DESC:FORM-2

THE PATENTS ACT, 1970
(39 OF 1970)
COMPLETE SPECIFICATION
(See section 10; rule 13)

OPHTHALMIC COMBINATION COMPOSITION

SUN PHARMACEUTICAL INDUSTRIES LIMITED

A company incorporated under the laws of India having their office at Sun House, 201 B/1, Western Express Highway, Goregaon (E), Mumbai-400063 Maharashtra, India.

The following specification particularly describes the invention and the manner in which it is to be performed.

FIELD OF INVENTION
The present invention is related to a quaternary ammonium compound free ophthalmic combination composition comprising of a carbonic anhydrase inhibitor and beta-adrenergic receptor blocker drug. The said composition is for decreasing of intraocular pressure (IOP) in adult patients with open-angle glaucoma or ocular hypertension for whom monotherapy provides insufficient IOP reduction.

BACKGROUND OF INVENTION
Elevated intraocular pressure (IOP) is one of the major risk factors of glaucoma, and decreased IOP can be associated with certain eye problems such as retinal detachment and uveitis. In children, high IOP can lead to corneal enlargement, tears in the Descemet's membrane, and corneal edema. In glaucoma management, percentage reduction in IOP is one of the important indices for different treatment protocols. Hasan Et al., Distribution of intraocular pressure and its determinants in an Iranian adult population; Int J Ophthalmol. 2016; 9(8): 1207–1214.
Thus, IOP control mechanisms have extensively been investigated in recent years. Studies also show that many systemic, ocular, and even biometric indices may be correlated with. Other major risk factors include age, sex, alcohol consumption, smoking, and family history of elevated IOP which have been reported to affect the distribution of IOP.
Commonly prescribed drugs for IOP reduction includes prostaglandin analogues such as Latanoprost, Bimatoprost, Travoprost; ß-blockers such as Timolol; Carbonic anhydrase inhibitors such as Brinzolamide, Dorzolamide and a-agonists drugs such as Brimonidine. Various drugs have been approved and used in varying permutations starting from single drug therapy to combination or multiple drug therapy, some of the examples include: Xalatan® (Latanoprost), Xalacom® (Latanoprost and timolol), Lumigan® (Bimatoprost), Duotrav® (Travoprost and timolol). Such approved products mostly use Benzalkonium chloride as surfactant or preservative.
However most formulation use aggressive preservatives or surfactants such as quaternary ammonium compounds, which makes there use for a long term hazardous to the patient population. Thereby leading either to worsening of associated side effects or discontinuation of therapy. One such technical problem identified by the present inventors, particularly associated with alkyl quaternary ammonium preservatives or compounds having surfactant like property such as benzalkonium chloride, benzdodecinium chloride and like and mixtures thereof.
Alkyl quaternary ammonium preservatives or surfactant such as Benzalkonium chloride leads to clinically adverse events like eye irritation, also known to discolour soft contact lenses. Benzalkonium chloride in eye drops has also been reported to cause punctate keratopathy and/or toxic ulcerative keratopathy, especially in frequent or prolonged use or in conditions where the cornea is compromised. EMEA public statement on antimicrobial preservatives in ophthalmic preparations for human use (EMEA/622721/2009). Benzalkonium chloride inhibits mitochondria of human corneal epithelial cells and cells bearing LHON mutations at pharmacologically relevant concentrations, and we suggest this is the basis of BAK's ocular toxicity. Prescribing BAK-containing eye drops should be avoided in patients with mitochondrial deficiency, including LHON patients, LHON carriers, and possibly primary open-angle glaucoma patients. Sandipan Datta Et al., The Eye Drop Preservative Benzalkonium Chloride Potently Induces Mitochondrial Dysfunction and Preferentially Affects LHON Mutant Cells, Invest Ophthalmol Vis Sci. 2017 Apr; 58(4): 2406–2412.
Despite all the side effects and associated toxicity in order to maintain sterility and prevent ocular infections from a contaminated eye formulations like drop, or suspension the addition of preservatives is mandatory in multi-dose ophthalmic formulations according to international standards. Of the many available preservatives, benzalkonium chloride (BAK), a quaternary ammonium salt, is the most common preservative used in eye drops till date.
U.S. Patent No. 6,071,904 discloses ophthalmic suspensions containing brinzolamide, tyloxapol, carbomer 974P, edetate disodium, benzalkonium chloride, mannitol, sodium chloride and purified water. U.S. Patent No. 9,044,484 and U.S. Patent No. 9,421,265 disclose multi-dose ophthalmic compositions containing brinzolamide, brimonidine, two or more different polyols in conjunction with borate and a low concentration of benzalkonium chloride.
Management of open angle glaucoma and ocular hypertension require long term treatment with eye drops containing preservatives such as benzalkonium chloride. Symptoms and signs of ocular surface disease such as ocular surface breakdown, irritation, burning, foreign body sensation, dryness, inadequate quantity of tears, etc. are prevalent in a large proportion of patients with open angle glaucoma and ocular hypertension. Further Patients experiencing hypersensitivity reactions with benzalkonium chloride should minimize use of commercially available brimonidine and timolol products containing benzalkonium chloride which is preserved even with 0.005% w/v benzalkonium chloride.
So there is a significant need for an alkyl quaternary ammonium compounds free ophthalmic formulation for reducing the intraocular pressure (IOP) in varying eye condition. Further such formulation has to be stable and effective without compromising on the sterility of the product.

