Stable antibody compositions

Abstract

The invention relates to an aqueous pharmaceutical composition as well as a preparation method and application thereof. The aqueous pharmaceutical composition comprises risankizumab or an antigen binding fragment of risankizumab, an antigen binding fragment of risankizumab, an antigen binding fragment of risankizumab and an antigen binding fragment of risankizumab, and a stabilizer, wherein the aqueous pharmaceutical composition does not include a polyol.

Description

[Technical field]

[0001] The present invention relates to a stable pharmaceutical composition comprising risankizumab or an antigen-binding fragment thereof, a method for preparing the composition, and medical use of the composition for treating various diseases.

[0002] [Prior Art]

[0003] Risankizumab is a highly effective and specific inhibitor of interleukin-23 (IL-23), and is a humanized immunoglobulin G1 (IgG1) monoclonal antibody against the p19 subunit of IL-23. Risankizumab can inhibit the effects of IL-23 in inducing and maintaining T helper (Th) 17 cells, innate lymphocytes, γδT cells and natural killer (NK) cells due to its binding to IL-23p19, which cause tissue inflammation, destruction and abnormal tissue recovery. Risankizumab is known to be effective in treating autoimmune diseases, especially inflammatory bowel disease (such as Crohn's disease and ulcerative colitis), multiple sclerosis, rheumatoid arthritis, ankylosing spondylitis, psoriatic arthritis or psoriasis (see WO2012 / 061448).

[0004] In order to effectively administer risankizumab, a pharmaceutical formulation with excellent stability is required. WO2021 / 048743 discloses a liquid pharmaceutical formulation comprising risankizumab and a polyol as a stabilizer and surfactant. However, it is known that polyols represented by sugars or sugar alcohols contain monosaccharides as impurities, and it is also known that polyols can be decomposed into monosaccharides depending on pH or temperature stress conditions (see The effect of sucrose hydrolysis on the stability of protein therapeutics during accelerated formulation studies. Journal of pharmaceutical sciences, December 2009; 98(12): 4501-10). However, it is known that monosaccharides can react with the amino acid groups of proteins and cause glycosylation, which has the potential risk of causing protein denaturation (aggregates, etc.) or affecting protein efficacy (see Glycation of polyclonal IgGs: Effect of sugar excipients during stability studies, European Journal of Pharmaceutics and Biopharmaceutics, Vol. 102, May 2016, pp. 185-190; Quantitative analysis of glycation and its impact on antigen binding, MAbs. 2018, Vol. 10, No. 3, pp. 406-415).

[0005]

Prior Technical Documents

[0006] [Patent Literature]

[0007] Patent Document 1: WO2012 / 061448

[0008] Patent document 2: WO2021 / 048743

[0009]

Non-patent literature

[0010] Non-patent literature 1: The effect of sucrose hydrolysis on the stability of protein therapeutics during accelerated formulation studies. Journal of pharmaceutical sciences, December 2009; 98(12): 4501-10

[0011] Non-patent literature 2: Quantitative analysis of glycation and its impact on antigen binding, MAbs. 2018, Vol. 10, No. 3, 406-415 [Summary of the invention]

[0012]

Technical issues

[0013] Therefore, there is a need to develop formulations of risankizumab that can further improve the stability of risankizumab while including stabilizers that can replace polyols.

[0014] One embodiment of the present invention provides a stable pharmaceutical composition comprising risankizumab or an antigen-binding fragment thereof, wherein the stable pharmaceutical composition is suitable for use as a medicament for treating an individual.

[0015] Another embodiment of the present invention provides a method of treating an autoimmune disease in an individual, comprising administering a pharmaceutical composition to the individual.

[0016] Another embodiment of the present invention provides a method for preparing a pharmaceutical composition.

[0017] [Solution to the problem]

[0018] One embodiment of the present invention provides an aqueous pharmaceutical composition comprising:

[0019] (a) risankizumab or an antigen-binding fragment thereof; and

[0020] (b) stabilizers,

[0021] wherein the aqueous pharmaceutical composition does not include a polyol.

[0022] Another embodiment of the present invention provides an aqueous pharmaceutical composition comprising:

[0023] (a) risankizumab or an antigen-binding fragment thereof; and

[0024] (b) an amino acid or a pharmaceutically acceptable salt thereof,

[0025] The pH of the aqueous pharmaceutical composition (c) is 5.0-7.0.

[0026] Another embodiment of the present invention provides a method for treating an autoimmune disease, comprising administering the aforementioned pharmaceutical composition to an individual.

[0027] [Advantageous effects of the present invention]

[0028] It was confirmed that the aqueous pharmaceutical composition including risankizumab or its antigen-binding fragment according to one embodiment has excellent stability under various conditions, including thermal stability, light stability, freezing and / or freeze-thaw stability, and stirring stability, by including an amino acid or its salt or metal salt (such as sodium chloride) as a stabilizer, while not including a polyol conventionally used as a stabilizer. In addition, since it has excellent stability even under conditions with or without a surfactant and / or a buffer, it has the advantage of being able to avoid side effects that may occur due to the inclusion of a surfactant and / or a buffer.

