DESC:Field of the invention
The present invention relates to a pharmaceutical formulation of anti-interleukin-5 (anti-IL-5) antibody.

Background of the invention
IL-5 plays a role in a number of different diseases such as asthma, severe eosinophilic asthma, severe asthma, uncontrolled eosinophilic asthma, eosinophilic asthma, sub- eosinophilic asthma, chronic obstructive pulmonary disease, eosinophilic granulomatosis with polyangiitis, hypereosinophilic syndrome, nasal polyposis, bullous pemphigoid and eosinophilic esophagitis. These serious diseases affect hundreds of millions of people worldwide. At present, mepolizumab and reslizumab are approved anti-IL-5 antibody used in the treatment of IL-5 related diseases as mentioned herein above. Mepolizumab is a monoclonal antibody that binds to soluble IL-5 and blocks the soluble IL-5 from binding to its receptor. WO 2017033121 discloses approved composition of mepolizumab for the treatment of IL-5 mediated diseases.
Present invention discloses novel compositions of anti-IL-5 antibody and its related method of preparation.

Summary of the invention
The present invention provides a pharmaceutical formulation of anti-interleukin-5 (anti-IL-5) antibody. In one aspect, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody and one or more buffer(s) wherein buffer(s) is selected from tromethamine buffer, succinate buffer, acetate buffer, arginine-arginine buffer, arginine-succinate buffer, arginine-acetate buffer, arginine-glutamate buffer, aspartate buffer, sodium glutamate buffer and suitable combination thereof optionally with other suitable excipient(s). In one more aspect, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody and chelating agent selected from pentetic acid or ethylene glycol tetraacetic acid (EGTA) optionally with other suitable excipient(s). In one of the aspects, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody with other suitable excipient(s) wherein the said formulation does not contain buffer. Mepolizumab is the preferred anti-IL-5 antibody according to the present invention. In one of the aspects, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of mepolizumab and amino acid optionally with other suitable excipient(s). In further aspect, the present invention provides method of preparing anti-IL-5 antibody composition with pharmaceutical excipient(s).

Brief description of drawing
Figure 1 illustrates the rate of formation of HMW species of mepolizumab protein formulated in different buffer(s) with other excipients as mentioned in formulation 9 (F-9), formulation 12 (F-12), formulation 15 (F-15) and formulation 16 (F-16) during storage under High temperature stressed condition 50 °C ± 2 °C, RH 75 % ± 5 % as assessed by HP-SEC. Control formulation plotted herein represents rate of formation of HMW species. Control formulation comprises 100 mg / mL of mepolizumab, 0.95 mg / mL of citric acid monohydrate, 4.16 mg / mL of sodium phosphate dibasic heptahydrate, 120 mg / mL of sucrose, 0.019 mg / mL of EDTA disodium dihydrate and 0.20 mg / mL of polysorbate 80 with a pH of 6.3 referred herein after “control mepolizumab formulation” or “control formulation”.
Figure 2 illustrates the rate of formation of HMW species of mepolizumab protein formulated in different buffer(s) with other excipients as mentioned in formulation 6 (F-6), formulation 7 (F-7), formulation 8 (F-8), formulation 9 (F-9), formulation 12 (F-12), formulation 13 (F-13), formulation 14 (F-14), formulation 15 (F-15), formulation 16 (F-16), formulation 49 (F-49), formulation 50 (F-50), formulation 51 (F-51), formulation 52 (F-52) during storage under High temperature stressed condition 50 °C ± 2 °C, RH 75 % ± 5 % as assessed by HP-SEC. Control formulation plotted herein represents rate of formation of HMW species.
Figure 3 illustrates the role of DTPA in rate of formation of HMW species of mepolizumab protein formulated in different buffer(s) with other excipients as mentioned in formulation 50 (F-50), formulation 51 (F-51), formulation 52 (F-52) during storage under High temperature stressed condition 50 °C ± 2 °C, RH 75 % ± 5 % as assessed by HP-SEC.

List of abbreviations used herein the specification
Anti-IL-5: anti-interleukin-5
DTPA: Diethylenetriaminepentaacetic acid
EDTA: Ethylenediaminetetraacetic acid
EGTA: Ethylene glycol tetraacetic acid
FT cycle: Freeze-thaw cycle
HMW: High molecular weight
LMW: Low molecular weight
mM: milimolar
NA: Not available
PEG: Polyethylene glycol

Definitions
The term “pharmaceutical formulation” refers to preparations which are in such form as to permit the biological activity of the active ingredients to be unequivocally effective, and which contain no additional components which are significantly toxic to the subjects to which the formulation would be administered. The term “pharmaceutical formulation”, “formulation”, “pharmaceutical composition”, or “composition” can be used here interchangeably.
The term “Control formulation” referred herein the current specification comprises 100 mg / mL of mepolizumab, 0.95 mg / mL of citric acid monohydrate, 4.16 mg / mL of sodium phosphate dibasic heptahydrate, 120 mg / mL of sucrose, 0.019 mg / mL of EDTA disodium dihydrate and 0.20 mg / mL of polysorbate 80 with a pH of 6.3 referred herein after “control mepolizumab formulation” or “control formulation”.
In a pharmacological sense, in the context of the present invention, a “therapeutically effective amount” or “effective amount” of an anti-IL-5 antibody refers to an amount effective in the prevention or treatment of a disorder for the treatment of which the anti-IL-5 antibody is effective. A “disorder” is any condition that would benefit from treatment with the antibody. This includes chronic and acute disorders or diseases including those pathological conditions, which predisposes the subject to the disorder in question.
The term “buffer” or “buffer solution” or “buffer system” refers to generally aqueous solution comprising a mixture of an acid (usually a weak acid) and conjugate base. A buffered solution prevents change of pH of the solution due to the “buffering capacity” imparted by the “buffering agent(s)”. The pH of a “buffer solution” will change very little upon addition of a small quantity of strong acid or base due to the “buffering effect” imparted by the “buffering agent”.
The term “buffer system” comprises one or more buffering agent(s) and / or an acid/base conjugate(s) thereof, and more suitably comprises one buffering agent only and an acid/base conjugate thereof. The overall pH of the composition comprising the relevant buffer system is generally a reflection of the equilibrium concentration of each of the relevant buffering species (i.e. the balance of buffering agent(s) to acid/base conjugate(s) thereof). The buffer system may comprise dual buffers where two separate buffers are mixed to obtain target pH.
The term “buffering agent” refers to an acid or base component (usually a weak acid or weak base) of a buffer or buffer solution. A buffering agent maintain the pH of a given solution at or near to a pre-determined value, and the buffering agents are generally chosen to complement the pre-determined value. The term “buffering agent” and “buffers” can be used here interchangeably.
The term “pharmaceutical excipient” or “suitable excipient” refers to an agent that may be added to a formulation to stabilize the active drug substance in the formulated form to adjust and maintain osmolality and pH of the pharmaceutical preparations. Examples of used excipient(s) include, but are not limited to, suitable buffer(s), suitable salt(s), suitable carbohydrate(s), suitable surfactant(s), suitable amino acid(s), suitable anti-oxidant(s), suitable chelating agent(s) and combination thereof.
The “lyophilized formulation” or “freeze dried” formulation is a dosage form, which is prepared by lyophilization or freeze drying process. The lyophilization was performed with conventional lyophilization technique known in the literatures involving steps such as freezing, primary drying, secondary drying and optionally annealing.
A “stable” formulation is one in which the active substance therein i.e. protein or antibody essentially retains its physical stability and/or chemical stability and/or biological activity upon storage. Active substance according to the present invention is an anti-IL-5 antibody. Preferably, the formulation essentially retains its physical and chemical stability, as well as its biological activity upon storage. The storage period is generally selected based on the intended shelf-life of the formulation. Various analytical techniques for measuring protein stability are available in the art. Stability can be measured at a selected temperature for a selected time period. The formulation of the current invention is stable between +2 °C and +8 °C for at least one month. Furthermore, the formulation of the current invention is preferably stable following freezing (to, e.g., - 70 °C) and thawing of the formulation, for example following 1, 2, 3 or 10 cycles of freezing and thawing. Stability can be evaluated qualitatively and/or quantitatively in a variety of different ways, including evaluation of aggregate formation (for example using size exclusion chromatography, by measuring turbidity, and/or by visual inspection); by assessing charge heterogeneity using cation exchange chromatography or capillary zone electrophoresis; amino-terminal or carboxy- terminal sequence analysis; mass spectrometric analysis; SDS-PAGE analysis to compare reduced and intact antibody; peptide map (for example tryptic or LYS-C) analysis; evaluating biological activity or antigen binding function of the antibody; etc. Instability may involve any one or more of: aggregation, deamidation (e.g. Asn deamidation), oxidation (e.g. Met oxidation), isomerization (e.g. Asp isomerization), clipping/hydrolysis/fragmentation (e.g. hinge region fragmentation), succinimide formation, unpaired cysteine(s), N-terminal extension, C-terminal processing, glycosylation differences, etc.
The term “tonicity agent” or “tonicifier” or “tonicity modifier” is a compound which renders the formulation isotonic. Carbohydrate(s) such as sugar(s) or sugar alcohol (polyols), salt(s), amino acid(s), glycerin may act as a tonicity agent. The term “isotonic” is known in the art and can mean, for example, that the formulation of interest has essentially the same osmotic pressure as human blood. Isotonicity can be measured using a vapor pressure or ice-freezing type osmometer. Cryoprotectant also contribute to the tonicity of the formulations.
The term “stabilizer” as used herein refers to a compound which stabilizes an active ingredient(s) (herein anti-IL-5 antibody) under various storage condition such as various temperature condition, for example, at 25°C or at 40 °C, or at 50 °C or during freezing or thawing condition, or during various pH condition such as under acidic pH condition or neutral or basic pH condition or under real-time storage condition or during stress condition, etc. Buffer(s), carbohydrate(s) such as sugar(s) or sugar alcohol (polyols), surfactant(s), salt(s), amino acid(s) may act as a stabilizers either alone or in suitable combination.
The term “anti-oxidant” as used herein refers to a compound which prevents oxidation of active ingredient(s) (herein anti-IL-5 antibody) under storage condition. Amino acid such as methionine, proline, arginine etc. and chelating agent such as pentetic acid (DTPA), EDTA, EGTA may act as anti-oxidant.
The term “about” as used herein refers to variation in the numerical quantity that can occur, for example, through typical measuring and handling procedures used for making compounds, compositions, concentrates or formulations; through inadvertent error in these procedures; through differences in the manufacture, source, or purity of starting materials or ingredients used to carry out the methods, and other similar considerations. The term “about” also encompasses amounts that differ due to aging of a formulation with a particular initial concentration or mixture, and amounts that differ due to mixing or processing a formulation with a particular initial concentration or mixture.
The term “patient” or “subject” is used in its conventional sense to refer to a living organism suffering from or prone to a condition that can be prevented or treated by administration of a complex or a composition of the present invention, and includes animals. The term “animal” refers to a human or non-human animal, including, but not limited to, farm animals such as cattle, sheep, pigs, goats and horses; domestic mammals such as dogs and cats; laboratory animals including rodents such as mice, rats and guinea pigs; birds, including domestic, wild and game birds such as chickens, turkeys and other gallinaceous birds, ducks, geese and non-human primates, including, but not limited to, monkeys, chimpanzees and other apes and monkey species. The term does not denote a particular age. Thus, adult, juvenile and newborn individuals are of interest.
The term “comprising” or “comprises” as used herein refers to the compositions and methods include the listed elements, but do not exclude other unlisted elements.
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.