SUMMARY OF INVENTION
The present invention provides a quaternary ammonium compound free ophthalmic combination composition comprising of a carbonic anhydrase inhibitor drug and beta-adrenergic receptor blocker drug for decreasing of intraocular pressure (IOP) in adult patients with open-angle glaucoma or ocular hypertension.
The present invention provides a quaternary ammonium compound free ophthalmic combination composition comprising of a carbonic anhydrase inhibitor drug and beta-adrenergic receptor blocker drug, with potassium sorbate in a pharmaceutically acceptable vehicle for decreasing of intraocular pressure (IOP) in adult patients with open-angle glaucoma or ocular hypertension.
In another embodiment the present invention provides an ophthalmic composition which is a suspension comprising carbonic anhydrase inhibitor drug and a beta-adrenergic receptor blocker drug, wherein the composition is free of quaternary ammonium compound.
In yet another embodiment the present invention provides an ophthalmic combination composition for treatment of elevated intraocular pressure in patients with ocular hypertension, the composition comprising carbonic anhydrase inhibitor drug and a beta-adrenergic receptor blocker drug in a therapeutically effective amount.
In another embodiment the present invention provides an ophthalmic combination composition for treatment of elevated intraocular pressure in patients with ocular hypertension, the composition comprising carbonic anhydrase inhibitor drug and a beta-adrenergic receptor blocker drug in a therapeutically effective amount along with potassium sorbate in a pharmaceutically acceptable vehicle.
In some embodiment the present invention provides a method of treating elevated intraocular pressure in patients by administering an ophthalmic combination composition comprising carbonic anhydrase inhibitor drug and a beta-adrenergic receptor blocker drug in a therapeutically effective amount along with potassium sorbate in a pharmaceutically acceptable vehicle.
In a further embodiment the present invention provides a method of reducing ocular hypertension or intraocular pressure by administering a combination composition comprising a carbonic anhydrase inhibitor drug, a beta-adrenergic receptor blocker drug and potassium sorbate, wherein the composition is characteristically free of quaternary ammonium compound.
In some embodiment the ophthalmic combination composition according to present invention is free of quaternary ammonium compound such as alkyl quaternary ammonium compound selected from benzalkonium chloride, benzdodecinium chloride and like and mixtures thereof.
In one embodiment the present invention provides an ophthalmic combination composition for lowering intraocular pressure in a patient comprising administering the composition comprising of brinzolamide or its pharmaceutically acceptable salts, timolol or its pharmaceutically acceptable salt, with one or more pharmaceutically acceptable excipients, wherein the composition is free of benzalkonium chloride.
In another embodiment the ophthalmic combination composition according to present invention comprises brinzolamide or a pharmaceutically acceptable salt thereof, timolol or a pharmaceutically acceptable salt thereof, along with one or more pharmaceutically acceptable excipients selected from buffers, viscosity modifying agents, tonicity adjusting agents, chelating agents, preservatives, polymer, surfactants, solvents, co-solvents, pH adjusting agents and/or combination thereof.

BRIEF DESCRIPTION OF DRAWINGS
Figure-1: Showing a comparative intraocular pressure (IOP) reduction of the present BKC free test formulation to that of Brinzotim®.