Brief Description of the Figures

[0030] Figure 1 The amino acid sequence of the light chain of risakizumab is shown (SEQ ID NO: 1).

[0031] Figure 2 The amino acid sequence of the heavy chain of risakizumab is shown (SEQ ID NO: 2).

[0032] Figure 3 Graph showing changes in HMW% (ΔHMW%) observed in thermal stability testing of risankizumab formulations including only stabilizers without buffers and surfactants at week 1, week 2, and week 4 relative to the initial state.

[0033] Figure 4 A graph showing stability modeling of the results of thermal stability testing of risankizumab formulations including proline as a stabilizer (SE-HPLC and WCX results) by designing 5 factors (protein concentration, pH, histidine buffer concentration, proline stabilizer concentration, and surfactant concentration) based on design of experiments (DoE) via DoE statistical analysis.

[0034] Figure 5 A graph showing stability modeling of the results of the stirring stability test (SE-HPLC and WCX results) of risankizumab formulations including proline as a stabilizer by designing 5 factors (protein concentration, pH, histidine buffer concentration, proline stabilizer concentration, and surfactant concentration) based on design of experiments (DoE) via DoE statistical analysis.

[0035] Figure 6A graph showing stability modeling of the results of photostability testing of risankizumab formulations including proline as a stabilizer (SE-HPLC, WCX and HIC results) by designing 5 factors (protein concentration, pH, histidine buffer concentration, proline stabilizer concentration and surfactant concentration) based on design of experiments (DoE) via DoE statistical analysis.

[0036] Figure 7 A graph showing stability modeling of the results of freeze-thaw stability testing of risankizumab formulations including proline as a stabilizer (SE-HPLC and WCX results) by designing 5 factors (protein concentration, pH, histidine buffer concentration, proline stabilizer concentration, and surfactant concentration) based on design of experiments (DoE) via DoE statistical analysis.

[0037] [Implementation Method]

[0038] Unless otherwise defined, all technical terms as used in this specification have the same meaning as those generally familiar with the relevant technology. In addition, suitable methods or samples are described in this specification, but similar or equivalent methods or samples are also within the scope of this specification. In addition, although not explicitly stated, the numerical values ​​described in this specification are considered to include the meaning of "about". The contents of all public cases mentioned in this invention are incorporated herein by reference in their entirety.

[0039] According to one embodiment of the present invention, an aqueous pharmaceutical composition comprises:

[0040] (a) risankizumab or an antigen-binding fragment thereof; and

[0041] (b) stabilizers,

[0042] wherein the aqueous pharmaceutical composition does not include a polyol.

[0043] The stabilizer may include an amino acid or a pharmaceutically acceptable salt thereof, or a metal salt.

[0044] The metal salt may be NaCl, KCl, NaF, KBr, NaBr, Na2SO4, NaSCN, CaCl2, MgCl2 or K2SO4. The metal salt may be, for example, NaCl or Na2SO4. The metal salt may be present in the pharmaceutical composition at a concentration of 0.5wt% to 1wt%. In one embodiment, the pharmaceutical composition may include 0.8wt% sodium chloride. The concentration of the metal salt may be freely adjusted within the range of maintaining the stability of the risankizumab or its antigen-binding fragment in the pharmaceutical composition, and may vary individually depending on the specific type of the metal salt.

[0045] According to another embodiment of the present invention, an aqueous pharmaceutical composition comprises:

[0046] (a) risankizumab or an antigen-binding fragment thereof; and

[0047] (b) an amino acid or a pharmaceutically acceptable salt thereof,

[0048] Wherein the pH of the aqueous pharmaceutical composition in (c) is 5.0 to 7.0. The pharmaceutical composition according to this embodiment may not include polyols.

[0049] Hereinafter, pharmaceutical compositions according to two embodiments will be described in more detail.

[0050] Amino acids can act as stabilizers and can include lysine, arginine, glycine, proline, histidine, alanine, valine, leucine, isoleucine, proline, phenylalanine, tyrosine, tryptophan, serine, threonine, cysteine, methionine, asparagine, glutamine, aspartic acid, glutamic acid, or pharmaceutically acceptable salts of the foregoing or mixtures of the foregoing, but are not limited thereto. In one embodiment, the amino acid can be lysine, arginine, glycine, proline, histidine, pharmaceutically acceptable salts of the foregoing or any combination of the foregoing. In one embodiment, the amino acid can be proline.

