WO2016162819A1

WO2016162819A1 - Stable aqueous pharmaceutical composition of anti-tnf alpha antibody

Info

Publication number: WO2016162819A1
Application number: PCT/IB2016/051971
Authority: WO
Inventors: Paresh VADGAMA; Kedar Gokhale; Rustom Sorab MODY; Deepali SONAWANE
Original assignee: Lupin Ltd
Current assignee: Lupin Ltd
Priority date: 2015-04-07
Filing date: 2016-04-07
Publication date: 2016-10-13
Anticipated expiration: 2017-10-07

Abstract

The present invention relates to the stable aqueous pharmaceutical compositions of Adalimumab or its biosimilar or its biobetter. More particularly, it relates to the stable aqueous pharmaceutical compositions of Adalimumab essentially free of surfactant. It also relates to the methods of manufacturing the pharmaceutical composition, method of administration and kits containing the same.

Description

STABLE AQUEOUS PHARMACEUTICAL COMPOSITION OF

ANTI-TNF ALPHA ANTIBODY

Field of Invention

The invention provides stable aqueous pharmaceutical compositions of Adalimumab or its biosimilar or its biobetter which is suitable for long term storage. More particularly the suitable stable aqueous pharmaceutical compositions comprising Adalimumab which is free of surfactant or essentially free of surfactant. The invention also provides methods of manufacturing the pharmaceutical composition, method of administration and kits containing the same.

Background of Invention

Tumor necrosis factor (TNF) alpha is a cytokine that promotes the inflammation and its associated signs by binding to its receptor. It is produced by macrophages and many other immune cells. It is involved in pathogenesis of many inflammatory disorders like rheumatoid arthritis, psoriatic arthritis, SLE, Crohn's disease etc. Hohmann et al (Hohmann et al. 1989 Biol Chem. 25, 14927-34) identified 2 distinct receptors of TNF-alpha which are present on different cell type's viz. myeloid cells and epithelial cells. Using monoclonal antibodies, Brockhaus et al (Brockhaus et al. 1990 Proc Natl Acad Sci U S A., 87(8), 3127-31) demonstrated that both TNF-alpha and beta bind to both the receptors with high affinity.

Tumor necrosis factor-alpha (TNF-alpha) is a central regulator of inflammation, and TNF- alpha antagonists may be effective in treating inflammatory disorders in which TNF-alpha plays an important pathogenic role. Inhibition of TNF has proven to be an effective therapy for patients with rheumatoid arthritis and other forms of inflammatory disease including psoriasis, psoriatic arthritis, and ankylosing spondylitis, inflammatory bowel disease. One such TNF-alpha antagonist is Adalimumab. Adalimumab is a recombinant human IgGl monoclonal antibody specific for human tumor necrosis factor (TNF). It is produced using phage display technology resulting in an antibody with human derived heavy and light chain variable regions and human IgGl: k constant regions. It is also known as D2E7. Adalimumab has been described in US6090382.

It is produced by recombinant DNA technology in a mammalian cell expression system and is purified by a process that includes specific viral inactivation and removal steps. It consists of 1330 amino acids and has a molecular weight of approximately 148 kilodaltons. Adalimumab interferes with TNF and acts as a TNF inhibitor due to which it can be used as a biopharmaceutical to treat autoimmune diseases such as rheumatoid arthritis, psoriatic arthritis, Crohn's disease, plaque psoriasis, juvenile idiopathic arthritis, ankylosing spondylitis, ulcerative colitis and hidradenitis suppurativa.

Dosage of Adalimumab depends on the disease, severity of condition, patient's clinical history, and response to the (prior) therapy, and will be adjusted and monitored by a physician. The pharmaceutical composition may be administered parenterally, such as subcutaneously, intramuscularly, intravenously, intraperitoneally, intracerebrospinally, intraarticularly, intrasynovially and/or intrathecally by either bolus injection or continuous infusion.

Commercially, Adalimumab is available in both single-use prefilled pen and single-use pen in aqueous form.

The most important feature of a composition is to help the protein to retain its structural conformation or its activity. The stability of protein in a composition can be related with long-term storage. It is understood to mean that the active polypeptide of the pharmaceutical composition does not substantially lose its activity as compared to the composition at the beginning of storage. All polypeptides have an Isoelectric Point (pi), which is generally defined as the pH at which a polypeptide carries no net charge. It is known in the art that protein solubility is typically lowest when the pH of the solution is equal to the isoelectric point (pi) of the protein. The melting temperature (Tm) of the Fab domain of a protein is a good indicator of the thermal stability of a protein and may further provide an indication of the shelf-life. Tm values of proteins determined by differential scanning calorimetry (DSC), give insight into heat-induced changes in protein conformation, mechanisms of protein unfolding and stabilization in solution. A lower Tm indicates less stability of a protein in the given solution, whereas a higher Tm indicates a better stability of the protein. The Tm of the protein will vary based on the formulation composition which in turn reflects its stability in respective formulation.