Embodiments of the invention
In first embodiment, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody and one or more buffer(s), wherein buffer(s) is selected from tromethamine buffer, succinate buffer, acetate buffer, arginine-arginine buffer, arginine-succinate buffer, arginine-acetate buffer, arginine-glutamate buffer, aspartate buffer, sodium glutamate buffer and suitable combination thereof optionally with other suitable excipient(s). In further embodiment of the first embodiment, the other suitable excipient(s) is selected from surfactant(s), carbohydrate(s), amino acid(s), salt(s), acid(s), base(s), chelating agent(s) and suitable combination thereof. In further embodiment of the first embodiment, anti-IL-5 antibody according to the present invention is mepolizumab.
In second embodiment, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody and chelating agent selected from pentetic acid or ethylene glycol tetraacetic acid (EGTA) optionally with other suitable excipient(s). In further embodiment of the second embodiment, the other suitable excipient(s) is selected from buffer(s), surfactant(s), carbohydrate(s), amino acid(s), salt(s), acid(s), base(s) and suitable combination thereof. In further embodiment of the second embodiment, anti-IL-5 antibody according to the present invention is mepolizumab.
In third embodiment, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody wherein the said formulation does not contain buffer. In a preferred embodiment of the third embodiment, the formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody in water for injection (WFI) optionally with other suitable excipient(s). In further embodiment of the third embodiment, the other suitable excipient(s) is selected from surfactant(s), carbohydrate(s), amino acid(s), salt(s), acid(s), base(s), chelating agent(s) and suitable combination thereof. In further embodiment of the third embodiment, anti-IL-5 antibody according to the present invention is mepolizumab.
In fourth embodiment, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of mepolizumab and amino acid(s) optionally with other suitable excipient(s). In further embodiment of the fourth embodiment, the other suitable excipient(s) is selected from buffer(s), surfactant(s), carbohydrate(s), salt(s), acid(s), base(s), chelating agent(s) and suitable combination thereof.
In further embodiment, the pharmaceutical formulation of the present invention is a liquid dosage form or a freeze dried dosage form or a frozen dosage form.
In a further embodiment, the present invention provides method of preparing anti-IL-5 antibody composition with pharmaceutical excipient(s).
Specific embodiments of the present invention:
a. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl;
b. 100 mg / mL of mepolizumab, 20 mM of sodium succinate;
c. 125 mg / mL of mepolizumab, 25 mM of Tris-Cl and 125 mM of sodium chloride;
d. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, pH 7.4 and 125 mM of sodium chloride;
e. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, pH 6.5 and 125 mM of sodium chloride;
f. 100 mg / mL of mepolizumab, 20 mM of sodium succinate and 0.1 % w / v of polysorbate 20;
g. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 0.1 % w / v of polysorbate 20 and 125 mM sodium chloride;
h. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 0.1 % w / v of polysorbate 20, 150 mM of L-proline and 10 mM of L-methionine;
i. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 0.1 % w / v of polysorbate 20, 125 mM of L-arginine HCl and 10 mM of L-methionine;
j. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 125 mM of sodium chloride, 10 mM of L-methionine and 1.0 % w / v PEG 4000;
k. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 50 mM of sodium chloride, 125 mM of L-arginine HCl and 0.1 % w / v of polysorbate 20;
l. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 125 mM of sodium chloride, 95 mM of L-arginine HCl, 10 mM of L-methionine and 0.02 % w / v of polysorbate 20;
m. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 50 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine and 0.1 % w / v of polysorbate 20;
n. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 125 mM of sodium chloride, 150 mM of L-proline, 10 mM of L-methionine and 0.02 % w / v of polysorbate 20;
o. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 50 mM of sodium chloride, 150 mM of L-proline, 10 mM of L-methionine and 0.1 % w / v of polysorbate 20;
p. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 50 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine and 0.1 % w / v of polysorbate 20.
q. 100 mg / mL of mepolizumab, 10 mM of sodium succinate, 112 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine and 0.05 % w / v of polysorbate 20.
r. 100 mg / mL of mepolizumab, 10 mM of sodium succinate, 112 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine, 0.2 mM DTPA and 0.05 % w / v of polysorbate 20.
s. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 50 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine, 0.2 mM DTPA and 0.1 % w / v of polysorbate 20.
t. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 125 mM of L-arginine HCl, 10 mM of L-methionine, 0.2 mM DTPA and 0.1 % w / v of polysorbate 20.