DESCRIPTION OF INVENTION
The terms “for example” and “such as,” and grammatical equivalences thereof, the phrase “and without limitation” is understood to follow unless explicitly stated otherwise. As used herein, the term “about” is meant to account for variations due to any experimental errors which may be commonly accepted in the analytical chemistry field. All measurements reported herein are understood to be modified by the term “about,” whether or not the term is explicitly used, unless explicitly stated otherwise. As used herein, the singular forms “a”, “an” and “the” include plural referents unless the context clearly dictates otherwise. For the values provided herein, the term “about” indicates a given number may vary by at least ±10%.
The terms “composition”, “formulation”, as used herein are interchangeably used to define a pharmaceutical composition according to present invention.
The term “buffered vehicle” or “pharmaceutically acceptable vehicle” as used herein are used interchangeably and means an aqueous or non-aqueous vehicle which comprises one or more of tonicity adjusting agent, buffering agents, chelating agents, viscosity modifiers, chelating agents, polymers as suspending agents or penetration enhancers, surfactants and their combinations.
The term “improved efficacy” as used herein is defined by therapeutic efficacy in terms of reduced side effects or reduction in toxicity or side effects associated with long term use of the quaternary ammonium compounds based preservative containing compositions for reduction in intraocular pressure or used for treatment of glaucoma.
The term beta-adrenergic receptor antagonists or beta-blockers as used herein are used interchangeably and includes beta blocker drugs.
Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Methods and materials are described herein for use in the present disclosure; other suitable methods and materials known in the art can also be used. The materials, methods and examples are illustrative only and not intended to be limiting by any means. All publications, patent applications, patents and other references mentioned herein are incorporated by reference in their entirety. In case of a conflict, the present specification, including definitions, will control.
The inventors surprisingly found that ophthalmic composition for reducing IOP can be prepared without including alkyl quaternary ammonium compounds like benzalkonium chloride, benzdodecinium chloride and like and mixtures thereof. Thus, the present invention provides an ophthalmic composition comprising carbonic anhydrase inhibitor with beta-adrenergic receptor blocker which contains dual benefit of improved efficacy and avoidance of undesirable effects of the alkyl quaternary ammonium compounds. Therefore the present invention provides an ophthalmic suspension which have a stability, efficacy, but whose antimicrobial efficacy is not compromised despite being free of alkyl quaternary ammonium preservatives.
The present invention relates to an ophthalmic composition for reducing intraocular pressure, the said composition is a combination of carbonic anhydrase inhibitor and beta-adrenergic receptor blocker and the said composition is free of alkyl quaternary ammonium compounds.
In one embodiment the present invention provides an ophthalmic composition comprising of carbonic anhydrase inhibitor drug and beta-adrenergic receptor blocker in a pharmaceutically acceptable vehicle.
In one embodiment the ophthalmic combination composition according to present invention comprises carbonic anhydrase inhibitor drug and beta-adrenergic receptor blocker, viscous polymers as suspending agent, non-ionic surfactants and stabilizers.
In another embodiment the present invention provides an ophthalmic composition comprising of carbonic anhydrase inhibitor drug, beta-adrenergic receptor blocker and potassium sorbate in a pharmaceutically acceptable vehicle.
In some embodiment the ophthalmic composition according to present invention is a multi-dose suspension without Benzalkonium chloride preservative.
The term “multi-dose” as used herein is defined as a composition / formulation wherein the composition can be administered multiple times from the same container despite being devoid of quaternary ammonium compound such as Benzalkonium chloride.
In some embodiment the carbonic anhydrase inhibitor drug is selected from brinzolamide, acetazolamide, methazolamide, dorzolamide, diclofenamide, ethoxzolamide, and zonisamide or a combination thereof.
In some embodiment the beta-adrenergic receptor blocker drug is selected from timolol, propranolol, nadolol, pindolol, labetalol, penbutolol, sotalol, carvedilol, metoprolol, Toprol, atenolol, acebutolol, betaxolol, esmolol, bisoprolol and nebivolol or a combination thereof.
In one embodiment the present invention provides an ophthalmic composition comprising of Brinzolamide or a pharmaceutically acceptable salt thereof, Timolol or a pharmaceutically acceptable salt thereof and potassium sorbate in a pharmaceutically acceptable vehicle.
In another embodiment the present invention provides an ophthalmic composition of Brinzolamide or a pharmaceutically acceptable salt thereof, Timolol or a pharmaceutically acceptable salt thereof and potassium sorbate in a pharmaceutically acceptable vehicle.
In some embodiment the composition according to the present invention is a solution, suspension, drop or an aqueous buffered mixture.
In another embodiment the composition according to the present invention comprises one of more active ingredient in the suspension form.
In yet another embodiment the composition according to present invention is a suspension.
In a further embodiment the present invention provides an ophthalmic suspension of Brinzolamide or a pharmaceutically acceptable salt thereof, Timolol or a pharmaceutically acceptable salt thereof and potassium sorbate in a pharmaceutically acceptable vehicle. In an aspect the composition is free of alkyl quaternary ammonium compounds.
In one embodiment the present invention provides alkyl quaternary ammonium compound free stable ophthalmic composition for reducing intraocular pressure, the disclosed composition is a combination of carbonic anhydrase inhibitor and beta-adrenergic receptor blocker.
In some embodiment the ophthalmic composition according to the present invention comprises a chelating agent selected from edetate disodium, ethylenediamine tetracetic acid, edetic acid, disodium edetate dihydrate, diethylenetriamine pentaacetic acid and/or combinations thereof.
In one embodiment the composition according to present invention is free of polyols selected from sorbitol, mannitol, maltitol, lactitol, xylitol, Isomalt, erythritol or mixture thereof.
In one preferred embodiment the present invention provides an ophthalmic formulation with reduced ocular toxicity for lowering intraocular pressure of eye comprising: about 1% w/v of a carbonic anhydrase inhibitor drug; about 0.5% w/v of a beta blocker drug; potassium sorbate, and an aqueous buffered vehicle, wherein the said formulation is characteristically free of any quaternary ammonium compound.
In another preferred embodiment the composition according to present invention has minimal or non-significant side effects being free of quaternary ammonium compounds like benzalkonium chloride.
In some embodiment according to present invention the particle size of the active ingredient in the present composition is ranging from, D10 in a range of 0.8 µm to 2 µm, D50 in a range of 2 µm to 7 µm, and D90 in a range of 7 µm to 15 µm.
In some embodiment according to present invention the particle size of the active ingredient in the present composition is ranging from, D10 in a range of 0.8 µm to 1.5 µm, D50 in a range of 2 µm to 5.5 µm, and D90 in a range of 7 µm to 15 µm.
In some embodiment the composition according to present invention is stable and is characterised by an assay of Brinzolamide and Timolol within 90.0% to 110% of label claim for at least 3 months at 40oC/75% RH.
In some embodiment the composition according to present invention is stable and is characterised by an assay of Brinzolamide and Timolol within 90.0% to 110% of label claim for at least 3 months at 30oC/75% RH.
In some embodiment the composition according to present invention is stable and is characterised by an assay of Potassium sorbate within 80.0% to 120% of label claim for at least 3 months at 40oC/75% RH.
In some embodiment the composition according to present invention is stable and is characterised by an assay of Potassium sorbate within 80.0% to 120% of label claim for at least 3 months at 30oC/75% RH.
In some embodiment the composition according to present invention is stable and is characterised by impurity level for Brinzolamide, as highest unknown impurity not more than 0.5%, Brinzolamide Related Compound A not more than 1.5% and total impurity (Excluding Brinzolamide compound-A) not more than 2.0%.
In some other embodiment the composition according to present invention is stable and is characterised by impurity level for Timolol, as highest unknown impurity not more than 1.0%, and total impurity not more than 2.0%.
The present invention also provides a method of treating glaucoma or ocular hypertension which comprises topically administering to an affected eye an ophthalmic suspension comprising therapeutically effective amount of a carbonic anhydrase inhibitor drug and one or more other therapeutic agents and suitable excipient(s) dissolved in a pharmaceutically acceptable vehicle, wherein the composition is free of alkyl quaternary ammonium compound.
In some embodiment the present invention provides method for reducing Intra ocular pressure which comprises topically administering to an affected eye an ophthalmic suspension comprising therapeutically effective amount of a carbonic anhydrase inhibitor drug and one or more other therapeutic agents and suitable excipient(s) dissolved in a pharmaceutically acceptable vehicle, wherein the composition is free of alkyl quaternary ammonium compound.