During long term storage, aqueous composition of proteins can lose active protein due to aggregation or degradation. Aggregation of the protein can lead to immunogenicity and is undesirable. Since the high concentration of Adalimumab used in the composition therefore, there is a likely possibility of protein aggregation during long term storage. To improve the stability of the protein either the concentration of the existing excipients can be varied or new excipients can be added to modify the composition.

Surfactant is extensively used and also considered as a most desirable choice to skilled person in order to maintain the stability or prevent aggregation of protein molecule.

US Patent No US8216583; US8916158; US8916157; US8802100; US8911741; US8932591; US20150191538, PCT applications WO2004016286; WO2014039903; WO2014099636 and WO2013164837 disclose pharmaceutical compositions of Adalimumab comprising a buffer, an isotonicity agent, a stabilizer and at least a surfactant. There is still need in the art to develop an economic and effective formulation in order to stabilize Adalimumab for long term storage. Therefore herein the present invention provides a formulation free or essentially free of surfactant to achieve the stability of Adalimumab. Summary of the Invention

In an embodiment, the invention is related to a stable aqueous pharmaceutical composition comprising Adalimumab, buffer system and said composition is free or essentially free of surfactant. In certain embodiments, the invention is related to a stable aqueous pharmaceutical composition comprising Adalimumab, buffer system, a tonicity agent, stabilizer and optionally an anti-aggregating agent wherein the said composition is free or essentially free of surfactant. In certain embodiments the present invention provides a stable aqueous pharmaceutical composition of Adalimumab free of surfactant or essentially free of surfactant which exhibits a long term stability.

In certain embodiments, the present invention provides a stable aqueous pharmaceutical composition buffered between pH 4 to 8.

In yet another embodiment, the composition may further by lyophilized.

In yet another embodiment, the invention is related to a stable aqueous pharmaceutical composition of Adalimumab free or essentially free of a surfactant comprising a buffer system selected from histidine, succinate, phosphate, citrate, acetate, maleate and tartrate buffers or combination thereof; tonicity agent is selected from sodium chloride or potassium chloride; stabilizer is selected from one or more polyols, sugars, cyclodextrins and optionally other excipients selected from stabilizers or anti-aggregating agents such as amino acids and having pH 4 to 8.

In yet another embodiment, the invention is related to a stable aqueous pharmaceutical composition of Adahmumab free or essentially free of a surfactant comprising buffer system comprises the combination of phosphate and citrate buffer selected from the group essentially consisting of sodium dihydrogen phosphate dihydrate, disodium hydrogen phosphate dihydrate, trisodium citrate dihydrate, citric acid monohydrate; sodium chloride; mannitol; optionally anti-aggregating agent is L-arginine hydrochloride; and having pH 5.2 ± 0.3. In another embodiment the stable aqueous pharmaceutical composition comprising Adahmumab in the concentration of more than 45 mg/ml and preferably about 45 mg/ml to about 160 mg/ml of Adahmumab; buffer system comprises about 1 mM to about 50 mM of sodium dihydrogen phosphate dihydrate, about 0.1 mM to about 50 mM of disodium hydrogen phosphate dihydrate, about 0.1 mM to about 50 mM of trisodium citrate dihydrate, about 1 mM to about 100 mM of citric acid monohydrate; about 10 mM to about 300 mM of sodium chloride; about 10 mM to about 100 mM of mannitol; about 5 mM to about 100 mM of L-arginine monohydrochloride and having pH 5.2 ± 0.3.

In another embodiment the stable aqueous pharmaceutical composition comprising Adahmumab in the concentration of more than 45 mg/ml and preferably about 45 mg/ml to about 160 mg/ml of Adahmumab; buffer system comprises about 1 mM to about 50 mM of sodium dihydrogen phosphate dihydrate, about 0.1 mM to about 50 mM of disodium hydrogen phosphate dihydrate, about 0.1 mM to about 50 mM of trisodium citrate dihydrate, about 1 mM to about 100 mM of citric acid monohydrate; about 10 mM to about 300 mM of sodium chloride; about 10 mM to about 100 mM of mannitol; about 5 mM to about 100 mM of glycine and having pH 5.2 ± 0.3.

