DESC:
FIELD OF THE INVENTION
The present invention relates to pharmaceutical formulations suitable for intravitreal administration comprising a vascular endothelial growth factor (VEGF) specific fusion protein antagonist and method for making and using such formulations for the treatment of methods of manufacture of the formulations, method of their administration, and kits containing the same. More specifically, the invention relates to stable aqueous pharmaceutical formulation for intravitreal administration comprising a therapeutically effective amount of aflibercept, buffer, bile acid salt, anti-aggregating agent, optionally tonicity modifier and one or more pharmaceutically acceptable excipients suitable for ophthalmic administration.

BACKGROUND OF THE INVENTION
Aflibercept is vascular endothelial growth factor (VEGF) Inhibitor and a recombinant fusion protein consisting of portions of human VEGF receptors 1 and 2 extracellular domains fused to the Fc portion of human IgG1.

US FDA approved Aflibercept containing products and commercially available under the brand names of Eylea® and Zaltrap®. Eylea® (aflibercept) is approved in the form of intravitreal injection and it is indicated for the treatment of Neovascular (Wet) Age-Related Macular Degeneration (AMD), Macular Edema Following Retinal Vein Occlusion (RVO), Diabetic Macular Edema (DME) and Diabetic Retinopathy (DR).

Aflibercept acts as a soluble decoy receptor that binds VEGF-A and placental growth factor (PlGF) with higher affinity than their natural receptors, and thereby can inhibit the binding and activation of these cognate VEGF receptors. Aflibercept’s high affinity for VEGF prevents the subsequent binding and activation of native VEGF receptors. Reduced VEGF activity leads to decreased angiogenesis and vascular permeability.

U.S. Patent No. 8,092,803 and US 10,464,992 discloses liquid ophthalmic formulation of aflibercept, sodium phosphate buffer, polysorbate, sucrose, sodium chloride with pH about 6.2-6.3.

U.S. Patent Application No. 20190030123 discloses liquid pharmaceutical composition comprising a) a histidine containing buffer, b) a non-ionic surfactant, c) a VEGF antagonist, d) an inorganic salt, and e) a carbohydrate.

U.S. Patent Application No. 20190160145 discloses ophthalmic pharmaceutical composition comprising a) aflibercept; b) an acetate salt buffering agent; c) a sugar or a sugar alcohol; and d) a surfactant in an aqueous medium, wherein the ophthalmic pharmaceutical composition is free of an ionic tonicity agent.

U.S. Patent Application No. 20190276528 discloses aqueous formulation comprising: a) aflibercept; b) a stabilizer; and c) an organic co-solvent, wherein the formulation is free of buffer.

Aflibercept ophthalmic formulations were disclosed in the art. There is still exists a need to develop an alternative, stable, ready to use pharmaceutical formulation of aflibercept. The inventors of the present invention have surprisingly found that a stable pharmaceutical formulation of aflibercept suitable for intravitreal administration can be developed using bile acid salt and anti-aggregating agent.

SUMMARY OF THE INVENTION
The invention is directed to a stable aqueous pharmaceutical formulation comprising a therapeutically effective amount of aflibercept, buffer, bile acid salt, anti-aggregating agent and optionally, tonicity modifier.

The object of present invention is to reduce aggregates or impurities or flocculation formation during formulation and storage. The inventors of the present invention found that the addition of bile acid salt and anti-aggregating agent in formulation makes the stable formulation by inhibiting aggregation and formation of degradant species.

In one aspect, the invention provides a stable aqueous pharmaceutical formulation suitable for intravitreal administration comprising a therapeutically effective amount of aflibercept, buffer, bile acid salt, anti-aggregating agent and optionally, tonicity modifier wherein the pharmaceutical formulation has a pH from about 5.0 to 7.0.

In another aspect, a stable aqueous pharmaceutical formulation suitable for intravitreal administration comprising:
(a) aflibercept;
(b) buffer;
(c) bile acid salt and
(d) anti-aggregating agent and optionally a tonicity modifier.

In another aspect, a stable aqueous pharmaceutical formulation suitable for intravitreal administration comprising:
(a) aflibercept;
(b) sodium phosphate or Histidine-Histidine HCl;
(c) sodium deoxycholate and
(d) sucrose and optionally, sodium chloride.