Detailed description of the invention
The present invention provides a pharmaceutical formulation of anti-interleukin-5 (anti-IL-5) antibody. The amount of anti-IL-5 antibody according to the present invention is in the range of 5.0 % w / v to 25 % w / v or its equivalent amount in mg / m L i.e. the amount of anti-IL-5 antibody according to the present invention is in the range of 50 mg / mL to 250 mg / mL. The amount of anti-IL-5 antibody according to present invention include each integer and non-integer number between the particular ranges. For example, 5.0 % w / v, 7.5 % w / v, 9.0 % w / v, 10.8 % w / v, 10.7 % w / v, 12.5 % w / v, 15 % w / v etc. The anti-IL-5 antibody according to the present invention include, but not limited to, mepolizumab, reslizumab, preferably, mepolizumab. Mepolizumab sequence has been disclosed in patent term extension application of US5693323 filed by GlaxoSmithKline LLC for mepolizumab product. In first embodiment, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody and one or more buffer(s) wherein buffer(s) is selected from tromethamine buffer or tris buffer, succinate buffer, acetate buffer, arginine-arginine buffer, arginine-succinate buffer, arginine-acetate buffer, arginine-glutamate buffer, aspartate buffer, sodium glutamate buffer and suitable combination thereof optionally with other suitable excipient(s). Buffers can be prepared by a skilled person by method known in the art. In one embodiment, the amount of buffer used herein the present invention is in the range of 0 mM to about 120 mM. In preferred embodiment, the amount of buffer used herein the present invention is in the range of 1 mM to about 120 mM. The amount of buffer in the range of about 0 mM to about 120 mM according to present disclosure include each integer and non-integer number between a particular range. For Example, the amount of buffer in the range of about 0 mM to about 120 mM includes about 0 mM, about 10 mM, about 20 mM, about 30 mM, about 40 mM, about 50 mM, about 60 mM, about 70 mM, about 80 mM, about 100 mM, about 120 mM. In one embodiment, the buffers included in the pharmaceutical formulation according to the present invention is a buffer that maintains the pH of the composition at a pH, ranging from about pH 4.0 to about pH 8.0, preferably about pH 5.0 to about pH 7.5. The pH range 4 to 8 according to the present invention includes pH 4.0, pH 5.0, pH 6.0, pH 7.0 or pH 8.0 or any non-integer number in between them (e.g., pH 6.3 or pH 6.5 or pH 6.7 or pH 6.8 or pH 7.4 or pH 7.5 and the like). The buffer(s) may act as stabilizer in the pharmaceutical formulation of the present invention. The other suitable excipient(s) of the formulation of first embodiment is selected from surfactant(s), carbohydrate(s), amino acid(s), salt(s), acid(s), base(s), chelating agent(s) and suitable combination thereof. In further embodiment of the first embodiment, anti-IL-5 antibody according to the present invention is mepolizumab.
In second embodiment, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody and chelating agent selected from pentetic acid or ethylene glycol tetraacetic acid (EGTA) optionally with other suitable excipient(s). The pentetic acid is also known as diethylenetriaminepentaacetic acid (DTPA). Pentetic acid or Ethylene glycol tetraacetic acid (EGTA) plays a role of chelating agent in the pharmaceutical formulation. In one embodiment, the amount of chelating agent(s) used herein the present invention is in the range of about 0.001 mM to about 50 mM. The amount of chelating agent(s) in the range of about 0.001 mM to about 50 mM according to present disclosure include each integer and non-integer number between a particular range. For Example, the amount of chelating agent(s) in the range of about 0.001 mM to about 50 mM includes about 0.001 mM, about 0.5 mM, about 1.0 mM, about 10 mM, about 20 mM, about 30 mM, about 40 mM, about 50 mM. The other suitable excipient(s) of formulation of the second embodiment is selected from buffer(s), surfactant(s), carbohydrate(s), amino acid(s), salt(s), acid(s), base(s) and suitable combination thereof. In further embodiment of the second embodiment, anti-IL-5 antibody according to the present invention is mepolizumab.
In a third embodiment, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of anti-interleukin-5 (anti-IL-5) antibody with other suitable excipient(s) wherein the said formulation does not contain buffer. The pharmaceutical formulation of the present invention does not contain buffer means to disclose 0 mM buffer. The other suitable excipient(s) of formulation of the third embodiment is selected from surfactant(s), carbohydrate(s), amino acid(s), salt(s), acid(s), base(s), chelating agent(s) and suitable combination thereof. In further embodiment of the third embodiment, anti-IL-5 antibody according to the present invention is mepolizumab.
In fourth embodiment, the pharmaceutical formulation of the present invention comprises therapeutically effective amount of mepolizumab and amino acid optionally with other suitable excipient(s). Examples of suitable amino acids include, but are not limited to, arginine, glycine, asparagine, glutamine, lysine, threonine, histidine, glutamic acid, aspartic acid, isoleucine, valine, leucine, alanine, phenylalanine, tyrosine, tryptophan, methionine, serine, proline, cysteine, their suitable salt form or combination of any of the above. The amino acid(s) used herein the present invention may have role as an aggregation inhibitor or as a stabilizer or as an anti-oxidant. In one embodiment, the amount of amino acid(s) used herein the present invention is in the range of 1 mM to 200 mM. The amount of amino acid in the range of about 1 mM to about 200 mM according to present disclosure include each integer and non-integer number between a particular range. For Example, the amount of amino acid in the range of about 1 mM to about 200 mM includes about 1 mM, about 10 mM, about 20 mM, about 30 mM, about 40 mM, about 50 mM, about 60 mM, about 70 mM, about 80 mM, about 100 mM, about 150 mM, about 170 mM, about 200 mM. The other suitable excipient(s) of formulation of the fourth embodiment is selected from buffer(s), surfactant(s), carbohydrate(s), salt(s), acid(s), base(s), chelating agent(s) and suitable combination thereof. Preferably, the pharmaceutical formulation prepared according to the present invention may further comprise excipient(s) selected from surfactant(s), carbohydrate(s), salt(s), acid(s), base(s) and suitable combination thereof. The present invention provides method of preparing anti-IL-5 antibody composition with suitable pharmaceutical excipient(s). In further embodiment, the pharmaceutical formulation of the present invention is a liquid dosage form or a freeze dried dosage form or a frozen dosage form, preferably liquid dosage form. In one of the embodiments, the present invention provides the pharmaceutical formulation of anti-IL-5 antibody which may be stable under 2 °C to 8 °C for at least one month. The pharmaceutical formulation of anti-IL-5 antibody according to the present invention may be stable under 2 °C to 8 °C for at least 3 months or for at least 6 months or for at least 12 months. In one of the embodiments, the pharmaceutical formulation of anti-IL-5 antibody, which may be stable under temperature, condition about +25 ?C to about +30 ?C for at least 1 month. In one of the embodiments, the pharmaceutical formulation of anti-IL-5 antibody which may be stable under stress conditions at about +40 ?C to +50 ?C. In one of the embodiments, the present invention provides pharmaceutical composition of anti-IL-5 antibody, preferably mepolizumab that has been observed to show reduced rate of formation of HMW species compared to the control formulation. The mepolizumab formulations of the present invention were observed to show reduced rate and extent of the formation of HMW species when compared to the control formulation in temperature-dependent stress study. Rate of formation of HMW species formed was observed under temperature stressed condition at 50 °C ± 2 °C, RH 75 % ± 5 % for at least 15 days. To estimate the level of aggregates formation, analytical HP-size exclusion chromatography (HP-SEC) was performed. The said analytical method used in the present invention is well known to a skilled person and a brief description of the same is provided herein below for the sake of reference.
In one of the embodiments, the pharmaceutical formulation of each embodiment of the present invention maintains stability of anti-IL-5 antibody at least after one freeze-thaw cycle, more preferably, maintains stability of anti-IL-5 antibody at least after 5 freeze- thaw cycle. In preferred embodiment, the pharmaceutical formulation of each embodiment of the present invention maintains structural integrity of mepolizumab upon multiple freeze-thaw cycles, preferably at least after one freeze-thaw cycle, more preferably, maintains structural integrity of mepolizumab at least after five freeze-thaw cycles.
Analytical methods used in the present invention:
Freeze-thaw study protocol:
In order to avoid the prolonged storage of mepolizumab between + 2 °C and + 8 °C, prior to analysis, samples were subjected to freezing and thawing processes. Samples are the mepolizumab formulations as mentioned herein the present invention.
Before initiating the freezing and thawing, all samples were stored at – 70 °C ± 10 °C. Freeze-thaw events were carried out in a forward order such that different aliquots had undergone multiple FT cycles. After completion of each FT cycle, sample was analysed for purity by HP-SEC along with FT-control sample.
For each FT cycle, freezing was performed at – 70 °C ± 10 °C for at least about 2 hours and thawing was performed rapidly at room temperature in about 10 – 15 min, manually. The FT-control (not subjected to FT) sample was stored under the same condition (+ 2 °C and + 8 °C).
To perform multiple repeated freezing and thawing cycles, each aliquots were filled with approximately 100 µL in a 0.5 mL cryo-tube as container closure system and subjected to FT study. After completion of respective FT cycles, each sample was analysed for its physiochemical properties such as purity by HP-SEC.
HP-SEC analysis:
To estimate the level of aggregates formation and level of total impurities formation, analytical HP-size exclusion chromatography (HP-SEC) was performed. The said analytical method used in the present invention is well known to a skilled person and a brief description of the same is provided below for the sake of reference.
Samples were analyzed to estimate the aggregates by HP-size exclusion chromatography (HP-SEC) using TSK gel G3000 SWXL column (7.8 mm I.D × 30 cm L). Samples were loaded and eluted isocratically in the presence of sodium phosphate buffer, at a flow rate of 0.5 mL / min. Elution was monitored with UV214 nm detection.