In yet another embodiment, the invention is related to the method of treating a disease using the stable aqueous pharmaceutical composition of the present invention wherein the disease is selected from rheumatoid arthritis, psoriatic arthritis, Crohn's disease, plaque psoriasis, juvenile idiopathic arthritis, active ankylosing spondylitis, ulcerative colitis and hidradenitis suppurativa.

In another embodiment the invention is related to a kit or a container containing the pharmaceutical composition of the invention.

The details of one or more embodiments of the invention set forth below are illustrative in nature only and not intended to limit to the scope of the invention. Other features, objects and advantages of the inventions will be apparent from the description and claims.

Detail Description of Invention

Definition:

As used herein, the term "surfactant" refers to an excipient which inhibits the aggregation of Adalimumab. The example of surfactant comprises but not limited to polysorbate 20, polysorbate 80, Brij 35, Triton X-10, Pluronic F127, and sodium doceyl Sulfate (SDS) and Pluronic F68 (poloxamer 188)

As used herein, the term "without" or "free" refers to a composition contains less than 0.1 mM surfactant or less than 0.01 mM or less than 0.0001 mM surfactant. The term "without" or "free" are interchangeable.

The term "essentially free" means that either no surfactant is present or only minimal, trace amounts of the surfactant is present which do not have any substantial impact on the properties of the composition. If reference is made to no amount of a surfactant, it should be understood as "no detectable amount".

As used herein, the term "buffer system" refers to a buffered solution that resists changes in pH by the action of its acid-base conjugate components. The example of suitable buffers system includes but are not limited to phosphate buffers, citrate-buffers, histidine -buffers, succinate-buffers, acetate -buffers, tartrate buffers, tromethamine buffers and the combinations thereof.

As used herein, the term "long-term storage" or "long term stability" is understood to mean that the pharmaceutical composition can be stored for three months or more, for six months or more, and preferably for one year or more, most preferably a minimum stable shelf life of at least two years. Generally speaking, the terms "long term storage" and "long term stability" further include stable storage durations that are at least comparable to or better than the stable shelf typically required for currently available commercial formulations of Adalimumab, without losses in stability that would render the formulation unsuitable for its intended pharmaceutical application. Long-term storage is also understood to mean that the pharmaceutical composition is stored either as a liquid at 2-8° C, or is frozen, e.g., at -20° C, or colder. It is also contemplated that the composition can be frozen and thawed more than once.

The term "stable" with respect to long-term storage is understood to mean that Adalimumab contained in the pharmaceutical compositions does not lose more than 20%, or more preferably 15%, or even more preferably 10%, and most preferably 5% of its activity of the composition at the beginning of storage.

The invention provides an improved stable aqueous pharmaceutical composition of Adalimumab free or essentially free of a surfactant. More particularly, the invention relates to the stable aqueous pharmaceutical composition of Adalimumab free or essentially free of a surfactant comprising a buffer system, a tonicity agent and a stabilizer and optionally other excipients selected from stabilizers or anti-aggregating agents which exhibits a lower degradation potential.

In an embodiment of the invention, an improved stable aqueous pharmaceutical composition of Adalimumab comprising a buffer system, a tonicity agent and a stabilizer and optionally other excipients which can be lyophilized wherein the composition is free of surfactant or essentially free of surfactant.

In embodiment, the stable aqueous pharmaceutical composition comprises a buffer system which plays critical role to regulate the stability of formulation and therefore selection of suitable buffer system is highly desirable to obtain the surfactant free stable formulation. The buffer system is selected from phosphate buffers, citrate -buffers, histidine -buffers, succinate - buffers, acetate -buffers, tartrate buffers, tromethamine buffers and the combinations thereof. In a certain embodiment the suitable buffer system comprises of a phosphate buffer. In preferred embodiment the suitable buffer system comprises a phosphate-citrate buffer.

In embodiment the pharmaceutical composition comprises; Adalimumab; buffer system comprises the combination of phosphate and citrate buffer selected from the group essentially consisting of sodium dihydrogen phosphate dihydrate, disodium hydrogen phosphate dihydrate, trisodium citrate dihydrate, citric acid monohydrate. In another embodiment, the stable aqueous pharmaceutical composition comprises tonicity agent(s) selected from sodium chloride or potassium chloride. In preferred embodiment the tonicity agent is sodium chloride.