In one preferred embodiment of the present invention, the ophthalmic suspension is free of surfactant or preservative or antimicrobial preservatives defined by the class of quaternary ammonium compounds, organic mercurial compounds, and substituted alcohol and phenol. Particularly, the ophthalmic suspension is free of quaternary ammonium compounds based surfactants and preservatives, such as for example, benzalkonium chloride. These classes of compounds are known to have a surfactant effect as well.
In one embodiment, the ophthalmic suspension of the present invention consisting essentially of therapeutically effective amount of at least one carbonic anhydrase inhibitor drug, at least one beta blocker drug, co-solvent(s) and pharmaceutically acceptable excipients selected from the group comprising of viscosity enhancing agents, tonicity adjusting agents, chelating agents, preservatives and buffers.
In some embodiment the ophthalmic composition according to present invention is free of preservatives, particularly, quaternary ammonium preservatives such as Benzalkonium Chloride (BAK or BKC), Benzethonium Chloride; free of other preservatives such as Benzyl Alcohol, Busan, Cetrimide, Chlorhexidine, Chlorobutanol, Mercurial Preservatives, or phenylmercuric Nitrate, Phenylmercuric Acetate, Thimerosal, phenylethyl Alcohol and like.
In some embodiment the ophthalmic composition according to present invention comprises the safer preservative systems and preservative efficacy enhancers such as edetate disodium, borates, pyruvates, parabens, stabilized oxychloro compounds, Sorbic Acid salts such as Potassium Sorbate, calcium sorbate, sodium sorbate; Polyaminopropyl Biguanide, Polyquaternium-1, Polyhexamethylene biguanide (PHMB), PVP-Iodine complex, metal ions, peroxides, aminoacids, arginine, tromethamine and mixtures thereof may be included within the scope of the present invention.
In certain embodiments of the present invention, the beta-adrenergic blocking agent may be selected from timolol maleate, betaxalol, levobunolol hydrochloride and their therapeutically active salts or esters. The most commonly used and first line drug for the treatment of glaucoma is timolol maleate. Timolol maleate is used in the composition of the present invention in therapeutically effective amounts. Timolol maleate may be used in an amount ranging from about 0.01% w/v to about 2.0 % w/v by weight of the composition, preferably from about 0.05 % w/v to about 1.0 % w/v by weight of the composition and most preferably from about 0.1 % w/v to about 0.5 % w/v by weight of the composition.
In some embodiments the other beta-adrenergic blocking agent, that is suitable for the present invention is levobunolol or its pharmaceutically acceptable salt. It is used in therapeutically effective amounts 0.5 %. In another embodiment, betaxolol or its pharmaceutically acceptable salt is used in amounts ranging from 0.1 % w/v to 0.8 % w/v, preferably, 0.5 % w/v of the ophthalmic composition of the present invention. The preferred amount of beta-adrenergic blocking agent may be included in the concentration of 0.1% w/v to 0.7% w/v, preferably from 0.25% w/v to 0.5% w/v.
The ophthalmic composition of the present invention comprises one or more water soluble excipients selected from a group of a water soluble polymer, an optional penetration enhancer and mixtures thereof. Examples of the water soluble polymers that may be used in the ophthalmic composition of the present invention, include, but are not limited to, natural and synthetic polymers, polysaccharides, polyaminoglycosides, cellulose derivatives, guar gum, xanthan gum, geltrite, dextran, hyaluroante, chondroitin sulfate, locust bean gum, polyvinyl alcohol, polyvinyl pyrrolidone, hydroxypropyl methyl cellulose, hydroxyethyl cellulose, carbopol, polycarbophil, polystyrene sulfonate and like and mixtures thereof.
The ophthalmic composition of the present invention may further comprise pharmaceutically acceptable excipients conventional to the pharmaceutical art. Typical of such pharmaceutically acceptable excipients include osmotic/tonicity-adjusting agents, one or more pharmaceutically acceptable buffering agents and pH-adjusting agents, viscosity enhancing agents, penetration enhancing vehicles and other agents conventional in art that may be used in formulating an ophthalmic composition or imparting a functional property such as gel-forming, bioadhesion, penetration enhancement and like.
The ophthalmic composition of the present invention may be required to be isotonic with respect to the ophthalmic fluids present in the human eye. The composition is having an osmolality of about 250 mOsm/kg to about 500 mOsm/kg, preferably an osmolality of about 275 mOsm/kg to about 475 mOsm/kg, more preferably about 300 mOsm/kg to about 450 mOsm/kg. Osmolality of the composition is adjusted by addition of an osmotic/tonicity adjusting agent. Osmotic agents that may be used in the composition of the present invention to make it isotonic with respect to the ophthalmic fluids present in the human eye, are selected from the group comprising sodium chloride, potassium chloride, calcium chloride, sodium bromide, sodium phosphate sodium sulfate, glycerol, propylene glycol, polyethylene glycols (PEG), PEG-400, PEG-200, PEG300 and the like, and mixtures thereof.
In a one embodiment of the invention the propylene glycol is used as the osmotic agent in an amount ranging from about 0.1 % to about 5.0 % by weight of the composition.
In some embodiments of the present invention, PEG-400 may be used as the osmotic agent. PEG-400 may be present in the composition of the present invention in an amount ranging from about 1.0 % to about 5.0 % by weight of the composition, preferably from about 2.5 % to about 4.0 % by weight of the composition and most preferably in an amount of about 3.0 % by weight of the composition.
In some embodiment of the invention the osmotic agent according to present invention may comprises a combination of osmotic agents in an amount ranging from about 0.1 to about 10% by weight of the composition.
In order to achieve, and subsequently maintain, an optimum pH, the ophthalmic composition may contain a pH adjusting agent and/or a buffering agent. The range of pH for an ophthalmic formulation is about 4.0 to about 8.0, the preferred pH is about 5.5-7.5, and more preferred pH is about 5 to about 7. The ophthalmic composition of the present invention comprises a pharmaceutically acceptable pH adjusting agents that may be selected from the group comprising acetic acid or salts thereof, boric acid or salts thereof, phosphoric acid or salts thereof, citric acid or salts thereof, tartaric acid or salts thereof, sodium hydroxide, potassium hydroxide, sodium carbonate, sodium hydrogen carbonate, trometamol, arginine, lysine, histidine, guanine and the like and mixtures thereof. Particularly, preferred pH adjusting agents that may be used in the ophthalmic composition of the present invention include acetic acid, hydrochloric acid, tromethamine, arginine and sodium hydroxide. These agents are used in amounts necessary to produce a pH ranging from about 4.5 to about 8.0, preferably 5 to about 7.5.
According to one embodiment the composition of the present invention comprises of one or more solvents or co-solvents. The pharmaceutically acceptable solvents may be selected from a group of alcohols, such as alcohols having 2 or 3 hydroxyl groups, ethanol, and glycols such as ethylene glycol, propylene glycol, polyethylene glycol, glycofurol and like.
Besides above mentioned ingredients, one embodiment of the present invention may comprise a number of additional components to provide various functional effects, as is well known in this field. For example, small organic acids may be included as buffers, polymers for ocular retention, co-preservatives etc.
One embodiment of the present invention further provides a process of preparation of an ophthalmic composition wherein the composition comprises a polymeric vehicle. In one embodiment, the suspension is prepared on a large scale batch such as more beta-adrenergic blocking agent is dissolved in a pharmaceutical vehicle, and preparing the polymeric vehicle separately. The solution may be slowly stirred to dissolve the swollen or gelatinized particles completely. Once the water soluble excipient such as the polymeric vehicle is prepared, the active ingredient phase is prepared that is, timolol maleate is separately dissolved in water for injection. Separately, one or more buffering agents such as boric acid may be added and dissolved in the above solution under stirring. Separately, the carbonic anhydrase inhibitor drug such as for example, Brinzolamide is taken and added to surfactant slurry such as Tyloxapol and stirred. This slurry of Brinzolamide is added to the Timolol maleate aqueous solution under stirring. The volume is made up with aseptically filtered water for injection and stirred. The pH is monitored and adjusted to 6.7-7.3, if required. Again the suspension is polish filtered through filter. The suspension is then filled into containers and the containers are subsequently sealed. The container may be purged with nitrogen.
In one embodiment, the process for the preparation of the ophthalmic composition of the present invention comprises:
i. preparation of a carbomer phase;
ii. preparation of a vehicle concentrate phase;
iii. mixing of the carbomer phase and vehicle concentrate phase to achieve a homogenous mixture phase;
iv. preparation of a solution of the beta blocker drug in an aqueous vehicle and mixing with the above mixture phase to get beta blocker solution phase;
v. preparation of a solution of the potassium salt of sorbic acid in an aqueous vehicle and mixing with the above beta blocker solution phase;
vi. adding and stirring the carbonic anhydrase drug in a solution of a non-ionic liquid polymer to get a homogenous dispersion;
vii. mixing the above homogenous dispersion with the beta blocker solution phase and adjusting the pH to get a bulk suspension; and
viii. polishing filter the bulk suspension to get the ophthalmic formulation.
While the present invention is disclosed generally above, additional aspects are further discussed and illustrated with reference to the examples below. However, the examples are presented merely to illustrate the invention and should not be considered as limitations thereto.

