DESC:
FIELD OF INVENTION
The present invention relates to a stable pharmaceutical composition comprising monoclonal antibody containing amino acid as a stabilizing excipient and buffer. More particularly, the invention relates to a stable pharmaceutical composition of monoclonal antibody that binds to human epidermal growth factor receptor (EGFR) such as HER 2 receptor, amino acid, buffer and surfactant.

BACKGROUND OF THE INVENTION
Antibodies can have exquisite specificity of target recognition and thus generate highly selective outcomes following their systemic administration. While antibodies can have high specificity, the doses required to treat patients, particularly for a chronic condition, are typically large. Fortunately, advances in production and purification capacities have allowed for the exceptionally large amounts of highly purified monoclonal antibodies to be produced. Additionally, genetic engineering of antibodies has provided a stable of antibody-like proteins that can be easier to prepare. Together, these advances have made antibody-based therapies one of the most commonly pursued pharmaceuticals in biotechnology pipelines. Therapeutic and diagnostic antibodies have become the fastest growing area of bio pharmaceutical applications. The majority of currently approved antibody drugs address previously unmet medical needs in the areas of cancer and inflammation.

In addition to novel antibody drugs and engineered antibody drugs that are currently under development, biosimilar antibody drugs are also being marketed. Optimization of drug composition, therefore, would ensure patient safety and also reduce the cost for pharmaceutical companies. Antibody drugs may be used either in the lyophilized form or as liquid composition; however, liquid composition are being increasingly used.

HER2 is a member of the human epidermal growth factor receptor family. Amplification or over-expression of this oncogene has been shown to play an important role in the development and progression of certain aggressive types of breast cancer. HER2 amplification and resultant HER2 protein overexpression have been linked to important tumor cell proliferation and survival pathways; several drugs have been developed to target the pathway; and, the detection of HER2 has become a routine prognostic and predictive factor in breast cancer.

Pertuzumab is a HER2 antibody useful for breast cancer therapy which binds to the HER2 receptor (antigen). Pertuzumab targets the extracellular dimerization domain (Subdomain II) of HER2 receptor. Inhibition of these signaling pathways can result in cell growth arrest and apoptosis, respectively. In addition, pertuzumab mediates antibody-dependent cell-mediated cytotoxicity (ADCC).

Pertuzumab was discovered and developed by Genentech, a subsidiary of Roche, and was first approved in 2012.

U.S. Patent No. 7,862,817 discloses a humanized anti-ErbB2 antibodies and methods for treating cancer with anti-ErbB2 antibodies (pertuzumab).

U.S. Patent No. 8,372,396 discloses a pharmaceutical formulation comprising an antibody that binds to domain II of HER2 in a histidine-acetate buffer at a pH from about 5.5 to about 6.5, a saccharide, and a surfactant.

Though compositions of HER2 antibody have been disclosed in the art, there still exists a need to develop an alternative stable pharmaceutical composition of monoclonal antibody. The inventors of the present invention have found that a stable pharmaceutical composition of HER2 antibody can be developed using amino acids.

SUMMARY OF THE INVENTION
The invention is directed to a stable pharmaceutical composition of monoclonal antibody susceptible to deamidation and/or aggregation comprising amino acid as a stabilizing excipient and buffer. The composition of the present invention retards degradation of the monoclonal antibody therein.

Monoclonal antibodies are prone to form aggregates, charged species and other degradant species in the composition. The inventors of the present invention found that the addition of amino acids in composition makes the antibody stable by inhibiting aggregation, charged species and other degradant species. It also improves the colloidal stability of the composition.

In one aspect, the invention provides a stable pharmaceutical composition comprising a monoclonal antibody, amino acid and buffer wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In another aspect, the invention relates to a stable pharmaceutical composition comprising
a. monoclonal antibody,
b. amino acid,
c. buffer, and
d. surfactant
wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In another aspect, the invention relates to a stable pharmaceutical composition comprising
a. HER2 antibody,
b. amino acid,
c. buffer, and
d. surfactant
wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In a further aspect, the invention relates to a stable liquid pharmaceutical composition comprising
a. Pertuzumab,
b. amino acid,
c. buffers selected from tris acetate and/or histidine hydrochloride, and
d. polysorbate 20
wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In yet further aspect, invention relates to a stable liquid pharmaceutical composition for intravenous administration comprising
a. Pertuzumab,
b. amino acid selected from glycine and/or arginine,
c. buffers selected from tris acetate and/or histidine hydrochloride, and
d. polysorbate 20,
wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