In another embodiment, the stable aqueous pharmaceutical composition comprises one or more suitable stabilizers. The suitable stabilizer is selected from polyols, sugars, cyclodextrins and the combinations thereof.

In a foregoing embodiment the polyols selected from mannitol, sorbitol, dextran, glycerol, arabitol, propylene glycol, polyethylene glycol and suitable combinations thereof. In preferred embodiment the suitable polyol is mannitol.

In foregoing embodiment the sugar is a monosaccharide or an oligosaccharide. Monosaccharide sugars are selected from glucose, fructose, galactose, mannose, sorbose, ribose, deoxyribose and the like or amino sugars, like neuraminic acid and suitable combinations thereof. An oligosaccharide is selected from sucrose, trehalose, lactose, maltose and raffinose and suitable combinations thereof.

In foregoing embodiment cyclodextrins or derivative thereof are selected from to a- cyclodextrin, β-cyclodextrin, γ-cyclodextrin and suitable combinations thereof. In another embodiment, the stable aqueous pharmaceutical composition comprises optionally other excipients selected from stabilizers or anti-aggregating agents. In such embodiment, other excipients includes but not limited to amino acids which can be used as stabilizers or anti-aggregating agents. In foregoing embodiment, the amino acid is selected from arginine, glycine, lysine, histidine, glutamic acid, aspartic acid, isoleucine, leucine, alanine, phenylalanine, tyrosine, tryptophane, methionine, serine, proline, cysteine/cystine and suitable combination thereof. In a preferred embodiment the amino acid is arginine. In more preferred embodiment amino acid is arginine monohydrochloride. In another preferred embodiment the amino acid is glycine.

In another embodiment, the stable aqueous pharmaceutical composition comprises pH value ranging from 4.0 to about 8.0. In a preferred embodiment the pH of formulation is maintained in the range of about 4.9 to 5.5. In a more preferred embodiment the pH is maintained to about 5.2.

In another embodiment, the stable aqueous pharmaceutical composition comprises Adalimumab generally present in a therapeutic amount of up to 160 mg/mL. In a preferred embodiment the therapeutic amount is about 1 mg/mL to about 100 mg/mL. In a more preferred embodiment the therapeutic amount is about 1 mg/mL to about 50 mg/mL.

In another embodiment, the stable aqueous pharmaceutical composition comprises suitable buffer system generally present in an amount of up to lOOmM. In a preferred embodiment the amount is about lmM to about 50 mM. In a more preferred embodiment the amount is about 15 mM to about 25 mM.

In another embodiment, the stable aqueous pharmaceutical composition comprises suitable tonicity agent generally present in an amount below 500mM. In a preferred embodiment the amount is about 30mM to about 150mM. In a more preferred embodiment the amount is about 60mM to about 120mM.

In another embodiment, the stable aqueous pharmaceutical composition comprises suitable stabilizer generally present in an amount below 500mM. In a preferred embodiment the amount is about lmM to about 200mM. In a more preferred embodiment the amount is about lOmM to about lOOmM.

In another embodiment, the stable aqueous pharmaceutical composition comprises suitable stabilizer or anti-aggregating agent in an amount below 500 mM. In a preferred embodiment the amount is about lmM to about lOOmM. In a more preferred embodiment the amount is about lOmM to about 50mM.

In certain embodiment stable aqueous pharmaceutical composition comprises free or essentially free of surfactant comprises 5 mg/mL to 160 mg/mL of Adalimumab, about 1 mM to 100 mM of phosphate and/or citrate buffer, 5 mM to 500 mM of sodium chloride and 5 mM to 500 mM mannitol and optionally lmM to about lOOmM L-arginine monohydrochloride or glycine and having a pH range of 4 to 8.

In certain embodiment stable aqueous pharmaceutical composition comprises free or essentially free of surfactant comprises 5 mg/mL to 160 mg/mL of Adalimumab, about 1 mM to 100 mM of phosphate and 1 mM to 100 mM citrate buffer, 5 mM to 500 mM of sodium chloride and 5 mM to 500 mM mannitol, and optionally lmM to about lOOmM L-arginine monohydrochloride or glycine and having a pH range of 4 to 8. In certain embodiment stable aqueous pharmaceutical composition comprises free or essentially free of surfactant comprises 5 mg/mL to 160 mg/mL of Adalimumab, about 1 mM to 100 mM phosphate buffer, 5 mM to 500 mM of sodium chloride and 5 mM to 500 mM mannitol, and optionally ImM to about lOOmM L-arginine monohydrochloride or glycine and having a pH range of 4 to 8.