In another aspect, a stable aqueous pharmaceutical formulation suitable for intravitreal administration comprising:
(a) aflibercept;
(b) sodium phosphate or L Histidine and L Histidine HCl;
(c) sodium cholate and
(d) sucrose and optionally sodium chloride.

The pH of the formulation may be between 5.5 to 7.5, preferably between 6.0 to 7.0. Also, preferably, the pH is 6.2 or 6.8.

In present invention, aflibercept formulation comprising combination of bile acid salt with anti-aggregating agent shows comparable stability over the formulation containing polysorbate. Eylea® (aflibercept) contains inactive ingredient such as sodium phosphate, sodium chloride, polysorbate 20, and sucrose, with a pH of 6.2. Further, the formulation comprising combination of bile acid salt with anti-aggregating agent reduces the low and high molecular weight impurities.

The invention further features ophthalmic formulations provided in a pre-filled syringe or vial or any device, particularly suitable for intravitreal administration.

Other objects and advantages will become apparent from a review of the ensuing detailed description.

DETAILED DESCRIPTION OF THE INVENTION
The present invention is described in detail by the way of reference only using the following definitions and examples. The present invention is not limited to particular methods, and experimental conditions described herein, as such methods and conditions may vary. The terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting unless indicated, since the scope of the present invention will be limited only by the appended claims. All technical and scientific terms and phrases used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention belongs.

DEFINITIONS
The term "VEGF antagonist" refers to a molecule, which specifically interacts with VEGF and inhibits one or more of its biological activities, e.g. its mitogenic, angiogenic and/or vascular permeability activity. It is intended to include both anti-VEGF antibodies and antigen-binding fragments thereof and non-antibody VEGF antagonists.

The term "aflibercept" is a recombinant fusion protein consisting of Vascular Endothelial Growth Factor (VEGF)-binding portions from the extracellular domains of human VEGF Receptors 1 and 2 fused to the Fc portion of the human IgG1. Aflibercept is a dimeric glycoprotein with a protein molecular weight of 97 kilodaltons (kDa) and contains glycosylation, constituting an additional 15% of the total molecular mass, resulting in a total molecular weight of 115 kDa. Amino acid and nucleic acid sequences of aflibercept are known in the art.

The term "pharmaceutical formulation" or "composition" as used herein refers to a mixture of a protein, such as a fusion protein, e.g., aflibercept, together with one or more additional components. In some embodiments, the additional components may include, e.g., one or more excipients, such as a buffer, anti-aggregating agent, tonicity modifier, bile acid salts, water and the like, e.g., a pharmaceutically acceptable carrier or excipient that is conventional in the art and which is suitable for administration to a subject for therapeutic, diagnostic, or prophylactic purposes. In one embodiment, the additional component is suitable for ophthalmic administration. For example, pharmaceutical compositions/formulations according to the present invention may be aqueous formulation suitable for intravitreal administration.

The term "buffer" refers to a buffered solution that resists changes in pH by the action of its acid-base conjugate components. In one example, a buffer consists of a weak acid and a conjugate salt or a weak base and a conjugate salt (e.g. sodium phosphate mono-basic and di-basic, Histidine-Histidine HCl). Buffers suitable for use in connection with this invention may have a pH in the range from about 4.0 to about 9.0; from about pH 4.0 to about 7.0; or from about pH 4.5 to about 6.5. A pH of any point in between the above ranges is also contemplated.

The term "stable" is understand to mean that aflibercept contained in the formulation or pharmaceutical compositions does not lose more than 20%, or more preferably 15%, or even more preferably 10%, and most preferably 5% of its activity relative to activity of the formulation or composition at the beginning of storage. The term “Stable” mean that the aflibercept formulations or compositions are at least comparable to, or better than commercially available aflibercept formulation, in terms of their stability and/or ability to resist formation of particulates during storage. In another embodiment, the term means that the aflibercept formulations or compositions are stable when agitated, mixed, or handled.

The term "intravitreal injection" refers to the administration of a pharmaceutical formulation in which the substance is injected directly into the eye. More specifically, the substance is injected into the vitreous humor (also called vitreous body or simply vitreous) which is the clear gel that fills the space between the lens and the retina of the eyeball of humans and other vertebrates.