EXAMPLE
Example 1: Composition of Ophthalmic suspension
S. No Ingredients Qty. (mg/ml)
1. Brinzolamide IP (Micronized And Sterile) 10
2. Timolol Maleate IP ( Eq. to Timolol 5mg) 6.83#
3. Carbomer Homopolymer Type B NF (Carbopol 974p) 1.2 – 5.7
4. Disodium Edetate IP/BP 0.31 – 0.75
5. Sodium Chloride IP/BP 0.28 – 0.7
6. Tyloxapol USP 0.23 – 0.79
7. Boric Acid IP/BP 2.9 – 6.1
8. Glycerin IP/Glycerol BP 4.5 – 10.4
9. Propylene Glycol IP/BP 2.5 – 6.5
10. Potassium Sorbate IP/NF 2.6 – 5.9
11. Sodium Hydroxide IP/BP q.s. to Adj pH
12. Hydrochloric Acid IP/BP q.s. to Adj pH
13. Water For Injection q.s.
#6.83 mg Timolol maleate is equivalent to 5 mg of Timolol
The ophthalmic suspension of was surprisingly found to remain stable in terms of chemical assay when stored at 30oC/75% RH for at least 3 month or up to 24 months.

Example 2: Composition of Ophthalmic suspension
S. No Ingredients Qty. % w/v
1. Brinzolamide IP (Micronized And Sterile) 1
2. Timolol Maleate IP ( Eq. To Timolol 5mg) 0.683#
3. Carbomer Homopolymer Type B NF(Carbopol 974p) 0.35
4. Disodium Edetate IP/BP 0.055
5. Sodium Chloride IP/BP 0.05
6. Tyloxapol USP 0.05
7. Boric Acid IP/BP 0.5
8. Glycerin IP/Glycerol BP 0.88
9. Propylene Glycol IP/BP 0.44
10. Potassium Sorbate IP/NF 0.47
11. Sodium Hydroxide IP/BP q.s. to Adj pH
12. Hydrochloric Acid IP/BP q.s. to Adj pH
13. Water for Injection q.s.
#0.683%w/v Timolol maleate is equivalent to 5 mg of Timolol
Preparation
Preparation of Carbomer phase
- Disperse carbomer in WFI under high speed stirring and continue until a homogeneous lump free liquid is formed.
Preparation of Vehicle concentrate phase
- Dissolve Edetate disodium di-hydrate, boric acid, sodium chloride, propylene glycol, glycol individually in portion of WFI with stirring. The dissolution of each ingredient was ensured visually; Combine the solutions to make a vehicle concentrate phase.
Phase III: Mixing phase (Carbomer phase and Vehicle Concentrate) and SIP
- the vehicle concentrate (Phase II) filtered using 0.45gm nylon capsule filter and add directly into this Carbomer 974P phase (Phase l) under stirring; Stir well to ensure homogeneous mixing;
- Sterilize this phase at 121oC for 35 minutes under stirring.
- Bring down the temperature to 20-25oC.
- Transfer the cooled phase into sterile manufacturing SS316L tank in sterile area.
Phase IV: Preparation of Timolol maleate solution phase
- Dissolve Timolol maleate in a portion of WFI under stirring. The solution was aseptically filtered with a set of 0.45µ &0.2 µ filter and directly add in phase III in sterile area. The contents are mixed under stirring.
Phase V: Preparation of Potassium sorbate solution phase its Mixing into polymer phase and pH adjustment
- Dissolve Potassium sorbate in a portion of WFI with stirring. Filter the solution using set of 0.45 µ & 0.2 µ filter and directly add in phase III.
- Mix the contents and adjust the pH to 7.0 ± 0.2 with sterile filtered 10% Sodium Hydroxide and/or hydrochloric acid solution.
Phase VI: Preparation of Brinzolamide -Tyloxapol Slurry
- Dissolve tyloxapol in a portion of WFI with stirring in a vessel.
- The solution was filtered using set of 0.45µ & 0.2µ filter and collected it in a sterile homogenizer tank in sterile area. Add sterile and micronized Brinzolamide aseptically added to the above tyloxapol solution under continuous stirring, followed by homogenizing the slurry by recirculating through inline homogenizer for 15 minutes for homogeneous dispersion.
Phase VII: Final bulk preparation
- the Brinzolamide slurry is aseptically transferred to the tank containing vehicle phase
- Rinse suspension container and add to the above phase.
- then the pH was checked and adjusted to 7.0 ± 0.3 with sterile filtered 10% w/v Sodium Hydroxide and/or hydrochloric acid solution the pH of Bulk was between 6.7 to 7.3.
- the volume was made up using sterile water for injection
- then the bulk suspension polish filtered through 70-micron PP filter and the filtered suspension collected into buffer tank for filling and transfer of remaining suspension to bulk suspension.

