmaceutical composition of any one of claims 1 to 7 wherein less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the active ingredient is released from the pharmaceutical composition within 24 hours of administration.

10. The pharmaceutical composition of any one of claims 1 to 7 wherein less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10% or less than 5% of the active ingredient is released from the pharmaceutical composition within 48 hours of administration.

11. The pharmaceutical composition of any one of claims 1 to 10 wherein the hyaluronan or derivative thereof has a molecular weight between 100,000 g/mol and 3,000,000 g/mol and the methylcellulose or derivative thereof has a molecular weight between 10,000 g/mol and 500,000 g/mol

12. A dosage form comprising between 1 ml_ and 100 ml_ of the pharmaceutical composition of any one of claims 1 to 11.

13. The dosage form of claim 12 comprising between 100 mg and 2000 mg of the active ingredient.

14. The dosage form of claim 12 comprising between 350 mg and 2000 mg of the active ingredient.

15. The dosage form of claim 12 comprising between 750 mg and 1500 mg of the active ingredient.

16. A method of treating or preventing pain comprising administering a therapeutically effective amount of the pharmaceutical composition of any one of claims 1 to 11 or a dosage form according to any one of claims 12 to 15 to a subject in need thereof.

17. A method of aiding wound healing comprising administering a therapeutically effective amount of the pharmaceutical composition of any one of claims 1 to 11 or a dosage form according to any one of claims 12 to 15 to a subject in need thereof.

18. A method of partially or fully sealing biological tissue comprising administering a therapeutically effective amount of the pharmaceutical composition of any one of claims 1 to 11 or a dosage form according to any one of claims 12 to 15 to a subject in need thereof.