In certain embodiment stable aqueous pharmaceutical composition comprises free or essentially free of surfactant comprises 5 mg/mL to 160 mg/mL of Adalimumab, about 1 mM to 100 mM citrate buffer, 5 mM to 500 mM of sodium chloride and 5 mM to 500 mM mannitol, and optionally ImM to about lOOmM L-arginine monohydrochloride or glycine and having a pH range of 4 to 8.

In certain embodiment stable aqueous pharmaceutical composition comprises free or essentially free of surfactant comprises 5 mg/mL to 160 mg/mL of Adalimumab, about 1 mM to 100 mM of phosphate and/or citrate buffer, 5 mM to 500 mM of sodium chloride, 5 mM to 500 mM mannitol and 5 mM to 100 mM of L-arginine monohydrochloride and having a pH range of 4 to 8.

In certain embodiment stable aqueous pharmaceutical composition comprises free or essentially free of surfactant comprises 5 mg/ml to 160 mg/mL of Adalimumab, about 1 mM to 100 mM of phosphate and/or citrate buffer, 5 mM to 500 mM of sodium chloride, 5 mM to 500 mM mannitol and 5 mM to 100 mM of glycine and having a pH range of 4 to 8. The present invention found that use of buffer system with at least more than one buffer like phosphate and citrate buffer in suitable concentration significantly reduced or completely eliminated the requirement of surfactant and provides long term stable Adalimumab composition. The invention further describes the use of sodium chloride as a tonicity agent, polyol as a stabilizing agent and optionally arginine or glycine as other excipients in combination with phosphate and citrate buffer to provide lone term stability in absence of surfactant. In embodiment the stable aqueous pharmaceutical composition exhibiting a long term stability and free or essentially free of surfactant comprising; Adalimumab; buffer system comprises the combination of phosphate and citrate buffer selected from the group essentially consisting of sodium dihydrogen phosphate dihydrate, disodium hydrogen phosphate dihydrate, trisodium citrate dihydrate, citric acid monohydrate; sodium chloride; mannitol; optionally anti-aggregating agent is L-arginine monohydrochloride and having pH 5.2 ± 0.3.

Accordingly to a certain embodiment of the present invention relates to the stable aqueous pharmaceutical composition comprising:

a) about 45 mg/ml to about 160 mg/ml of Adalimumab;

b) buffer system comprises about 1 mM to about 50mM of sodium dihydrogen phosphate dihydrate about 0.1 mM to about 50 mM of disodium hydrogen phosphate dihydrate about 0.1 mM to about 50 mM of trisodium citrate dihydrate, about 1 mM to about 100 mM of citric acid monohydrate and having pH 5.2 ± 0.3;

c) about 10 mM to about 300 mM of sodium chloride;

d) about 10 mM to about 100 mM of mannitol;

e) about 5 mM to about 100 mM of L-arginine monohydrochloride and

having pH 5.2 ± 0.3

a) about 45 mg/ml to about 160 mg/ml of Adalimumab;

b) buffer system comprises about 1 mM to about 50 mM of sodium dihydrogen phosphate dihydrate, about 0.1 mM to about 50 mM of disodium hydrogen phosphate dihydrate, about 0.1 mM to about 50 mM of trisodium citrate dihydrate, about 1 mM to about 100 mM of citric acid monohydrate;

c) about 10 mM to about 300 mM of sodium chloride;

d) about lOmM to about 100 mM of mannitol;

e) about 5 mM to about 100 mM of glycine and

having pH 5.2 ± 0.3 Accordingly to a certain embodiment of the present invention relates to the stable aqueous pharmaceutical composition comprising:

a) 50 mg/ml of Adalimumab;

b) buffer system comprises 5.5 mM of sodium dihydrogen phosphate dihydrate, 8.6 mM of disodium hydrogen phosphate dihydrate, 1 mM of trisodium citrate dihydrate, 6.2 mM of citric acid monohydrate;

c) 105.4mM of sodium chloride;

d) 65.9mM of Mannitol;

e) 23.7 mM of L-arginine monohydrochloride and

having pH 5.2 ± 0.3

a) 50 mg/ml of Adalimumab;

b) buffer system comprises 5. 5 mM of sodium dihydrogen phosphate dihydrate, 8.6 mM of disodium hydrogen phosphate dihydrate, 1 mM of trisodium citrate dihydrate, 6.2 mM of citric acid monohydrate;

c) 105.4mM of sodium chloride;

d) 65.9mM of mannitol;

e) 23.7 mM of glycine and

having pH 5.2 ± 0.3

In foregoing embodiment the invention provides pharmaceutical composition essentially comprising of Adalimumab free or essentially free of surfactant.