Example 3: Composition of Ophthalmic suspension
S. No Ingredients Qty. (mg/ml)
3a 3b 3c 3d
1. Brinzolamide (Micronized & Sterile) 10 10 10 10
2. Timolol Maleate IP ( Eq. to Timolol 5mg) 6.83# 6.83# 6.83# 6.83#
3. Carbomer Homopolymer Type B (Carbopol 974p) 3.5 3.5 3.5 3.5
4. Disodium Edetate IP/BP 0.55 NIL 0.55 0.55
5. Sodium Chloride IP/BP 0.5 0.5 0.5 0.5
6. Tyloxapol USP 0.5 0.5 0.5 0.5
7. Boric Acid IP/BP 5 5 NIL 5
8. Glycerin IP/Glycerol BP 8.8 8.8 8.8 NIL
9. Propylene Glycol IP/BP 4.4 4.4 4.4 NIL
10. Potassium Sorbate IP/NF 4.7 4.7 4.7 4.7
11. Sodium Hydroxide IP/BP q.s to 7 q.s to 7 q.s to 7 q.s to 7
12. Hydrochloric Acid IP/BP q.s to 7 q.s to 7 q.s to 7 q.s to 7
13. Water for Injection q.s. q.s. q.s. q.s.
#6.83 mg Timolol maleate is equivalent to 5 mg of Timolol
Study:
The example 3 samples were prepared by omitting components of the formulation, and to evaluate the impact of such change in formulation on the overall composition.
Observation:
Various test parameters were evaluated such as PET, viscosity, osmolality and pH for the samples of example 3, and it was observed that the sample according to present invention compiles with the specification and found suitable, for preparing a stable dosage form.

Example 4: Stability Study
Samples of example 3a were kept for stability study under stress condition of 40oC/75% RH and 30oC/75%RH, the study results are shown below:
Stage Assay
of BZ % Assay of TM %
RS by LC of BZ in % RS by LC
of TM in % pH Osmolality
RC-A Highest unk. Imp Total imp. Highest unk. Imp Total imp.
Spec. 90-110% (Label claim) 90-110% (Label claim)
NMT 1.5% NMT 0.5% NMT 2.0% NMT 1.0% NMT 2.0% pH 6.2 – 8.2 300-450mOsm/Kg
0 M 99.35 99.40
ND 0.05 0.045 0.017 0.017 7.10 375
3M – 40oC/75% RH 100.74 94.89
0.328 0.061 0.077 0.288 0.468 7.14 381
6M – 40oC/75% RH 97.26 96.02
0.474 0.145 0.266 0.34 0.927 7.22 392
3M – 30oC/75% RH 98.43 94.66
0.030 0.05 0.052 0.14 0.173 7.15 379
6M – 30oC/75% RH 95.38 99.54
0.109 0.06 0.094 0.198 0.285 7.23 385
12M – 30oC/75% RH 97.66 99.45
0.128 0.053 0.129 0.331 0.483 7.14 384
RH: Relative Humidity, NMT: Not more than, BZ: Brinzolamide; TM: Timolol; RC-A: Related compound A; Unk. Imp.: Unknown Impurity; Imp.: Impurity; M: Month; RS: Related Substance
Samples of test formulation were found to be stable and within specification throughout the stability period. The samples were found stable for at least 6 months at 30oC/75%RH or 40oC/75% RH.