The term “excipient” used here in refers to additional component may include, e.g., one or more excipients, such as water, buffer, bile acid salt, anti-aggregating agent, tonicity modifier etc. and the like, e.g., a pharmaceutically acceptable carrier or excipient that is conventional in the art and which is suitable for administration to a subject for therapeutic, diagnostic, or prophylactic purposes. In one embodiment, the additional component is suitable for ophthalmic administration. For example, pharmaceutical formulations according to the present invention may be aqueous formulation suitable for intravitreal administration.

The term “high molecular weight” (HMW) species, refers to higher order oligomeric species like dimers, trimers, tetramers, etc., formed by addition of monomers that can be either covalently or non-covalently linked, which consist of misfolded monomers in which surfaces of the monomer are exposed that typically would not be in the folded monomeric form.

The term “low molecular weight” (LMW) species, refers to clipped species of intact antibodies composed of heavy and light chains.

In one embodiment, the aflibercept used as an active ingredient in the pharmaceutical formulation of the present invention may be included in a therapeutically effective amount. For example, aflibercept may be present in a concentration ranging from 10 to 100 mg/ml, preferably in a concentration ranging from 20 to 50 mg/ml, more preferably in a concentration of about 40 mg/ml, but not limited thereto.

In one embodiment, the buffer is selected from the group consisting of acetate, histidine, phosphate, citrate, succinate, tartrate, maleate and combination thereof. In another embodiment, the buffer is a phosphate buffer, for example, sodium phosphate.

In one embodiment, the buffer is present at a concentration in the range of about 1 mM to 150 mM. In a preferred embodiment, the buffer is in a concentration in the range of 1 mM to 40 mM, preferably of 2 mM to 35 mM, more preferably of 3 mM to 30 mM, even more preferably of 5 mM to 20 mM and more preferably of 8 mM to 15 mM. Most preferably, the buffer is present at a concentration of about 10 mM.
In one embodiment, sodium phosphate buffer is present at a concentration in the range of about 5-40 mM of. In a particular preferred embodiment, the buffer is in a concentration of 10 mM.

Bile acid salts are naturally occurring cholesterol derivatives with a steroidal skeleton and specific self-assembly and interface behaviors. Cholic acid is a primary bile acid and salts of cholic acid are called cholates. Deoxycholic acid is one of the secondary bile acids.

In one embodiment, bile acid salt is selected from the group consisting of sodium deoxycholate and sodium cholate. In another embodiment, bile acid salt is sodium deoxycholate.

In one embodiment, the bile acid salt is present in a concentration ranging from 0.001% w/v to 0.1 w/v %. In a particular preferred embodiment, the bile acid salt is present in a concentration ranging from 0.01 to 0.05 w/v %, preferably from 0.01 to 0.03 w/v%.

In one embodiment, the anti-aggregating agent is selected from the group consisting of amino acids, sugars, polyols, polymers, and combinations.

In one embodiment, the anti-aggregating agent is a sugar. In another embodiment, the anti-aggregating agent is sucrose, sorbitol, trehalose, glycerol, or mannitol. In another preferred embodiment, the anti-aggregating agent is sucrose.

In one embodiment, the sugar may be present in a concentration ranging from 5% w/v to 12% w/v, preferably from 6% w/v to 10% w/v, more preferably about =8% w/v, most preferably about 9% w/v.

A tonicity modifier is an ingredient that contributes to the osmolality of a solution.

In one embodiment, tonicity modifier is selected from the group consisting of sugars, amino acids or inorganic salt, which have osmoregulatory properties. Preferred inorganic salts for use in the pharmaceutical formulation of the present invention are potassium chloride, calcium chloride, sodium chloride, sodium phosphate, potassium phosphate and sodium bicarbonate. Preferably, the inorganic salt is a sodium salt; more preferably, it is sodium chloride (NaCl).

The concentration of the inorganic salt or sodium chloride in the pharmaceutical formulation used in the present invention is preferably in the range of 0 to 150 mM, more preferably in the range of 0 to 80 mM, even more preferably the inorganic salt has a concentration in the range of 0 to 60 mM or 0 to 45 mM, and most preferably the concentration is 0 mM to 40 mM.

In a most preferred embodiment, the inorganic salt is sodium chloride with a concentration of 0 to 40 mM.

STABILITY
Pharmaceutical formulations containing proteins has multiple stability challenges in view of the unique chemical and physical properties of protein. Physical instability involves aggregates/ impurities/ flocculation. Inventors of this present invention found that by using bile acid salt with buffer, anti-aggregating agent and optionally, tonicity modifier in aflibercept formulation prevents aggregates/ impurities/ flocculation resulting in stable formulation.