The pharmaceutical composition is a sterile and stable for long period of time at 2-8 C. In certain embodiment the pharmaceutical composition is a sterile and stable for long period of time at 25 C. In certain embodiment the pharmaceutical composition is a sterile and stable for long period of time at 40 C. In certain embodiments the pharmaceutical composition is stable at non-refrigerated temperature for extended period of time. In certain embodiment pH of the formulation is maintained about 5.2 ± 0.3 for at least 15 days, preferably one month, more preferably 3 months most preferably six months to two years.

In certain embodiment formulation comprises impurities below 10% or preferably below 5% or more preferably below 1% for at least 15 days, preferably one month, more preferably 3 months and most preferably six months to two years.

In certain embodiment formulation comprises acidic variant below 25% or preferably below 15% or more preferably below 10% for at least 15 days, preferably one month, more preferably 3 months and most preferably six months to two years. In certain embodiment formulation maintains purity at least 75% or preferably more than 85% for at least 15 days, preferably one month, more preferably 3 months and most preferably six months to two years.

In another embodiment, the invention relates to a kit comprising Adalimumab; buffer system comprises the combination of phosphate and citrate buffer selected from the group essentially consisting of sodium dihydrogen phosphate dihydrate, disodium hydrogen phosphate dihydrate, trisodium citrate dihydrate, citric acid monohydrate and having pH 5.2 ± 0.3; sodium chloride; mannitol; optionally anti-aggregating agent is L-arginine monohydrochloride.and instructions for use of the present composition.

In a preferred embodiment, the composition is contained in a pre-filled syringe. In a further preferred embodiment, the composition is contained in a pre-filled vial. In Another preferred embodiment is a pre-filled syringe within an autoinjector. The kit may comprise one or more unit dosage forms containing the pharmaceutical composition of the invention.

Any suitable syringe or vial or cartridge may be used. The kit may also comprise of the pharmaceutical composition according to the invention in another secondary container, such as in an autoinjector. The prefilled syringe may contain the composition in aqueous form. Described syringe may be further supplied with an autoinjector or pen, which often is a disposable article for single use only, and may have a volume between 0.1 and 1 mL. The described vial may contain the composition in lyophilised or aqueous state, and may serve as a single or multiple use devices. The vial may have a volume between 1 and 10 mL. The invention further relates to a stable pharmaceutical composition, wherein the composition is liquid or lyophilized. The invention is further related to a stable pharmaceutical composition in a pre-filled syringe, vial or cartridge.

In another embodiment, the method may further comprise a lyophilization step, which may be before or after adding the at least one tonicity modifier, and/or an excipient as defined above.

The active ingredient Adalimumab, which was used in the described examples of the present invention, is produced from CHO cells by using recombinant DNA technology method which is well known in the general art. The CHO cells were cultured in a fed-batch process known in the art. Adalimumab was purified from the cell free harvest by standard purification and filtration process known in the art including affinity chromatography and further chromatographic and filtration steps. All patents, patent applications and publications cited in this application are hereby incorporated by reference in their entirety for all purposes to the same extent as if each individual patent, patent application or publication were so individually denoted.

The examples are only for further illustrative purpose of the invention and are not intended to be limiting. Although certain embodiments and examples have been described in detail above, those having ordinary skill in the art will clearly understand that many modifications are possible in the embodiments and examples without departing from the teachings thereof. Experimental Section

Example 1: Process for Preparation of stable pharmaceutical composition of

Adalimumab

The drug substance of purified Adalimumab is obtained and formulated by using various excipients in the present example which is shown in Table 1. The concentration of 50 mg/ml Adalimumab is formulated with excipients in absence of polysorbate 80. Only phosphate buffer is used in Formulation 2 along with other excipients sodium chloride, mannitol at pH of 5.2 ± 0.3 adjusted by 2M sodium hydroxide as mentioned in Table 1 and two buffers phosphate and citrate buffer are used in formulation 1 with other excipients sodium chloride, mannitol at pH of 5.2 ± 0.3 adjusted by 2M sodium hydroxide. Both formulation 1 and formulation 2 lack surfactant polysorbate 80.