BRIEF DESCRIPTION OF YHE DRAWINGS
FIG 1 shows % Normalized HMW and LMW in 20 mM Tris-Acetate containing glycine
FIG 2 shows % Normalized HMW and LMW in 20 mM Tris-Acetate containing arginine
FIG 3 shows % Normalized HMW and LMW in 20 mM Histidine HCl containing glycine
FIG 4 shows % Normalized HMW and LMW in 20 mM Histidine HCl containing arginine

DETAILED DESCRIPTION OF THE INVENTION
The present invention relates to a stable pharmaceutical composition comprising monoclonal antibody.

As monoclonal antibodies are prone to form aggregates, charged species and other product degradant that may adversely affect its therapeutic value. Among these product variants and aggregates are known to pose safety risk due to immunogenicity, while other impurities may affect efficacy. Hence these impurities are required to be controlled to ensure the physical and chemical stability.

The inventors of the present invention have found that a stable pharmaceutical composition of HER2 antibody can be developed using amino acids.

The present invention relates to a stable pharmaceutical composition comprising monoclonal antibody, specifically HER2 antibody containing one or more amino acid as a stabilizing excipient and buffer. The invention more particularly relates to a stable pharmaceutical composition of monoclonal antibody specifically HER 2 antibody such as pertuzumab containing one or more amino acid as a stabilizing excipient and buffer wherein the said composition have pH from about 5.0 to about 7.0.

As used herein, the term ‘stable pharmaceutical composition’ refers to a preparation covering a biological active ingredient to be effective and which contains no additional components which are unacceptably toxic to a subject.

As used herein the term ‘monoclonal antibody’ refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies comprising the population are identical and/or bind the same epitope, except for possible variants that may arise during production of the monoclonal antibody, such variants generally being present in minor amounts.

The human epidermal growth factor receptor (HER) family comprises four homologous members: ErbB-1 (epidermal growth factor [EGF] receptor [EGFR] or HER-1), ErbB-2 (HER-2), ErbB-3 (HER-3) and ErbB-4 (HER-4). The activation of these receptors triggers a complex series of signal transduction pathways which affect pivotal tumorigenic processes. The deregulation of HER signaling is seen in several human malignancies. HER-2 is now recognized as a key oncogene in breast cancer pathogenesis. HER receptors achieve activation by forming ligand-bound homo-and/or heterodimeric receptor complexes. The HER complexes signal from the cell surface to the nucleus through numerous downstream pathways which affect the transcription of target genes, regulating critical tumorigenic processes including proliferation, differentiation, apoptosis, angiogenesis, and migration.

As used herein, the term ‘HER 2’ is a member of the human epidermal growth factor receptor family which contains an extracellular ligand binding domain, a transmembrane domain, and an intracellular domain that can interact with a multitude of signaling molecules and exhibit both ligand-dependent and ligand-independent activity.

As used herein, HER2 antibody refers to antibody that binds to antigen binding site of HER2 and includes pertuzumab and/or trastuzumab.

In an embodiment, a stable pharmaceutical composition of the present invention may comprise HER 2 antibody in the concentration ranging from about 10 mg/mL to about 200 mg/mL, preferably about 5 mg/mL to about 100 mg/mL, more preferably about 5 mg/mL to about 50 mg/mL.

As monoclonal antibodies are prone to form aggregates, charged species and other product degradant that may adversely affect its therapeutic value. The downstream processes are developed to control most common protein based impurities includes aggregates such as high molecular weight (HMW) and low molecular weight (LMW) species, charged variants such as acidic variants and basic variants, and host-cell proteins.

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

As used herein, ‘Low Molecular Weight’ species, refers to clipped species of intact antibodies composed of heavy and light chains.

As used herein, ‘High Molecular Weight’ species, refers to higher order oligomeric species like dimers, trimers, tetramers, etc., formed by addition of monomers that can be either covalently or non-covalently linked, which consist of misfolded monomers in which surfaces of the monomer are exposed that typically would not be in the folded monomeric form.