Most common protein based impurities includes aggregates such as high molecular weight (HMW) and low molecular weight (LMW) species, charged variants such as acidic variants and basic variants, and host-cell proteins.

High molecular weight and low molecular weight species have the potential to affect the safety and efficacy of biopharmaceuticals. As a result, the levels of these protein impurities in biologic drug substances and drug products must be controlled and are typically considered as critical quality attributes. The properties of the different contaminants vary significantly.

In one aspect of the invention, a stable aqueous formulation is a formulation having less than about 10%, or less than about 5% of the protein being present as aggregate in the formulation.

In present invention, VEGF antagonist aqueous formulation comprising a bile acid salt such as sodium cholate, sodium deoxycholate or combination thereof; surprisingly shows lower protein based impurities such as high molecular weight (HMW) and low molecular weight (LMW) species formation over longer duration.

In preferred embodiment aqueous aflibercept formulation comprising sodium deoxycholate in concentration 0.01% w/v to 0.05% w/v, shows lower percentage of HMW and LMW formation than control formulation. The control formulation is Eylea® (aflibercept) by Regeneron Pharmaceuticals, Inc.

A protein in an aqueous formulation can be stable at frozen storage temperatures (-20°C or -80°C), liquid storage temperatures such as refrigeration (e.g. 2-8°C) and room temperature (about 25 °C), and under accelerated liquid stability testing temperatures (e.g. 40 °C.).

Various analytical techniques for measuring protein stability, including techniques for measuring the type and degree of particulates that may be present in protein formulations, are available in the art and are reviewed in Peptide and Protein Drug Delivery, 247-301 (Vincent Lee ed., New York, N.Y., 1991) and Jones, 1993 Adv. Drug Delivery Rev. 10: 29-90, for examples. Stability can be measured at a selected temperature for a selected time period, e.g., as described in the examples below.

The Size exclusion chromatography (SEC) analysis was conducted on formulations of the present invention to measure the changes in stability.

The osmolality of a pharmaceutical formulation is adjusted to minimize discomfort to the patient upon administration. Additionally, osmolality of a formulation may affect the active ingredient's stability and can be adjusted to improve the stability. It is generally preferred that a pharmaceutical formulation for intravitreal administration to a patient be isotonic with vitreous, i.e., have the same or similar osmolality, which is achieved by addition of a tonicity modifier. However, hypertonic formulations which would then be diluted in an isotonic vehicle are also within the scope of this invention.

In one embodiment, the osmolality of the present formulations is from about 180 to about 1000mOsM. In another embodiment, the osmolality of the present formulations is from about 300 mOsM.

The present invention preferably has a pH in the range from about 6.0 to about 7.0, preferably from about 6.1 to about 6.8, more preferably from about 6.0 to 6.5, even more preferably from about 6.2 to 6.5 and most preferably has a pH of about 6.2 or 6.8.

In one embodiment, the stable pharmaceutical formulation of the present invention is a liquid or aqueous formulation.

The liquid ophthalmic formulation of the present invention may be administered by various routes to the eye; such as topical, local ocular (subconjunctival, intravitreal, retrobulbar, intracameral), and systemic, depending on purposes. The present ophthalmic formulation is preferably administered through intravitreal injection for the treatment of ophthalmic diseases.

The invention further features ophthalmic formulations provided in a pre-filled syringe or vial, particularly suitable for intravitreal administration.

The suitable amount of administration of the liquid formulation of the present invention may vary depending on factors such as formulation methods, administration modes, age, weight, sex, and disease state of patients, foods, administration time, administration routes, excretion rates, and reaction sensitivity. Physicians with ordinary skills may easily determine and prescribe the amount of administration effective for desired therapy or prevention.

The liquid protein formulation is prepared by buffer exchange techniques like dialysis or diafiltration, wherein the excipient such as bile acid salt; can be added during the buffer exchange process to avoid any aggregation due to shear involved in processing.

The bulk processed aflibercept material was formulated using procedures mentioned below.