Table 1 : Composition of the formulations

Ingredients Formulation 1 f ormulation 2

Adalimumab 50 mg/mL 50 mg/mL

Polysorbate 80 - -

Sodium dihydrogen phosphate, 0.86 mg/mL 3.243 mg/mL

dihydrate

Disodium hydrogen phosphate, 1.53 mg/mL 0.037 mg/mL

dihydrate

Trisodium citrate dihydrate 0.30 mg/mL -

Citric acid monohydrate 1.30 mg/mL -

Sodium Chloride 6.16 mg/mL 6.16 mg/mL

Mannitol 12 mg/mL 12 mg/mL

PH 5.2 ± 0.3 5.2 ± 0.3

Example 2: Process for Preparation of stable pharmaceutical composition of

Adalimumab

The drug substance of purified Adalimumab is obtained and formulated by using various excipients in the present example which is shown in Table 2. The concentration of 50 mg/mL Adalimumab is formulated with excipients in absence of polysorbate80. Only phosphate buffer is used in Formulation 4 along with other excipients sodium chloride, mannitol, L-Arginine HC1 at pH5.2 ± 0.3 adjusted by 2M sodium hydroxide as mentioned in Table 2 and two buffers phosphate and citrate buffer are used in formulation 3 with other excipients sodium chloride, mannitol, L-Arginine HC1 at pH of 5.2 ± 0.3 adjusted by 2M sodium hydroxide. Both formulation 3 and formulation 4 lack surfactant polysorbate 80.

Table 2: Composition of the formulations

The stability data of are summarized for each condition in below tables: Table 2a

Table 2b

Table 2c

Example 3: Process for Preparation of stable pharmaceutical composition of

Adalimumab

The drug substance of purified Adalimumab is obtained and formulated by using various excipients in the present example which is shown in Table 3.

The concentration of 50 mg/mL Adalimumab is formulated with excipients in absence of polysorbate 80. Only phosphate buffer is used in Formulation 6 along with other excipients sodium chloride, mannitol, Glycine at pH of 5.2 ± 0.3 adjusted by 2M sodium hydroxide as mentioned in Table 3 and two buffers phosphate and citrate buffer are used in formulation 5 with other excipients sodium chloride, mannitol, Glycine at pH5.2 ± 0.3 adjusted by 2 M sodium hydroxide. Both formulation 5 and formulation 6 lack surfactant polysorbate 80.

Table 3: Composition of the selected formulations

Ingredients Formulation 5 Formulation 6

Adalimumab 50 mg/mL 50 mg/mL

Polysorbate 80

Sodium dihydrogen 0.86 mg/mL 3.243 mg/mL

phosphate, dihydrate

Disodium hydrogen 1.53 mg/mL 0.037 mg/mL

phosphate, dihydrate

Trisodium Citrate 0.30 mg/mL

dihydrate

Citric acid monohydrate 1.30 mg/mL

Sodium Chloride 4.75 mg/mL 4.75 mg/mL

Mannitol 12 mg/mL 12 mg/mL

Glycine 1.78 mg/mL 1.78 mg/mL

PH 5.2 ± 0.3 5.2 ± 0.3 Various studies were performed to evaluate the desire effect of formulated Adalimumab obtained from Example 1-3 of the present invention: Shaking Study: The formulated compositions of Adalimumab were placed in a glass vial and the vial was positioned on plate shaker. The plate shaker was operated at 500 RPM and post 72 hours the samples were withdrawn and using SE-HPLC and CE-HPLC methods.

Freeze Thaw Study: In order to investigate the impact of freeze-thaw stress, each formulation in PFS were frozen at (-20 ± 5) °C and thawed at room temperature (23 ± 3) °C for four times. Post the four freeze-thaw cycles, protein stability was assessed using stability indicating methods like SE-HPLC and CE-HPLC.

Stability data after Heat Stress at 25 °C

The formulations of Adalimumab remain stable after 14 days of storage at 25 °C.

The above studies indicate that there is no significant increase in impurities of Adalimumab, indicating that the formulation remains stable after multiple freeze-thaw cycles, during stress conditions, and shaking studies and that makes the formulation of the present invention stable for long storage.

Claims

1. A stable aqueous pharmaceutical composition comprising Adalimumab and a buffer system wherein the composition is free of surfactant or substantially free of surfactant.

2. The stable aqueous pharmaceutical composition according to claim 1 wherein the buffer system is selected from the group consisting of histidine, succinate, phosphate, citrate, acetate, maleate and tartrate buffers including combinations thereof.