Amino acids used in the present invention decreases aggregates/impurities formation during storage of the composition. Examples of amino acid includes, but not limited to arginine, lysine, aspartic acid, glycine, glutamic acid and/or combinations thereof. The preferred amino acid is arginine and/or glycine. The composition of the present invention may comprise amino acid in the range of about 30 mM to about 200 mM, preferably from about 50 mM to 150 mM, more preferably from about 50 mM to about 130 mM.

As used herein, ‘buffer’ refers to a buffered solution which is an aqueous solution consisting of a mixture of a weak acid and its conjugate base, or vice versa that resists changes in pH by the action of its acid-base conjugate components. Examples of buffers includes, but not limited to Tris, succinate, phosphate, histidine, citrate, maleate, acetate, other organic acid buffers and/or combinations thereof. The preferred buffer herein is Tris or histidine buffer. Examples of histidine buffers includes, but not limited to histidine hydrochloride, histidine acetate, histidine phosphate, histidine sulfate. The preferred histidine buffer is histidine hydrochloride. Examples of Tris buffers includes, but not limited to Tris acetate, Tris maleate, Tris succinate and/or combinations thereof. The preferred Tris buffer is Tris acetate. The composition of the present invention may comprise buffer in the range of about 5 mM to about 50 mM, preferably from about 5 mM to 30 mM, more preferably from about 5 mM to about 25 mM.

The term ‘surfactant’ as used herein is intended to mean a substance that functions to reduce the surface tension of a liquid dissolved therein, to suppress or control agglomeration, particle formation and/or surface adsorption in liquid composition. The surfactant can be anionic, cationic and/or nonionic. Examples of surfactants herein includes, but not limited to polysorbate (for example, polysorbate 20 and, polysorbate 80); poloxamer (e.g. poloxamer 188); Triton; sodium dodecyl sulfate; sodium laurel sulfate; sodium octyl glycoside; lauryl-, myristyl-, linoleyl-, or stearyl-sulfobetaine; lauryl-, myristyl-, linoleyl- or stearyl-sarcosine; linoleyl-, myristyl-, or cetyl-betaine; lauroamidopropyl-, cocamidopropyl-, linoleamidopropyl-, myristamidopropyl-, palmidopropyl-, or isostearamidopropyl-betaine (e.g. lauroamidopropyl); myristamidopropyl-, palmidopropyl-, or isostearamidopropyl-dimethylamine; sodium methyl cocoyl-, or disodium methyl oleyl-taurate; the polyethyl glycol, polypropyl glycol, copolymers of ethylene, propylene glycol and/or combinations thereof. The preferred surfactants are polysorbate 20, polysorbate 80, poloxamer, triton, sodium dodecyl sulfate, sodium laurel sulfate, propylene glycol and/or combinations thereof. More preferably the surfactant is polysorbate 20. The composition of the present invention may comprise surfactant in the range of about 0.001 % (w/v) to about 1.0 % (w/v), preferably from about 0.01 % (w/v) to about 0.1 % (w/v), more preferably from about 0.01 % (w/v) to about 0.05 % (w/v).

The pH of the stable pharmaceutical composition of the present invention is in the range from about 5.0 to about 7.0, preferably from about 5.5 to about 6.5, for example from about 5.8 to about 6.2, and most preferably has a pH from about 6.0.

In an embodiment the stable pharmaceutical composition of the present invention is a liquid composition.

In another aspect, the invention relates to a stable pharmaceutical composition comprising monoclonal antibody, amino acid, buffer, and surfactant; wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In another aspect, the invention relates to a stable pharmaceutical composition comprising HER2 antibody, amino acid, buffer, and surfactant; wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In another aspect, the invention relates to a stable pharmaceutical composition comprising pertuzumab and/or trastuzumab, amino acid, buffer, and surfactant; wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In another aspect, the invention relates to a stable pharmaceutical composition comprising pertuzumab, amino acid, buffer, and surfactant; wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In a further aspect, the invention relates to a stable liquid pharmaceutical composition comprising pertuzumab, amino acid, buffers selected from tris acetate and/or histidine hydrochloride, and polysorbate 20; wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In yet further aspect, invention relates to a stable liquid pharmaceutical composition comprising pertuzumab, amino acid selected from glycine and/or arginine, buffers selected from tris acetate and/or histidine hydrochloride, and polysorbate 20; wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