The aflibercept material was buffered exchanged following the dialysis procedure using tangential flow filtration. After dialysis the osmotic pressure and the pH was checked, if the pH of the sample was not within range pH units, the formulation buffer was made more acidic or basic, by repeat dialysis or by addition of HCl or NaOH, until the pH target was reached. Following pH adjustment if needed, the sample was concentrated above the target formulation concentration. The pH, osmotic pressure and the protein concentration was then measured once more. The sample was then diluted with the formulation buffer to reach the target formulation protein concentration within 10%. The protein concentration was then measured once more to ensure the diluted sample was within 10% of the target concentration. The last step was the addition of the bile acid salt (diluted in water) to the sample based on weight, if required.

In a specific preferred embodiment, the stable liquid ophthalmic formulation for intravitreal administration comprises about 40-100 mg/ml aflibercept, about 5-40 mM sodium phosphate or Histidine - Histidine HCl buffer pH of about 5.8 to about 7.0, 0.001% w/v to 0.1% w/v Sodium deoxycholate, about 5% w/v-12% w/v sucrose, and optionally about 40-150 mM sodium chloride, pH of formulation about 6.0-7.0.

The stability data of formulation containing sodium deoxycholate (Example 2), polysorbate (Example 6) and Control formulation, which is Eylea® (aflibercept) is mentioned in below Table 1.

Table 1: Comparative stability data

Parameter Time Point Control formulation Example 2 Example 6
%HMW Day 0 1.67 1.73 1.73
1 Month 1.80 1.91 1.88
2 Month 1.96 2.03 1.94
3 Month 2.01 1.74 1.98
6 Month 2.51 1.13 2.27
9 Month 5.47 3.80 3.93
%LMW Day 0 0.82 0.92 1.05
1 Month 1.06 0.85 0.39
2 Month 3.05 3.50 2.18
3 Month 2.16 8.70 1.46
6 Month 3.02 40.42 0.82
9 Month 2.37 1.41 1.97

The inventors of present invention observed that aflibercept drug product at real time conditions (2°C to 8°C) at 9 months, formulation containing sodium deoxycholate has lesser %HMWs and %LMW than the controlled formulation as well as formulation containing polysorbate resulting in stable formulation. This is unexpected results.

The following examples illustrate the invention and they do not any way limit the scope of the invention.

EXAMPLES

Example 1:
An ophthalmic liquid formulation containing 40 mg/ml Aflibercept, 10 mM, Sodium Phosphate buffer system 0.001% to 0.1% Sodium deoxycholate, 0 to 40 mM NaCl, 8-10% sucrose, and pH 6.8 ±0.3 pH, was stored at 5°C in 3 ml container and samples tested as per ICH guideline. Stability was determined by SE-HPLC.

No. Ingredient Concentration Amount/L
1. Aflibercept 40mg/mL 40 g/L
2. Sodium Phosphate monobasic, monohydrate 6.21 mM 0.852 g/L

3. Sodium phosphate dibasic, heptahydrate 3.84 mM 1.03 g/L

4. Sodium Deoxycholate 0.01% w/v 0.1 g/L
5. Sucrose 10 % w/v 100 g/L
6. Sodium Chloride 40 mM 2.34 g/L
7. Water q.s. to 1 mL q.s. to 1L
8. pH 6.8±0.3

Example 2:
An ophthalmic liquid formulation containing 40 mg/ml Aflibercept, 10 mM, Sodium Phosphate buffer system 0.001% to 0.1% Sodium deoxycholate, 8-10% sucrose, and pH 6.2 ± 0.3 pH, was stored at 5°C in 3 ml container and samples tested as per ICH guideline. Stability was determined by SE-HPLC.

No. Ingredient Concentration Amount/L
1. Aflibercept 40 mg/mL 40 g/L
2. Sodium Phosphate monobasic, monohydrate 8.23 mM 1.13 g/L
3. Sodium phosphate dibasic, heptahydrate 1.79 mM 0.48 g/L
4. Sodium Deoxycholate 0.005% w/v 0.05 g/L
5. Sucrose 9% w/v 90 g/L
6. Water q.s. to 1mL q.s. to 1L
7. pH 6.2 ± 0.3

Example 3:
An ophthalmic liquid formulation containing 40 mg/ml Aflibercept, 10 mM, Sodium Phosphate buffer system 0.001% to 0.1% Sodium deoxycholate, 8-10% sucrose, and pH 6.8 ± 0.3 pH, was stored at 5°C in 3 ml container and samples tested as per ICH guideline. Stability was determined by SE-HPLC.