Example 5: Pre-clinical Efficacy Study
To compare efficacy of potassium sorbate preserved formulation of brinzolamide (1%) and Timolol Maleate (0.5%) v/s benzalkonium chloride preserved ophthalmic suspension of brinzolamide (1%) plus timolol maleate (0.5%) - Brinzotim® in normotensive New Zealand White (NZW) rabbits (n=12, i.e. 6 in each group) following single ocular instillation.

Methods:
• Twelve male rabbits of 9-12 months weighing 3.4 - 4.4kg were divided in two groups of six each. Each group received either test drug or reference drug.
• Two days of pre-treatment IOP (Intra ocular pressure) was recorded and average IOP was calculated. Reference drug or test drug of 35µl was instilled in left eye of each animal on day 0.
• Subsequent IOP readings were taken using pneumatonometer at 2,4,6,8 and 24hrs (day 1) post instillation. Percentage change in IOP was calculated with respect to initial IOP readings in each group.

Table 1: Study Design:

Group Design Concentration of Drugs Dose (µg/eye) Volume
(µg/ml) single dose No. of animals in group
(B) (T) B T
A Test item *
[Brinzolamide (B) + Timolol (T)] potassium sorbate preserved 1% 0.5% 350 175 35 6
B Reference drug (Brinzotim)#
BKC Preserved 1% 0.5% 350 175 35 6
# BKC: Benzalkonium Chloride; B: Brinzolamide; T: Timolol

Table 2: Activity Schedule for assessment:

Activity Day 2 Day 2 & 1 Time Point (Hours)
Day 0 Day 1
9 am 5 pm 0 2 4 6 8 24
Body Weight ?
Grouping of Animals ?
IPO measurement (baseline) ? ? ?
Test Items / reference items instillation ?
IPO measurement (After treatment) ? ? ? ? ?

Statistical Analysis:
• All statistical analysis was carried with PRISM (GraphPad version 5.03, December 10, 2009) and p<0.05 was considered as statistically significant
• The IOP data were also analyzed using Two-way ANOVA followed by Bonferroni posttests (n=6) for comparison between the groups.

Result:
Table 3: Comparison of IOP reduction potential

Time Points Test Item without BKC Brinzotim® with BKC
Hours Mean IOP
(mmHg) ± SEM ?IOP
(Compared to Baseline) % Change Mean IOP
(mmHg) ± SEM ?IOP
(Compared to Baseline) % Change
0 19.78±0.61 - - 20.54±1.18 - -
2 14.73*±0.41 -5.0 -25.5 15.90*±0.74 -4.6 -22.6
4 16.08*±0.37 -3.7 -18.7 17.73*±0.77 -2.8 -13.7
6 17.04*±0.76 -2.7 -13.8 18.75*±0.97 -1.8 -8.7
8 19.05±0.99 -0.7 -3.7 20.27±1.00 -0.3 -1.3
24 19.68±0.44 -0.1 -0.5 20.73±0.99 0.2 0.9
*p<0.001 Differences were statistically significant as compared to initial mean IOP value.
Single ocular instillation of both Test drug and Brinzotim showed significant reduction of IOP up to 6 hrs. in normotensive NZW rabbits. No significant difference was observed between two groups at all-time points (Figure 1). However the toxic effect of Benzalkonium chloride based formulation as can be seen in long term use (Christophe Baudouin et al., Preservatives in eye drops: The good, the bad and the ugly, Elsevier, Progress in Retinal and Eye Research 29(2010) 312-334), depicts that the present formulation without BKC is therapeutically more desirable with lesser side effects and associated toxicity of benzalkonium based preparations as used in comparator.

Example 6: Pre-Clinical Ocular Toxicity Study

Study 1:
Repeated Dose 28-Day Ocular Toxicity Study of Brinzolamide and Timolol Ophthalmic Suspension (1% w/v + 0.5% w/v) in New Zealand White Rabbits.
Method:
New Zealand White rabbits were randomly divided into six groups (G1 to G6), each group comprised of three male rabbits and three female rabbits. Rabbits were treated with repeated once daily ocular instillation of Brinzolamide and Timolol Ophthalmic Suspension for a minimum period of 28 consecutive days. The selected dose levels for rabbit study viz. 0.8 mg/kg, 1.6 mg/kg and 3.2 mg/kg body weight were approximately 1, 2 and 4 times respectively of the human dose considering the rabbit body surface area.
Study 2:
Repeated Dose 28-Day Ocular Toxicity Study of Brinzolamide and Timolol Ophthalmic Suspension (1% w/v + 0.5% w/v) in Wistar Rats.
Method:
Wistar rats were randomly divided into six groups (G1 to G6), each group comprised of three male rats and three female rats. Rats were treated with repeated once daily ocular instillation of Brinzolamide and Timolol Ophthalmic Suspension for a minimum period of 28 consecutive days. The selected dose levels for rat study viz. 1.55 mg/kg, 3.1 mg/kg and 6.2 mg/kg body weight were approximately 1, 2 and 4 times respectively of the human dose considering the rat body surface area.
Assessment Parameters:
- Clinical signs and adverse reaction related to treatment;
- Ophthalmologic examination:
o tonometry (intraocular pressure/tension),
o slit lamp examination (cornea, iris, and aqueous humor),
o fluorescent dye (sodium fluorescein, 0.25 to 1.0%) test for detecting defects in surface epithelium of cornea and conjunctiva.
- Body weight and feed consumption.
- Biochemical and hematological analysis of blood sample.
- Urine analysis.
- Organ weight (internal & external), pathological and histopathological examination of the body organ.
Conclusion:
i. The repeated dose ocular instillation of Brinzolamide and Timolol Ophthalmic Suspension for a period of 28 days with low dose, mid dose, high dose and recovery high dose did not show any ocular lesions.
ii. No significant differences in the body weight, feed consumption.
iii. Clinical biochemistry and hematological parameters did not show any significant changes.
iv. Urine analysis parameters did not reveal any treatment related changes.
v. No treatment related abnormalities in gross pathological and histopathological examination seen.
Hence, high dose of Brinzolamide and Timolol Ophthalmic Suspension (4 times respectively of the human dose) was concluded as NOAEL (No Observed Adverse Effect Level).