Detailed description of excipients which applies to each formulation of each embodiment described herein above of the present invention

Buffers
Buffers suitable for use in each of the formulations of the present invention include buffers that are compatible with the anti-IL-5 antibody, preferably mepolizumab and suitable for administration. Buffers of each of the formulations of the present invention are selected from aspartate buffer, aspartate-glutamate buffer, phosphate buffer (e.g., sodium phosphate etc), arginine-phosphate buffer, histidine buffer, arginine-arginine buffer, arginine-histidine buffer, glycine buffer, arginine buffer, citrate buffer (e.g., sodium citrate etc), arginine-citrate buffer, succinate buffer (e.g. succinic acid, sodium succinate etc), arginine-succinate buffer, acetate buffer, arginine-acetate buffer, arginine-glutamate buffer, arginine-aspartate buffer, tromethamine buffer, citrate and phosphate buffer and combinations thereof. Buffers of each of the formulations of the present invention can be prepared by a skilled person by method known in the art.

Salts
The salt according to each of the formulations of the present invention is an organic salt and inorganic salt. Organic salt refers to any salt that contains C-H bonds while inorganic salt refers to any salt that does not contain C-H bonds. Preferred salt according to each of the formulations of the present invention is inorganic salt. Inorganic salts according to the present invention may include, but are not limited to, sodium chloride, potassium chloride, potassium sulfate and sodium sulfate. However, a skilled person can also use amino acids as a salt. Salt(s) is used to maintain osmolality of the pharmaceutical formulation. It may also have a role of tonicity modifier. In one embodiment, the amount of salt(s) used herein the present invention is in the range of 0 mM to 250 mM. In preferred embodiment, the amount of salt(s) used herein the present invention is in the range of 1 mM to 250 mM. The amount of salt in the range of about 0 mM to about 250 mM according to present disclosure include each integer and non-integer number between a particular range. For Example, the amount of salt in the range of about 1 mM to about 250 mM includes about 1 mM, about 10 mM, about 20 mM, about 30 mM, about 40 mM, about 50 mM, about 60 mM, about 70 mM, about 80 mM, about 100 mM, about 125 mM, about 175 mM, about 200 mM, about 225 mM, about 250 mM. In one of the embodiments, the formulation according to the current invention may not comprise salt.

Carbohydrates
The carbohydrate according to each of the formulations of the present invention may include sugar, sugar alcohol (polyol), a derivatized sugar, an esterified sugar and sugar polymer. Carbohydrate may use as a bulking agent or tonicity modifier or stabilizer herein the present invention. Some carbohydrate of interest include, but are not limited to, for example, monosaccharides, such as fructose, maltose, galactose, glucose, D-mannose, sorbose, and the like; disaccharides, such as lactose, sucrose, trehalose and the like; polysaccharides, such as raffinose, melezitose, maltodextrins, dextrans, starches, and the like; sugar alcohol such as mannitol, xylitol, maltitol, lactitol, xylitol, sorbitol (glucitol), pyranosyl sorbitol, myoinositol, and the like. Preferred sugars suitable for use in each of the formulations of the present invention include sugars that are compatible with the pharmaceutical formulation of anti-IL-5 antibody of the present invention suitable for administration to a subject. Examples of suitable sugars include, but not limited to, sucrose, mannitol, sorbitol, maltose, trehalose, hydroxypropyl-beta-cyclodextrin (ß-HPCD), hydroxypropyl-gamma-cyclodextrin (?-HPCD) lactose, glucose, fructose, galactose, glucosamine, and the like, and suitable combinations thereof. In certain instances, the sugar is a disaccharide. For example, the disaccharide may be sucrose or trehalose. In one embodiment, the amount of sugar(s) used herein the present invention is in the range of 0 mM to 250 mM. The amount of sugar in the range of about 0 mM to about 100 mM according to present disclosure include each integer and non-integer number between a particular range. For Example, the amount of sugar in the range of about 1 mM to about 100 mM includes about 1 mM, about 10 mM, about 20 mM, about 30 mM, about 40 mM, about 50 mM, about 60 mM, about 70 mM, about 80 mM, about 100 mM, about 150 mM, about 200 mM, about 250 mM. In one of the embodiments, the formulation according to the current invention may not comprise carbohydrate(s).

Surfactants or detergents
Surfactants suitable for use in each of the formulations of the present invention include surfactants that are compatible with the pharmaceutical formulation of anti-IL-5 antibody. Surfactant(s) may use as a stabilizer or an aggregation inhibitor in the pharmaceutical formulation of the present invention. Examples of surfactants according to each of the formulations of the present invention include, but not limited to polyoxyethylensorbitan fatty acid esters (Tween), polyoxyethylene alkyl ethers (e.g. Brij), alkylphenylpolyoxyethylene ethers (e.g. Triton-X), polyoxyethylene-polyoxypropylene copolymer (e.g. Poloxamer, Pluronic), polyethylene glycol (PEG), polyethyleneimine, sodium dodecyl sulphate (SDS) and the like. In one embodiment of the present invention, the pharmaceutical formulation of anti-IL-5 antibody according to each of the formulations of the present invention comprises surfactant, wherein the surfactant is polyoxyethylensorbitan-fatty acid esters (Tweens). In another embodiment of the present invention, the pharmaceutical formulation of anti-IL-5 antibody according to each of the formulations of the present invention comprises polysorbate 20 or polysorbate 80. The polysorbate 20 is sold under the trademark Tween 20™. In another embodiment of the present invention, the amount of surfactant according to each of the formulations of the present invention is in the range of about 0.001 mg / mL to 5 mg / mL. The amount of surfactant according to each of the formulations of the present invention include each integer and non-integer number between particular concentration range. For Example, the amount of surfactant about 0.001 mg / mL to about 5 mg / mL include 0.001 mg / mL, 0.005 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.3 mg / mL, 0.5 mg / mL, 1.0 mg / mL, 2.0 mg / mL, 5.0 mg / mL, or any integer or non-integer number in between them.

Acids and Bases
Acids and bases according to the present invention can be a part of buffering agent or acid/base conjugate or as a pharmaceutical excipient. Examples of acid according to the present invention may include, but are not limited to hydrochloric acid, acetic acid, phosphoric acid, citric acid, malic acid, lactic acid, formic acid, trichloroacetic acid, succinic acid, sulfuric acid, fumaric acid, and any combinations thereof. Examples of bases according to the present invention include, but are not limited to sodium hydroxide, sodium acetate, ammonium hydroxide, potassium hydroxide, ammonium acetate, potassium acetate, sodium phosphate, potassium phosphate, sodium citrate, sodium succinate, sodium format, sodium sulfate, potassium sulfate, potassium fumarate, and any combinations thereof. In further embodiment of the present invention, the buffering agent(s) or buffering species or buffering component(s) used in each of the formulations according to the present invention is present in amount about 0.01 mg / mL to about 100 mg / mL. The concentration range of the buffering agent(s) or buffering species or buffering component(s) used in the formulations of present invention include each integer and non-integer number between particular range. For Example, the concentration range about 0.01 mg / mL to about 100 mg / mL includes 0.01 mg / mL, 0.1 mg / mL, 1.0 mg / mL, 1.08 mg / mL, 1.28 mg / mL, 2 mg / mL, 5 mg / mL, 10 mg / mL, 20 mg / mL, 50 mg / mL, 100 mg / mL or any integer and non-integer number between said concentration range.

Additional excipients of the current invention
Additional excipient(s) according to the present invention may include, but are not limited to, antimicrobial agent(s), preservative(s), anti-oxidant(s), water (e.g., water for injection (WFI)), alcohol(s), glycerin, vegetable oil(s), phospholipid(s) and any combination(s) thereof.
The amount of active substance i.e. anti-IL-5 antibody and pharmaceutical excipient(s) used herein the present invention, in % w / v or its equivalent mg / mL or its equivalent mM can be used interchangeably here in the present invention.
The composition according to the present invention finds therapeutic use in single-dose form or in multi-dose form. In one of the embodiments, the pharmaceutical formulation according to the present invention may comprises suitable preservatives. The present invention includes stable frozen, liquid or freeze dried anti-IL-5 antibody composition, preferably mepolizumab composition. The frozen, liquid or freeze dried composition of anti-IL-5 antibody, preferably mepolizumab according to the present invention comprises buffer(s), sugar(s), surfactant(s) and other suitable excipients. The lyophilization process to prepare formulation of the present invention can be performed by a skilled person using the techniques available in the art, which includes various steps such as freezing, primary drying, secondary drying and optionally annealing.
The formulations of the invention may be suitable for any use, including both in vitro and in vivo uses. In one embodiment, the formulation of the present invention is suitable for administration to a subject via a mode of administration, including, but not limited to, subcutaneous, intravenous, intradermal, transdermal, intraperitoneal, intranasal and intramuscular administration. The formulations of the invention may be used in the treatment of a disorder in a subject. Also included in the invention are devices that may be used to deliver the formulation of the invention. Examples of such devices include, but are not limited to, a glass vial, a syringe, a pen, an implant, an ampoule, a needle-free injection device and a patch.
The following non-limiting examples describe different formulations of anti-IL-5 antibody, which can be prepared as per the present invention. It will be appreciated that other excipients can also be added as necessary to these formulations and such addition of excipients are considered to be within the scope of a person skilled in the art and are to be included within the scope of the present invention. The following examples describe experiments relating to present invention.