No. Ingredient Concentration Amount/L
1. Aflibercept 40 mg/mL 40 g/L
2. Sodium Phosphate monobasic, monohydrate 6.21 mM 0.852 g/L

3. Sodium phosphate dibasic, heptahydrate 3.84 mM 1.03 g/L

4. Sodium Deoxycholate 0.01% w/v 0.1 g/L
5. Sucrose 9% w/v 90 g/L
6. Water q.s. to 1mL q.s. to 1L
7. pH 6.8 ± 0.3

Example 4:
An ophthalmic liquid formulation containing 40 mg/ml Aflibercept, 10 mM, Sodium Phosphate buffer system 0.001% to 0.1% Sodium cholate, 8-10% sucrose, and pH 6.2 ± 0.3 pH, was stored at 5°C in 3 ml container and samples tested as per ICH guideline. Stability was determined by SE-HPLC.

No. Ingredient Concentration Amount/L
1. Aflibercept 40 mg/mL 40 g/L
2. Sodium Phosphate monobasic, monohydrate 8.23 mM 1.13 g/L
3. Sodium phosphate dibasic, heptahydrate 1.79 mM 0.48 g/L
4. Sodium Cholate 0.01% w/v 0.1 g/L
5. Sucrose 9% w/v 90 g/L
6. Water q.s. to 1mL q.s. to 1L
7. pH 6.2 ± 0.3

Example 5:
An ophthalmic liquid formulation containing 40 mg/ml Aflibercept, 10 mM, Histidine buffer system 0.001% to 0.1% Sodium deoxycholate, 8-10% sucrose, and pH 6.2 ± 0.3 pH, was stored at 5°C in 3 ml container and samples tested as per ICH guideline. Stability was determined by SE-HPLC.

No. Ingredient Concentration Amount/L
1. Aflibercept 40 mg/mL 40 g/L
2. L Histidine 5.865 mM 0.91 g/L
3. L Histidine HCl 3.768 mM 0.794 g/L
4. Sodium Deoxycholate 0.01% w/v 0.1 g/L
5. Sucrose 10% 100 g/L
6. Water q.s. to 1mL q.s. to 1L
7. pH 6.2 ± 0.3

Example 6:
An ophthalmic liquid formulation containing 40 mg/ml Aflibercept, 10 mM, Sodium Phosphate buffer system 0.001% to 0.1% Polysorbate 20, 8-10% sucrose as anti-aggregating agent, and pH 6.2 ± 0.3 pH, was stored at 5°C in 3 ml container and samples tested as per ICH guideline. Stability was determined by SE-HPLC.

No. Ingredient Concentration Amount/L
1. Aflibercept 40 mg/mL 40 g/L
2. Sodium Phosphate monobasic, monohydrate 8.23 mM 1.13 g/L
3. Sodium phosphate dibasic, heptahydrate 1.79 mM 0.48 g/L
4. Polysorbate 20 0.03% w/v 0.3 g/L
5. Sucrose 9% w/v 90 g/L
6. Water q.s. to 1mL q.s. to 1L
7. pH 6.2 ± 0.3

,CLAIMS:
1. A stable aqueous pharmaceutical formulation comprising:
(a) a therapeutically effective amount of aflibercept,
(b) buffer
(c) bile acid salt
(d) anti-aggregating agent and optionally tonicity modifier
wherein, the pH of said formulation is about pH 6.0 to pH 7.0.

2. The formulation of claim 1, wherein the aflibercept is present in a concentration ranging from 10 mg/ml to 100 mg/ml.

3. The formulation of claim 1, wherein buffer is selected from the group consisting of acetate, histidine, phosphate, citrate, succinate, tartrate, maleate and combination thereof.

4. The formulation of claim 3, wherein the buffer is present in a concentration ranging from about 5mM to 40mM.

5. The formulation of claim 1, wherein bile acid salt is selected from sodium deoxycholate, sodium cholate or combination thereof.

6. The formulation of claim 5, wherein bile acid salt is present in a concentration ranging from about 0.001% w/v to 0.1% w/v.

7. The formulation of claim 1, wherein anti-aggregating agent is selected from the group consisting of amino acids, sugars, polyols, polymers and combinations thereof.

8. The formulation of claim 7, wherein anti-aggregating agent is present in a concentration ranging from about 5% w/v to 12% w/v.

9. The formulation of claim 1 is intravitreal injection provided in a pre-filled syringe or vial.