,CLAIMS:We Claim:
1. An ophthalmic formulation for lowering intraocular pressure of eye comprising:
about 1% w/v of a carbonic anhydrase inhibitor drug;
about 0.5% w/v of a beta blocker drug;
a potassium sorbate, and
an aqueous buffered vehicle,
wherein the said formulation is characteristically free of any quaternary ammonium compound.
2. The ophthalmic formulation as claimed in claim 1, wherein the said carbonic anhydrase inhibitor drug is brinzolamide.
3. The ophthalmic formulation as claimed in claim 1, wherein the said beta blocker drug is timolol.
4. The ophthalmic formulation as claimed in claim 1, wherein the said quaternary ammonium compound is benzalkonium chloride.
5. The ophthalmic formulation as claimed in claim 1, wherein the formulation is characterized by reduced ocular toxicity.
6. A stable ophthalmic formulation of a carbonic anhydrase inhibitor drug for lowering intraocular pressure of eye comprising about 1% w/v of a carbonic anhydrase inhibitor drug, about 0.5% w/v of a beta blocker drug, and potassium sorbate in an aqueous buffered vehicle, wherein the formulation is characteristically free of quaternary ammonium compound.
7. A stable ophthalmic formulation of a carbonic anhydrase inhibitor drug for lowering intraocular pressure of eye comprising:
about 1% w/v of a carbonic anhydrase inhibitor drug,
about 0.5% w/v of a beta blocker drug, and
potassium sorbate in an aqueous buffered vehicle,
wherein the formulation D50 of the drug is not more than 10 µm.
8. The stable ophthalmic formulation as claimed in claim 6 or 7, wherein the formulation characterized by a level of related compound A is not more than 1.5% upon storage at 30oC/75% RH for at least 12 months and upon storage at 40oC/75% RH for at least 6 months.
9. The stable ophthalmic formulation as claimed in claim 6 or 7, wherein the formulation characterized by a level of highest unknown impurity is not more than 0.5% upon storage at 30oC/75% RH for at least 12 months and upon storage at 40oC/75% RH for at least 6 months.
10. The stable ophthalmic formulation as claimed in claim 6 or 7, wherein the formulation characterized by a level of total impurity is not more than 2.0% upon storage at 30oC/75% RH for at least 12 months and upon storage at 40oC/75% RH for at least 6 months.
11. A benzalkonium chloride free composition for lowering intraocular pressure in an eye comprising:
about 1% w/v brinzolamide;
about 0.5% w/v timolol;
about 0.3% w/v to 0.5% w/v Potassium Sorbate, and
an aqueous buffered vehicle.
12. The composition as claimed in claim 11 is a suspension for lowering intraocular pressure in an eye comprising:
about 1% w/v brinzolamide;
about 0.5% w/v timolol;
about 0.47% w/v Potassium Sorbate, and
an aqueous buffered vehicle.
13. A process for preparation of an ophthalmic formulation of carbonic anhydrase inhibitor drug, beta blocker drug and potassium sorbate, wherein the process comprises steps of:
i. preparation of a carbomer phase;
ii. preparation of a vehicle concentrate phase;
iii. mixing of the carbomer phase and vehicle concentrate phase to achieve a homogenous mixture phase;
iv. preparation of a solution of the beta blocker drug in an aqueous vehicle and mixing with the above mixture phase to get beta blocker solution phase;
v. preparation of a solution of the potassium sorbate in an aqueous vehicle and mixing with the above beta blocker solution phase;
vi. adding and stirring the carbonic anhydrase drug in a solution of a non-ionic liquid polymer to get a homogenous dispersion;
vii. mixing the above homogenous dispersion with the beta blocker solution phase and adjusting the pH to get a bulk suspension; and
viii. filtering the bulk suspension to get the ophthalmic formulation.
14. The process as claimed in claim 13, wherein the pH of the formulation is in a range of pH 4.0 to pH 8.
15. A benzalkonium chloride free stable ophthalmic suspension for lowering intraocular pressure in an eye comprising:
about 1% w/v brinzolamide;
about 0.5% w/v timolol;
about 0.47% w/v Potassium Sorbate;
carbomer;
tonicity adjusting agents;
surfactant;
pH adjusting agents, and
pharmaceutically acceptable vehicle.
16. The suspension as claimed in claim 15, wherein the tonicity adjusting agent is selected from sodium chloride, glycerine, propylene glycol, mixture of dihydroxyl and trihydroxyl alcohols, inorganic salts, or mixture thereof.
17. The suspension as claimed in claim 15, wherein the surfactant is Tyloxapol.
18. The suspension as claimed in claim 15, wherein the pH adjusting agent is selected from sodium hydroxide, hydrochloric acid or mixture thereof.
19. The suspension s claimed in claim 15, wherein the suspension further comprises buffer as boric acid.
20. The suspension is claimed in claim 15, wherein the suspension further comprises chelating agent as ethylenediamine tetracetic acid.
21. The suspension as claimed in claim 15, wherein the vehicle is an aqueous vehicle.

Dated this 28th day of January, 2022


Dr. Gaurav Sahal
Senior General Manager
Sun Pharmaceutical Industries Limited