Example
Example 1: Mepolizumab formulations with different buffers
Table 1 : Details of Formulation F-1 and F-2
Formulation No. Mepolizumab Buffer
F- 1 90 mg / mL 25 mM
Tris-Cl, pH 6.5
F- 2 100 mg / mL 20 mM
sodium succinate, pH 6.3

Mepolizumab was formulated in Tris-Cl buffer and sodium succinate buffer at pH about 6.3 ± 0.3 as mentioned in table 1. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulation was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions.

Results and discussion:
Table 2: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st Freeze-thaw 3rd
Freeze-thaw 5th
Freeze-
thaw 10th Freeze-thaw
F-1 HMW 0.80 % 0.87 % 0.86 % 1.03 % 1.09 %
Principal peak 99.06 % 99.00 % 99.00 % 98.82 % 98.77 %
LMW 0.14 % 0.13 % 0.13 % 0.16 % 0.14 %
F- 2 HMW 1.05 % 1.10 % 1.16 % 1.22 % 1.53 %
Principal peak 98.86 % 98.82 %
98.75 %
98.70 %
98.39 %

LMW 0.08 % 0.07 % 0.09 % 0.08 % 0.08 %

Results demonstrated in table 2 clearly indicate that mepolizumab in tris buffer and succinate buffer remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein.

Example 2: Mepolizumab formulation with mepolizumab > 100 mg / mL
Table 3 : Details of Formulation F-3
Formulation No. Mepolizumab Buffer Salt
F- 3 125 mg / mL 25 mM Tris-Cl, pH 7.4 125 mM
Sodium chloride

Mepolizumab was formulated in Tris-Cl buffer at pH about 7.4. Other excipient(s), like sodium chloride can be added as shown in table 3. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulations was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions
Results and discussion:
Table 4: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st Freeze-thaw 2nd Freeze-thaw 3rd
Freeze-thaw 4th
Freeze-thaw 5th Freeze-thaw
F-3 HMW 0.71 % 0.72 % 0.74 % 0.76 % 0.77 % 0.68 %
Principal peak 99.21 % 99.21 % 99.17 % 99.18 % 99.14 % 99.24 %
LMW 0.08 % 0.07 % 0.09 % 0.07 % 0.08 % 0.08 %

Results demonstrated in table 4 clearly indicate that mepolizumab at high protein concentration (> 100 mg / mL) in tris buffer remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein.

Example 3: Mepolizumab formulations with buffer and salt at different pH
Table 5 : Details of Formulation F-4 and F-5
Formulation No. Mepolizumab Buffer Salt
F-4 90 mg / mL 25 mM Tris-Cl, pH 7.4 125 mM
Sodium chloride
F-5 90 mg / mL 25 mM Tris-Cl, pH 6.5 125 mM
Sodium chloride

Mepolizumab was formulated in Tris-Cl buffer at pH about 7.4 ± 0.3 and pH about 6.3 ± 0.3 as mentioned in table 5. Other excipient(s), like sodium chloride can be added as shown in table 5. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulations was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions.

Results and discussion:
Table 6: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st Freeze-thaw 3rd
Freeze-thaw 5th Freeze-thaw 10th Freeze-thaw
F-4 HMW 0.68 % 0.77 % 0.71 % 0.79 % 0.77 %
Principal peak 99.21 % 99.11 % 99.18 % 99.07 % 99.12 %
LMW 0.11 % 0.12 % 0.11 % 0.13 % 0.11 %
F-5 HMW 0.80 % 0.85 % 0.83 % 0.96 % 0.92 %
Principal peak 99.06 % 99.03 % 99.04 % 98.90 % 98.93 %
LMW 0.14 % 0.12 % 0.13 % 0.15 % 0.14 %

Results demonstrated in table 6 clearly indicate that mepolizumab formulated in tris buffer and sodium chloride salt remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein.

Example 4: Mepolizumab formulation with buffer and surfactant
Table 7 : Details of Formulation F-6
Formulation No. Mepolizumab Buffer Surfactant
F-6 100 mg / mL 20 mM Sodium succinate,
pH 6.3 0.1 %
Polysorbate 20

Mepolizumab was formulated in sodium succinate buffer at pH about 6.3 ± 0.3 as mentioned in table 7. Other excipient(s), like polysorbate 20 can be added as shown in table 7. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulations was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions.

Results and discussion:
Table 8: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st Freeze-thaw 3rd
Freeze-thaw 5th Freeze-thaw 10th Freeze-thaw
F-6 HMW 1.06 % 1.10 % 1.19 % 1.24 % 1.49 %
Principal peak 98.86 % 98.81 % 98.73 % 98.68 %
98.43 %
LMW 0.09 % 0.09 % 0.09 % 0.08 % 0.08 %

Results demonstrated in table 8 clearly indicate that mepolizumab formulated in succinate buffer and polysorbate 20 remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein. Further, Formulation 6 was evaluated under temperature stress conditions at 50 °C ± 2 °C; RH 75 % ± 5 % for 30 days to assess the extent of degradation of mepolizumab, mainly in terms of formation of HMW species as compared to the control mepolizumab formulation. Results are illustrated in Figure 2. The formulation-6 (F-6) was observed to show reduced rate and extent of the formation of HMW species when compared to the control formulation.

Example 5: Mepolizumab formulation with buffer, surfactant and salt
Table 9 : Details of Formulation F-7
Formulation No. Mepolizumab Buffer Surfactant Salt
F-7 100 mg / mL 20 mM Sodium succinate,
pH 6.3 0.1 %
Polysorbate 20 125 mM Sodium chloride

Mepolizumab was formulated in sodium succinate buffer at pH about 6.3 ± 0.3 as mentioned in table 9. Other excipient(s), like polysorbate 20 and sodium chloride can be added as shown in table 9. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulations was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions.

Results and discussion:
Table 10: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st Freeze-thaw 3rd
Freeze-thaw 5th Freeze-thaw 10th Freeze-thaw
F-7 HMW 1.00 % 1.01 % 1.09 % 1.12 % 1.20 %
Principal peak 98.91 % 98.91 % 98.83 % 98.79 %
98.72 %

LMW 0.08 % 0.08 % 0.09 % 0.08 % 0.08 %

Results demonstrated in table 10 clearly indicate that mepolizumab formulated in succinate buffer, polysorbate 20 and sodium chloride salt remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein. Further, Formulation 7 was evaluated under temperature stress conditions at 50 °C ± 2 °C; RH 75 % ± 5 % for 30 days to assess the extent of degradation of mepolizumab, mainly in terms of formation of HMW species as compared to the control mepolizumab formulation. Results are illustrated in Figure 2. The formulation-7 (F-7) was observed to show reduced rate and extent of the formation of HMW species when compared to the control formulation.

Example 6: Mepolizumab formulation with buffer, surfactant and amino acid
Table 11 : Details of Formulation F-8 and F-9
Formulation No. Mepolizumab Buffer Surfactant amino acid amino acid
F- 8 100 mg / mL 20 mM Sodium
Succinate, pH 6.3 0.1 %
Polysorbate 20 150 mM
L-Proline 10 mM
L-methionine
F- 9 100 mg / mL 20 mM Sodium Succinate, pH 6.3 0.1 %
Polysorbate 20 125 mM
L-arginine HCl 10 mM
L-methionine

Mepolizumab was formulated in sodium succinate buffer at pH about 6.3 ± 0.3 as mentioned in table 11. Other excipient(s), like polysorbate 20 and amino acids such as L-proline, L-methionine and L- arginine HCl can be added as shown in table 11. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulations was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions.