In yet further aspect, invention relates to a stable liquid pharmaceutical composition for intravenous administration comprising pertuzumab, amino acid selected from glycine and/or arginine, buffers selected from tris acetate and/or histidine hydrochloride, and polysorbate 20; wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

The stable pharmaceutical composition of the present invention is administered to a human patient in accord with known methods, such as intravenous administration, e.g., as a bolus or by continuous infusion over a period of time, by intramuscular, intraperitoneal, intracerobrospinal, Subcutaneous, intra-articular, intrasynovial routes. The preferred routes are intravenous, intramuscular or subcutaneous and the most preferred route is with intravenous administration.

Stability of present pharmaceutical composition, refers to the retention of structure and/or function of a polypeptide within a composition. A polypeptide in a composition of the invention exhibit attributes such as resistance to change or deterioration that affect stability or function and therefore maintain consistent functional characteristics over time. Accordingly, composition of the invention will exhibit, for example, reliability and safety with respect to activity per volume or activity units.

The stability of present pharmaceutical composition can be tested by evaluating physical stability, chemical stability, and/or biological activity of the antibody in the composition around the time of composition as well as following storage at the noted temperatures. Physical and/or chemical 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 etc.

The following examples illustrate the invention and they do not any way limit the scope of the invention. A person skilled in the art would easily modify the process for manufacturing the said pharmaceutical composition or could modify the composition with similar materials and finally a person skilled in the art could modify the method of administering the said composition of this invention.

EXAMPLES

Example 1: Pertuzumab composition (FA):

Table 1

Sr. No. Ingredients Concentration Amount/L
1 Pertuzumab 30 mg/mL 30 gm
2 Tris 20 mM 2.42 gm
3 Glacial acetic acid 21 mM 1.202 mL
4 Polysorbate 20 0.02% (w/v) 0.2 mL

Pertuzumab was formulated in pH 6.0 buffer at a concentration of 30 mg/mL into 20 mM Tris-Acetate and 0.02% (w/v) polysorbate 20. Each composition was filtered and transferred into USP Type 1 glass vials and stoppered. The stoppered vial containing pertuzumab solution was hermetically sealed with aluminum flip-off seals.

Example 2: Pertuzumab composition (FB):

Table 2
Sr. No. Ingredients Concentration Amount/L
1 Pertuzumab 30 mg/mL 30 gm
2 Histidine Hydrochloride 20 mM 4.19 gm
3 Polysorbate 20 0.02% (w/v) 0.2 mL
4 Sodium Hydroxide (10% w/v) q.s to adjust pH 6 q.s to adjust pH 6

Pertuzumab was formulated in pH 6.0 buffer at a concentration of 30 mg/mL into 20 mM Histidine-HCl and 0.02% (w/v) polysorbate 20. Each composition was filtered and transferred into USP Type 1 glass vials and stoppered. The stoppered vial containing pertuzumab solution was hermetically sealed with aluminum flip-off seals.

Example 3: Stable Pertuzumab composition (F1):

Table 3
Sr. No. Ingredients Concentration Amount/L
1 Pertuzumab 30 mg/mL 30 gm
2 Tris 20 mM 2.42 gm
3 Glacial acetic acid 21 mM 1.202 mL
4 Glycine 100 mM 7.51 gm
5 Polysorbate 20 0.02% (w/v) 0.2 mL

Pertuzumab was formulated in pH 6.0 buffer at a concentration of 30 mg/mL into 20 mM Tris-Acetate, 100 mM glycine and 0.02% (w/v) polysorbate 20. Each composition was filtered and transferred into USP Type 1 glass vials and stoppered. The stoppered vial containing pertuzumab solution was hermetically sealed with aluminum flip-off seals.

Example 4: Stable Pertuzumab composition (F2):

Table 4
Sr. No. Ingredients Concentration Amount/L
1 Pertuzumab 30 mg/mL 30 gm
2 Tris 20 mM 2.42 gm
3 Glacial acetic acid 21 mM 1.202 mL
4 Arginine 100 mM 17.42 gm
5 Polysorbate 20 0.02% (w/v) 0.2 mL

Pertuzumab was formulated in pH 6.0 buffer at a concentration of 30 mg/mL into 20 mM Tris-Acetate, 100 mM arginine and 0.02% (w/v) polysorbate 20. Each composition was filtered and transferred into USP Type 1 glass vials and stoppered. The stoppered vial containing pertuzumab solution was hermetically sealed with aluminum flip-off seals.