3. The stable aqueous pharmaceutical composition according to claim 1 wherein the buffer system comprises combination of phosphate and citrate buffer.

4. The stable aqueous pharmaceutical composition according to claim 1 further comprises tonicity agent, stabilizer and optionally anti aggregation agent.

5. The stable aqueous pharmaceutical composition according to claim 4 wherein the tonicity agent is selected from sodium chloride or potassium chloride.

6. The stable aqueous pharmaceutical composition according to claim 5 wherein the tonicity agent is sodium chloride.

7. The stable aqueous pharmaceutical composition according to claim 4 wherein the stabilizer is selected from polyols, sugars, cyclodextrins and the combinations thereof.

8. The stable aqueous pharmaceutical composition according to claim 7 wherein the polyol is selected from mannitol, sorbitol, dextran, glycerol, arabitol, propylene glycol, polyethylene glycol and suitable combinations thereof.

9. The stable aqueous pharmaceutical composition according to claim 7 wherein the sugar is selected from glucose, fructose, galactose, mannose, sorbose, ribose, deoxyribose, amino sugars, neuraminic acid, sucrose, trehalose, lactose, maltose and raffinose and suitable combinations thereof.

10. The stable aqueous pharmaceutical composition according to claim 4 wherein the anti-aggregating agent is selected from arginine, glycine, lysine, histidine, glutamic acid, aspartic acid, isoleucine, leucine, alanine, phenylalanine, tyrosine, tryptophane, methionine, serine, proline, cysteine/cystine and suitable combination thereof.

11. The stable aqueous pharmaceutical composition according to claim 10 wherein the anti-aggregating agent is arginine.

12. The stable aqueous pharmaceutical composition according to claim 11 wherein the anti-aggregating agent is arginine monohydrochloride.

13. The stable aqueous pharmaceutical composition according to claim 10 wherein the anti-aggregating agent is glycine.

14. The stable aqueous pharmaceutical composition according to any of the preceding claim comprises pH in the range from pH 4 to pH 8.

15. The stable aqueous pharmaceutical composition according to any of the preceding claim comprises;

a) Adalimumab b) buffer system comprises the combination of phosphate and citrate buffer selected from the group essentially consisting of sodium dihydrogen phosphate dihydrate, disodium hydrogen phosphate dihydrate, trisodium citrate dihydrate, citric acid monohydrate; c) sodium chloride; d) mannitol; e) optionally anti-aggregating agent is arginine monohydrochloride and

having pH 5.2 ± 0.3.

16. The stable aqueous pharmaceutical composition according to claim 15 comprising: a) about 45 mg/ml to about 160 mg/ml of Adalimumab;

b) buffer system comprises about 1 mM to about 50 mM of sodium dihydrogen phosphate dihydrate, about 0.1 mM to about 50 mM of disodium hydrogen phosphate dihydrate, about 0.1 mM to about 50 mM of trisodium citrate dihydrate, about 1 mM to about 100 mM of citric acid monohydrate

c) about 10 mM to about 300 mM of sodium chloride;

d) about 10 mM to about 100 mM of mannitol;

e) about 5 mM to about 100 mM of L- arginine monohydrochloride and

having pH 5.2 ± 0.3.

17. The stable aqueous pharmaceutical composition according to any of the preceding claim comprising:

a) about 45 mg/ml to about 160 mg/ml of Adalimumab;

c) about 10 mM to about 300 mM of sodium chloride;

d) about 10 mM to about 100 mM of mannitol;

e) about 5 mM to about 100 mM of glycine and

having pH 5.2 ± 0.3.

18. The stable aqueous pharmaceutical composition according to claim 16 comprising: a) 50 mg/ml of Adalimumab;

c) 105.4mM of sodium chloride;

d) 65.9mM of mannitol;

e) 23.7 mM of L-arginine monohydrochloride and

having pH 5.2 ± 0.3.

19. The stable aqueous pharmaceutical composition according to claim 17 comprising: a) 50 mg/ml of Adalimumab;

b) buffer system comprises 5.5mM of sodium dihydrogen phosphate dihydrate, 8.6mM of disodium hydrogen phosphate dihydrate, ImM of trisodium citrate dihydrate, 6.2mM of citric acid monohydrate;

c) 105.4 mM of sodium chloride;

d) 65.9 mM of mannitol;

e) 23.7 mM of glycine and

having pH 5.2 ± 0.3.

20. The stable aqueous pharmaceutical composition according at any of preceding claim is administered with injection device selected from syringe, Pen, auto-injector.