Results and discussion:
Table 12: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st Freeze-thaw 3rd
Freeze-thaw 5th Freeze-thaw 10th Freeze-thaw
F-8 HMW 0.90 % 0.83 % 0.85 % 0.89 % 0.92 %
Principal peak 99.02 %
99.07 %
99.07 %
99.03 % 99.00 %

LMW 0.08 % 0.09 % 0.09 % 0.08 % 0.08 %
F-9 HMW 0.74 % 0.73 % 0.64 % 0.65 % 0.74 %
Principal peak 99.18 %
99.19 % 99.28 %
99.27 %
99.18 %

LMW 0.08 % 0.08 % 0.08 % 0.08 % 0.08 %

Results demonstrated in table 12 clearly indicate that mepolizumab formulated in succinate buffer, polysorbate 20 and different amino acids such as L-proline, L-methionine and L- arginine HCl as mentioned in table 11 remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein. Further, Formulation 9 was evaluated under temperature stress conditions at 50 °C ± 2 °C; RH 75 % ± 5 % for 15 days and 30 days to assess the extent of degradation of mepolizumab, mainly in terms of formation of HMW species as compared to the control mepolizumab formulation. Results are illustrated in Figure 1 and Figure 2. Further, Formulation 8 was evaluated under temperature stress conditions at 50 °C ± 2 °C; RH 75 % ± 5 % for 30 days to assess the extent of degradation of mepolizumab, mainly in terms of formation of HMW species as compared to the control mepolizumab formulation. Results are illustrated in Figure 2.
The formulation-8 (F-8) and formulation-9 (F-9) were observed to show reduced rate and extent of the formation of HMW species when compared to the control formulation.

Example 7: Mepolizumab formulation with buffer, salt, amino acid and polyethylene glycol (PEG)
Table 13 : Details of Formulation F-10
Formulation No. Mepolizumab Buffer Salt Amino acid Surfactant
F-10 90 mg / mL 25 mM
Tris-Cl
pH 6.5 125 mM
Sodium chloride 10 mM
L-methionine 1.0 %
PEG 4000

Mepolizumab was formulated in Tris-Cl buffer at pH about 6.3 ± 0.3 as mentioned in table 13. Other excipient(s), like sodium chloride, L- methionine and polyethylene glycol (PEG) 4000 can be added as shown in table 13. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulations was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions.

Results and discussion:
Table 14: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st
Freeze-thaw 3rd
Freeze-thaw 5th Freeze-thaw 10th
Freeze-thaw
F-10 HMW 0.73 % 0.75 % 0.73 % 0.84 % 0.80 %
Principal peak 99.13 % 99.12 % 99.14 % 99.00 % 99.05 %
LMW 0.14 % 0.13 % 0.13 % 0.17 % 0.15 %

Results demonstrated in table 14 clearly indicate that mepolizumab formulated in tris buffer, sodium chloride salt, L-methionine and PEG remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein.

Example 8: Mepolizumab formulation with buffer, salt, amino acid and polysorbate 20
Table 15 : Details of Formulations F-11 to F-16
Formulation No. Mepolizumab Buffer Salt Amino acid Amino acid Surfactant
F-11 90 mg / mL 25 mM
Tris-Cl
pH 6.2 50 mM
Sodium chloride 125 mM
L-arginine HCL NA 0.1 % polysorbate 20
F-12 90 mg / mL 25 mM
Tris-Cl
pH 6.4 125 mM
Sodium chloride 95 mM
L-arginine HCL 10 mM
L-methionine 0.02 % polysorbate 20
F-13 90 mg / mL 25 mM
Tris-Cl
pH 6.2 50 mM
Sodium chloride 125 mM
L-arginine HCL 10 mM
L-methionine 0.1 % polysorbate 20
F- 14 90 mg / mL 25 mM
Tris-Cl
pH 6.5 125 mM
Sodium chloride 150 mM
L-proline 10 mM
L-methionine 0.02 % polysorbate 20
F-15 100 mg / mL 20 mM SodiumSuccinate, pH 6.3 50 mM
Sodium chloride 150 mM
L-proline
10 mM
L-methionine
0.1 % polysorbate 20
F-16 100 mg / mL 20 mM SodiumSuccinate, pH 6.3 50 mM
Sodium chloride 125 mM
L-arginine HCl 10 mM
L-methionine
0.1 % polysorbate 20

Mepolizumab was formulated in Tris-Cl buffer and succinate buffer at pH about 6.3 ± 0.3 as mentioned in table 15. Other excipient(s), like sodium chloride, L-methionine, L-proline, L-arginine HCL and polysorbate 20 can be added as shown in table 15. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulations was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions. Results are summarized in table 16.

Results and discussion:
Table 16: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st
Freeze-thaw 3rd
Freeze-thaw 5th Freeze-thaw 10th
Freeze-thaw
F-11 HMW 0.85 % 0.79 % 0.78 % 0.84 % 0.86 %
Principal peak 99.09 % 99.09 % 99.12 % 99.04 % 99.04 %
LMW 0.06 % 0.12 % 0.11 % 0.11 % 0.10 %
F-12 HMW 0.58 % 0.55 % 0.54 % 0.55 % 0.53 %
Principal peak 99.28 % 99.32 % 99.34 % 99.30 % 99.35 %
LMW 0.13 % 0.12 % 0.13 % 0.15 % 0.13 %
F-13 HMW 0.81 % 0.74% 0.77 % 0.81 % 0.85 %
Principal peak 99.12 % 99.15 % 99.12 % 99.10 % 99.05 %
LMW 0.07 % 0.11 % 0.11 % 0.09 % 0.10 %
F-14 HMW 0.66 % 0.66 % 0.63 % 0.68 % 0.66 %
Principal peak 99.21 % 99.22 % 99.24 % 99.17 % 99.21 %
LMW 0.13 % 0.13 % 0.12 % 0.15 % 0.13 %
F-15 HMW 0.88 % 0.82 % 0.82 % 0.84 % 0.89 %
Principal peak 99.04 % 99.09 %
99.09 %
99.07 %
99.03 %
LMW 0.08 % 0.09 % 0.09 % 0.09 % 0.08 %
F-16 HMW 0.73 % 0.70 % 0.62 % 0.66 % 0.74 %
Principal peak 99.19 %
99.21 %
99.29 %
99.26 %
99.18 %

LMW 0.08 % 0.08 % 0.09 % 0.08 % 0.08 %

Results demonstrated in table 16 clearly indicate that mepolizumab formulated in tris buffer and succinate buffer along with other excipients as mentioned in table 15 remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein. Further, formulation 12, formulation 15 and formulation 16 were evaluated under temperature stress conditions at 50 °C ± 2 °C; RH 75 % ± 5 % for 15 days to assess the extent of degradation of mepolizumab, mainly in terms of formation of HMW species as compared to the control mepolizumab formulation. Results are illustrated in Figure 1. The formulation 12, formulation 15 and formulation 16 were observed to show reduced rate and extent of the formation of HMW species when compared to the control formulation.
In addition, a skill person can prepare mepolizumab formulations as exemplified in example 9 to example 11 as mentioned herein below.

Example 9: Mepolizumab formulations with amino acid
Table 17 : Details of Formulations F-17 to F-36
Formulation No. Active Ingredient Amino acid
F-17 Mepolizumab
25 mg / mL to 250 mg / mL Arginine
1 mM to 200 mM
F-18 Mepolizumab
25 mg / mL to 250 mg / mL Glycine
1 mM to 200 mM
F-19 Mepolizumab
25 mg / mL to 250 mg / mL Asparagine
1 mM to 200 mM
F-20 Mepolizumab
25 mg / mL to 250 mg / mL Glutamine
1 mM to 200 mM
F-21 Mepolizumab
25 mg / mL to 250 mg / mL Lysine
1 mM to 200 mM
F-22 Mepolizumab
25 mg / mL to 250 mg / mL Threonine
1 mM to 200 mM
F-23 Mepolizumab
25 mg / mL to 250 mg / mL Histidine
1 mM to 200 mM
F-24 Mepolizumab
25 mg / mL to 250 mg / mL Glutamic acid
1 mM to 200 mM
F-25 Mepolizumab
25 mg / mL to 250 mg / mL Aspartic acid
1 mM to 200 mM
F-26 Mepolizumab
25 mg / mL to 250 mg / mL Isoleucine
1 mM to 200 mM
F-27 Mepolizumab
25 mg / mL to 250 mg / mL Valine
1 mM to 200 mM
F-28 Mepolizumab
25 mg / mL to 250 mg / mL Leucine
1 mM to 200 mM
F-29 Mepolizumab
25 mg / mL to 250 mg / mL Alanine
1 mM to 200 mM
F-30 Mepolizumab
25 mg / mL to 250 mg / mL Phenylalanine
1 mM to 200 mM
F-31 Mepolizumab
25 mg / mL to 250 mg / mL Tyrosine
1 mM to 200 mM
F-32 Mepolizumab
25 mg / mL to 250 mg / mL Tryptophan
1 mM to 200 mM
F-33 Mepolizumab
25 mg / mL to 250 mg / mL Methionine
1 mM to 200 mM
F-34 Mepolizumab
25 mg / mL to 250 mg / mL Serine
1 mM to 200 mM
F-35 Mepolizumab
25 mg / mL to 250 mg / mL Proline
1 mM to 200 mM
F-36 Mepolizumab
25 mg / mL to 250 mg / mL Cysteine
1 mM to 200 mM