Example 5: Stable Pertuzumab composition (F3):

Table 5
Sr. No. Ingredients Concentration Amount/L
1 Pertuzumab 30 mg/mL 30 gm
2 Histidine Hydrochloride 20 mM 4.19 gm
3 Glycine 100 mM 7.51 gm
4 Polysorbate 20 0.02%(w/v) 0.2 mL
5 Sodium Hydroxide (10% w/v) q.s to adjust pH 6 q.s to adjust pH 6

Pertuzumab was formulated in pH 6.0 buffer at a concentration of 30 mg/mL into 20 mM Histidine-HCl, 100 mM glycine and 0.02% (w/v) polysorbate 20. Each composition was filtered and transferred into USP Type 1 glass vials and stoppered. The stoppered vial containing pertuzumab solution was hermetically sealed with aluminum flip-off seals.

Example 6: Stable Pertuzumab composition (F4):

Table 6
Sr. No. Ingredients Concentration Amount/L
1 Pertuzumab 30 mg/mL 30 gm
2 Histidine Hydrochloride 20 mM 4.19 gm
3 Arginine 100 mM 17.42
4 Polysorbate 20 0.02% (w/v) 0.2 mL
5 Sodium Hydroxide (10% w/v) q.s to adjust pH 6 q.s to adjust pH 6

Pertuzumab was formulated in pH 6.0 buffer at a concentration of 30 mg/mL into 20 mM Histidine-HCl, 100 mM arginine and 0.02% (w/v) polysorbate 20. Each composition was filtered and transferred into USP Type 1 glass vials and stoppered. The stoppered vial containing pertuzumab solution was hermetically sealed with aluminum flip-off seals.

The compositions of the above examples were further subjected to stress stability studies at 40°C. Further samples were analyzed for aggregates to determine HMW and LMW species by size exclusion chromatography. The results of composition with amino acid (F1-F4) were compared with the composition without amino acids (FA and FB). It was found that level of HMW and LMW species was less in composition containing amino acids compared to compositions without amino acids.
Analytical Method:
The compositions were further subjected to stress stability studies at 40°C. The samples were analyzed for aggregates and LMW species by Size Exclusion Chromatography. A mobile phase containing 100 mM disodium hydrogen phosphate and 250 mM sodium perchlorate in water at pH 6.5 was used at a flow rate of 0.5 mL/min for 45 minutes to separate HMW and LMW species of pertuzumab at a UV detection of 214 nm.
,CLAIMS:
1. A stable pharmaceutical composition comprising a monoclonal antibody, amino acid, buffer and surfactant.

2. The stable pharmaceutical composition according to claim 1, wherein the monoclonal antibody is HER 2 antibody.

3. The stable pharmaceutical composition according to claim 2, wherein the HER 2 antibody is Pertuzumab or Trastuzumab.

4. The stable pharmaceutical composition according to claim 1, wherein the amino acid is selected from arginine, glycine, cysteine, proline, lysine, phenylalanine, leucine and combinations thereof.

5. The stable pharmaceutical composition according to claim 1, wherein the buffer is selected from tris, succinate, succinate, phosphate, histidine, citrate, maleate, acetate and combinations thereof.

6. The stable pharmaceutical composition according to claim 1, wherein the surfactant selected from polysorbate 20, polysorbate 80, poloxamer, triton, sodium dodecyl sulfate, sodium laurel sulfate, propylene glycol and combinations thereof.

7. The stable pharmaceutical composition according to claim 1, wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

8. A stable pharmaceutical composition comprising;
a. HER2 antibody,
b. amino acid selected from glycine and arginine,
c. buffer selected from tris acetate and histidine hydrochloride, and
d. surfactant,
wherein the pharmaceutical composition has a pH from about 5.0 to 7.0.

9. The stable pharmaceutical composition of claim 8, wherein the composition is for intravenous administration.

10. The stable pharmaceutical composition of claim 8, wherein the composition comprises amino acid from about 50 mM to 130 mM, buffer from about 5 mM to 25 mM and surfactant from about 0.01 % (w/v) to 0.05 % (w/v).