Mepolizumab formulations can be prepared as mentioned in table 17. Further excipients can be added which are selected from buffer(s), surfactant(s), carbohydrate(s), salt(s), acid(s), base(s), chelating agent(s) and suitable combination thereof. The amount of excipients can be selected in the ranges as described here in the present application. A person skilled in the art can prepare mepolizumab formulations and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) by following the suitable process as described in examples 1-8 of the current application, for storage at suitable temperature for further use. Further, the mepolizumab formulations as mentioned here in table 17 can be prepared by a skilled person wherein the said formulations do not contain buffer. The pH of the formulations as mentioned herein table 17 may be in the range of between about pH 5.0 to 7.5.

Example 10: Mepolizumab formulations with chelating agent
Table 18 : Details of Formulations F-37 and F-38
Formulation No. Active Ingredient Chelating agent
F-37 Mepolizumab
25 mg / mL to 250 mg / mL Pentetic acid
0.001 mM to 50 mM
F-38 Mepolizumab
25 mg / mL to 250 mg / mL Ethylene glycol tetraacetic acid (EGTA)
0.001 mM to 50 mM

Mepolizumab formulations can be prepared as mentioned in table 18. Further excipient(s) can be added which are selected from buffer(s), surfactant(s), carbohydrate(s), amino acid(s), salt(s), acid(s), base(s) and suitable combination thereof. The amount of excipients can be selected in the ranges as described here in the present application. A person skilled in the art can prepare mepolizumab formulations and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) by following the suitable process as described in examples 1-8 of the current application, for storage at suitable temperature for further use. Further, the mepolizumab formulations as mentioned here in table 18 can be prepared by a skilled person wherein the said formulations do not contain buffer. The pH of the formulations as mentioned herein table 18 may be in the range of between about pH 5.0 to 7.5.

Example 11: Mepolizumab formulations with buffer
Table 19 : Details of Formulations F-39 and F-48
Formulation No. Active Ingredient Buffer
F-39 Mepolizumab
25 mg / mL to 250 mg / mL Succinate buffer
1 mM to 120 mM
F-40 Mepolizumab
25 mg / mL to 250 mg / mL Acetate buffer
1 mM to 120 mM
F-41 Mepolizumab
25 mg / mL to 250 mg / mL Arginine-arginine buffer
1 mM to 120 mM
F-42 Mepolizumab
25 mg / mL to 250 mg / mL Arginine-succinate buffer
1 mM to 120 mM
F-43 Mepolizumab
25 mg / mL to 250 mg / mL Arginine-acetate buffer
1 mM to 120 mM
F-44 Mepolizumab
25 mg / mL to 250 mg / mL Arginine-glutamate buffer
1 mM to 120 mM
F-45 Mepolizumab
25 mg / mL to 250 mg / mL Aspartate buffer
1 mM to 120 mM
F-46 Mepolizumab
25 mg / mL to 250 mg / mL Aspartate-glutamate buffer
1 mM to 120 mM
F-47 Mepolizumab
25 mg / mL to 250 mg / mL Sodium glutamate buffer
1 mM to 120 mM
F-48 Mepolizumab
25 mg / mL to 250 mg / mL Sodium adipic buffer
1 mM to 120 mM

Mepolizumab formulations can be prepared as mentioned in table 19. Further excipient(s) can be added which are selected from surfactant(s), carbohydrate(s), amino acid(s), salt(s), acid(s), base(s), chelating agent(s) and suitable combination thereof. The amount of excipients can be selected in the ranges as described here in the present application. A person skilled in the art can prepare mepolizumab formulations and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) by following the suitable process as described in examples 1-8 of the current application, for storage at suitable temperature for further use. The pH of the formulations as mentioned herein table 19 may be in the range of between about pH 5.0 to pH 7.5.
The formulations prepared according to the examples 9 to 11 can be evaluated further under freeze-thaw analysis and under temperature stress conditions.

Example 12: Mepolizumab formulation with buffer, salt, amino acid and polysorbate 20
Table 20 : Details of Formulations F-49
Formulation No. Mepolizumab Buffer Salt Amino acid Amino acid Surfactant
F-49 100 mg / mL 10 mM Sodium succinate, pH 6.3 112 mM Sodium chloride 125 mM L-arginine HCl 10 mM L-methionine 0.05 % Polysorbate 20
Mepolizumab was formulated in succinate buffer at pH about 6.3 ± 0.3 as mentioned in table 20. Other excipient(s), like sodium chloride, L-methionine, L-arginine HCL and polysorbate 20 can be added as shown in table 20. A person skilled in the art can prepare mepolizumab formulation and can fill the formulated solution in suitable container-closure system (like vials, cartridges, syringes etc.) for storage at suitable temperature for further use. Stability of mepolizumab in the said formulations was evaluated through repeated freezing and thawing study and analyzed for the size variants of the monoclonal antibody by HP-SEC, under native conditions.

Results and discussion:
Table 21: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st
Freeze-thaw 3rd
Freeze-thaw 5th Freeze-thaw
F-49 HMW 0.43 0.41 0.40 0.40
Principal peak 99.53 99.56 99.56 99.58
LMW 0.05 0.03 0.04 0.02

Results are summarized in table 21 which clearly indicate that mepolizumab formulated in succinate buffer, L-arginine HCl, L-methionine and polysorbate 20 remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein. Further, formulation 49 was evaluated under temperature stress conditions at 50 °C ± 2 °C; RH 75 % ± 5 % for 30 days to assess the extent of degradation of mepolizumab, mainly in terms of formation of HMW species as compared to the control mepolizumab formulation. Results are illustrated in Figure 2. The formulation 49 was observed to show reduced rate and extent of the formation of HMW species when compared to the control formulation.

Example 13: Mepolizumab formulations in chelating agent with other excipients
Table 22: Details of Formulations F-50 to F-52
Formulation No. Mepolizumab Buffer Salt Amino acid Amino acid Chelating agent Surfactant
F-50 100 mg / mL 10 mM Sodium succinate, pH 6.3 112 mM sodium chloride 125 mM L-arginine HCl 10 mM L-methionine 0.2 mM DTPA 0.05 % Polysorbate 20
F-51 100 mg / mL 20 mM Sodium succinate buffer, pH 6.3 50 mm sodium chloride 125mM L-arginine HCl 10 mM L-methionine 0.2 mM DTPA 0.1 % Polysorbate 20
F-52 100 mg / mL 20 mM Na-Succinate, pH 6.3 NA 125mM L-arginine HCl 10 mM L-methionine
0.2 mM DTPA 0.1 % Polysorbate 20

Results and discussion:
Table 23: Purity of mepolizumab upon repeated freeze-thaw stress
Formulation No. % Purity by HP-SEC
Initial 1st
Freeze-thaw 3rd
Freeze-thaw 5th Freeze-thaw
F-50 HMW 0.42 0.45 0.44 0.42
Principal peak 99.55 99.50 99.50 99.57
LMW 0.03 0.04 0.06 0.02
F-51 HMW 1.16 0.86 1.00 1.09
Principal peak 98.73 99.04 98.93 98.82
LMW 0.10 0.09 0.07 0.09
F-52 HMW 1.15 0.92 1.03 1.12
Principal peak 98.76 98.98 98.87 98.78
LMW 0.10 0.10 0.10 0.10

Results are summarized in table 23 which clearly indicate that mepolizumab formulated in succinate buffer, L-arginine HCl, L-methionine, DTPA and polysorbate 20 remains stable and can withstand multiple freeze-thaw stress without exhibiting any significant degradation of protein. Further, formulation 50, 51 and 52 were evaluated under temperature stress conditions at 50 °C ± 2 °C; RH 75 % ± 5 % for 30 days to assess the extent of degradation of mepolizumab, mainly in terms of formation of HMW species as compared to the control mepolizumab formulation. Results are illustrated in Figure 2. The formulations 50, 51 and 52 were observed to show reduced rate and extent of the formation of HMW species when compared to the control formulation.
In addition, formulations 50, 51 and 52 were evaluated against formulations 49, 16 and 9 respectively under temperature stress conditions at 50 °C ± 2 °C; RH 75 % ± 5 % for 30 days to assess the extent of degradation of mepolizumab, mainly in terms of formation of HMW species. Results are illustrated in Figure 3.The formulations 50, 51 and 52 were observed to show reduced rate and extent of the formation of HMW species when compared to the formulations 49, 16 and 9 which indicates DTPA reduces rate and extent of the formation of HMW species in mepolizumab formulation.
A person skilled in the art may evaluate formulated mepolizumab as described in Example 1 to Example 13 at various temperature conditions like, between +2 ?C and +8 ?C, at about +25 ?C and +30 ?C, under stress conditions at about +40 ?C or above.

Incorporation by reference
The entire disclosure of each of the patent documents and scientific articles referred to herein is incorporated by reference for all purposes.

Equivalents
The invention may be embodied in other specific forms without departing from the spirit or essential characteristics thereof. The foregoing embodiments are therefore to be considered in all respects illustrative rather than limiting the invention described herein. Scope of the invention is thus indicated by the appended claims rather than by the foregoing description, and all changes that come within the meaning and range of equivalency of the claims are intended to be embraced therein.

,CLAIMS:We claim:-
1. A pharmaceutical formulation of anti-interleukin-5 (anti-IL-5) antibody comprises therapeutically effective amount of anti-IL-5 antibody and one or more buffer(s), wherein buffer(s) is selected from tromethamine buffer, succinate buffer, acetate buffer, arginine-succinate buffer, arginine-acetate buffer, arginine-glutamate buffer, sodium glutamate buffer and suitable combination thereof optionally with other suitable excipient(s).
2. The suitable excipient(s) as claimed in claim 1 is selected from surfactant, carbohydrate, amino acid, salt, chelating agent and suitable combination thereof.
3. A pharmaceutical formulation of anti-interleukin-5 (anti-IL-5) antibody comprises chelating agent, wherein chelating agent is selected from pentetic acid (DTPA) or ethylene glycol tetraacetic acid (EGTA) optionally with other suitable excipient(s).
4. The suitable excipient(s) as claimed in claim 3 is selected from buffer, surfactant, carbohydrate, amino acid, salt and suitable combination thereof.
5. The pharmaceutical formulation as claimed in claim 1 and claim 4 wherein the buffer is in the range of about 1 mM to about 120 mM.
6. The pharmaceutical formulation as claimed in claim 2 and claim 3 wherein the chelating agent is in the range of about 0.001 mM to about 50 mM.
7. The pharmaceutical formulation as claimed in any preceding claim wherein the amino acid is selected from arginine, glycine, asparagine, glutamine, lysine, threonine, histidine, glutamic acid, aspartic acid, isoleucine, valine, leucine, alanine, phenylalanine, tyrosine, tryptophan, methionine, serine, proline, cysteine, their suitable salt form and suitable combination thereof wherein the amount of amino acid is in the range of about 1 mM to 200 mM.
8. The pharmaceutical formulation as claimed in any preceding claim wherein the salt is selected from sodium chloride, potassium chloride, potassium sulfate and sodium sulfate wherein the amount of salt is in the range of 1 mM to 250 mM.
9. The pharmaceutical formulation as claimed in claim 2 and claim 4 wherein the carbohydrate(s) is sugar(s) or derivatized sugar(s) or polyol(s) selected from fructose, maltose, galactose, glucose, D-mannose, sorbose, lactose, sucrose, trehalose, cellobiose, raffinose, melezitose, maltodextrins, dextrans, starches, mannitol, xylitol, maltitol, lactitol, xylitol, sorbitol (glucitol), pyranosyl sorbitol, myoinositol, hydroxypropyl-beta-cyclodextrin (ß-HPCD) and suitable combination thereof wherein the amount of sugar is in the range of about 0 mM to about 100 mM.
10. The pharmaceutical formulation as claimed in any preceding claim wherein the surfactant is selected from polyoxyethylensorbitan fatty acid esters (Tween), polyoxyethylene alkyl ethers (e.g. Brij), alkylphenylpolyoxyethylene ethers (e.g. Triton-X), polyoxyethylene-polyoxypropylene copolymer (e.g. Poloxamer, Pluronic), polyethylene glycol (PEG), polyethyleneimine (PEI), sodium dodecyl sulphate (SDS) and suitable combination thereof, preferably polyoxyethylensorbitan-fatty acid esters (Tweens), more preferably selected from polysorbate 20 or polysorbate 80 or PEG wherein the amount of surfactant is in the range of 0.001 mg / mL to 5 mg / mL.
11. The anti-IL-5 antibody as claimed in any preceding claims wherein the amount of anti-IL-5 antibody is in the range of 5.0 % w / v (50 mg / mL) to 25 % w / v (250 mg / mL).
12. The anti-IL-5 antibody as claimed in any preceding claim is mepolizumab.
13. The pharmaceutical formulation as claimed in any preceding claim is a liquid dosage form or a Freeze dried dosage form or a frozen dosage form, preferably liquid dosage form.
14. The pharmaceutical formulation as claimed in any of preceding claims selected from
a. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl;
b. 100 mg / mL of mepolizumab, 20 mM of sodium succinate;
c. 125 mg / mL of mepolizumab, 25 mM of Tris-Cl and 125 mM of sodium chloride;
d. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, pH 7.4 and 125 mM of sodium chloride;
e. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, pH 6.5 and 125 mM of sodium chloride;
f. 100 mg / mL of mepolizumab, 20 mM of sodium succinate and 0.1 % w / v of polysorbate 20;
g. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 0.1 % w / v of polysorbate 20 and 125 mM sodium chloride;
h. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 0.1 % w / v of polysorbate 20, 150 mM of L-proline and 10 mM of L-methionine;
i. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 0.1 % w / v of polysorbate 20, 125 mM of L-arginine HCl and 10 mM of L-methionine;
j. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 125 mM of sodium chloride, 10 mM of L-methionine and 1.0 % w / v PEG 4000;
k. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 50 mM of sodium chloride, 125 mM of L-arginine HCl and 0.1 % w / v of polysorbate 20;
l. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 125 mM of sodium chloride, 95 mM of L-arginine HCl, 10 mM of L-methionine and 0.02 % w / v of polysorbate 20;
m. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 50 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine and 0.1 % w / v of polysorbate 20;
n. 90 mg / mL of mepolizumab, 25 mM of Tris-Cl, 125 mM of sodium chloride, 150 mM of L-proline, 10 mM of L-methionine and 0.02 % w / v of polysorbate 20;
o. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 50 mM of sodium chloride, 150 mM of L-proline, 10 mM of L-methionine and 0.1 % w / v of polysorbate 20;
p. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 50 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine and 0.1 % w / v of polysorbate 20.
q. 100 mg / mL of mepolizumab, 10 mM of sodium succinate, 112 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine and 0.05 % w / v of polysorbate 20.
r. 100 mg / mL of mepolizumab, 10 mM of sodium succinate, 112 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine, 0.2 mM DTPA and 0.05 % w / v of polysorbate 20.
s. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 50 mM of sodium chloride, 125 mM of L-arginine HCl, 10 mM of L-methionine, 0.2 mM DTPA and 0.1 % w / v of polysorbate 20.
t. 100 mg / mL of mepolizumab, 20 mM of sodium succinate, 125 mM of L-arginine HCl, 10 mM of L-methionine, 0.2 mM DTPA and 0.1 % w / v of polysorbate 20.
15. The pharmaceutical formulation as claimed in any of preceding claims maintains structural integrity of mepolizumab upon multiple freeze-thaw cycles, preferably at least after one freeze-thaw cycle, more preferably, at least after five freeze-thaw cycles.
16. The pharmaceutical formulation as claimed in any of preceding claims maintains stability under 2 °C to 8 °C storage condition for at least one month.
17. The pharmaceutical formulation as claimed in any preceding claim has a pH in the range of about pH 4.0 to about pH 8.0, preferably about pH 5.0 to about pH 7.5.
18. The pharmaceutical formulation as claimed in any of preceding claims is administered to a subject by subcutaneous, intravenous, intradermal, transdermal, intraperitoneal or intramuscular administration.

Dated this 14th day of September 2022.

(HARIHARAN SUBRAMANIAM)
IN/PA-93
Of SUBRAMANIAM & ASSOCIATES
ATTORNEYS FOR THE APPLICANTS