[0051] The concentration of the amino acid as a stabilizer may be 0.1 mM to 300.0 mM, 0.5 mM to 300.0 mM, 1.0 mM to 300.0 mM, 5.0 mM to 300.0 mM, 10.0 mM to 300.0 mM, 25.0 mM to 300.0 mM, 30.0 mM to 300.0 mM, 50.0 mM to 300.0 mM, 80.0 mM to 300.0 mM, 100.0 mM to 300.0 mM, 120.0 mM to 300.0 mM, 0.1 mM to 250.0 mM, 0.5 mM to 250.0 mM, 1.0 mM to 250.0 mM, 5.0 mM to 250.0 mM, 10.0 mM to 250.0 mM, 2 5.0mM~250.0mM, 30.0mM~250.0mM, 50.0mM~250.0mM, 80.0mM~250.0mM, 100 .0mM~250.0mM, 120.0mM~250.0mM, 0.1mM~200.0mM, 0.5mM~200.0mM, 1.0mM ~200.0mM, 5.0mM~200.0mM, 10.0mM~200.0mM, 25.0mM~200.0mM, 30.0mM~20 0.0mM, 50.0mM~200.0mM, 80.0mM~200.0mM, 100.0mM~200.0mM, 120.0mM~200 .0mM, 0.1mM~160.0mM, 0.5mM~160.0mM, 1.0mM~160.0mM, 5.0mM~160.0mM, 1 0.0mM~160.0mM, 25.0mM~160.0mM, 30.0mM~160.0mM, 50.0mM~160.0mM, 80. 0mM~160.0mM, 100.0mM~160.0mM, 120.0mM~160.0mM, 130.0mM~150.0mM, 0. 1mM~100.0mM, 0.5mM~100.0mM, 1.0mM~100.0mM, 5.0mM~100.0mM, 10.0mM~10 0.0mM, 25.0mM~100.0mM, 30.0mM~100.0mM, 50.0mM~100.0mM, 80.0mM~100. 0mM, 0.1mM~50.0mM, 0.5mM~50.0mM, 1.0mM~50.0mM, 5.0mM~50.0mM, 10.0mM~ 50.0mM, 25.0mM~50.0mM, 30.0mM~50.0mM, 0.1mM~40.0mM, 0.5mM~40.0mM, 1 .0mM~40.0mM, 5.0mM~40.0mM, 10.0mM~40.0mM, 25.0mM~40.0mM, 30.0mM~40.Alternatively, the concentration of the amino acid may be 1 wt % to 3 wt %. In one embodiment, the amino acid may be 100 mM to 200 mM histidine, in particular 150 mM histidine, a pharmaceutically acceptable salt thereof, or a mixture thereof. In one embodiment, the amino acid may be lysine, arginine, glycine, or proline at a concentration of 1 wt% to 3 wt%. The concentration of the amino acid may be adjusted within a range that provides stability of risakizumab or its antigen-binding fragment without affecting the desired pH of the pharmaceutical composition, and may vary depending on the specific amino acid. In one embodiment, the amino acid may be 2.5 wt% proline. .

[0052] The polyol not included in the pharmaceutical composition may be, for example, sorbitol, sucrose, trehalose, mannose, maltose, mannitol, or a mixture of the foregoing. The pharmaceutical composition may have excellent stability by including a metal salt or an amino acid as a stabilizer while excluding a polyol (see Test Example 3, formulations 5-9, 14-18, 20 and 22; and Test Example 4, formulations 5-8). This is an unexpected effect when considering that pharmaceutical formulations containing risankizumab in this technology typically include polyols to have stability. In addition, polyols may include monosaccharides as impurities, and monosaccharides are known to have the potential risk of causing risankizumab to denature (aggregates, etc.) or affect the efficacy of proteins (see non-patent documents 1 and 2). In this regard, the pharmaceutical composition can avoid the potential risk of monosaccharides destabilizing the protein active ingredient due to the absence of polyols.

[0053] In addition, the pharmaceutical composition may or may not include a surfactant. Excellent stability can be achieved with or without a surfactant (see Test Example 3, Formulations 5-9 and 20 versus Formulations 14-18 and 22; Test Example 4, Formulations 5-8).

[0054] In addition, the pharmaceutical composition may or may not include a buffer. Excellent stability can be achieved with or without a buffer (see Test Example 3, Formulations 5-9 and 20 versus Formulations 14-18 and 22; Test Example 4, Formulations 5-8).

[0055] In one embodiment, the pharmaceutical composition may include neither a polyol nor a surfactant.

[0056] As used in the present specification, the expression "the pharmaceutical composition does not include ingredient A" may mean that the pharmaceutical formulation does not include or substantially does not include ingredient A. The expression "substantially does not include ingredient A" may be interpreted to cover the case where ingredient A is completely absent, the case where ingredient A (if present) is present in a trace amount so as not to substantially affect the characteristics of the pharmaceutical composition, or the case where ingredient A is present in an undetectable amount.

[0057] The surfactant may be, for example, a polysorbate, a poloxamer, a sorbitan ester of another fatty acid, or a mixture of the foregoing. The polysorbate may be, for example, polysorbate 20, polysorbate 80, or a mixture of the foregoing. In one embodiment, the surfactant may be polysorbate 20. The concentration of the surfactant in the pharmaceutical composition may be 0.001 wt% to 2 wt% of the pharmaceutical composition, and may vary depending on the specific type of the surfactant. In one embodiment, the surfactant may be 0.02 wt% polysorbate 20.

[0058] The pH of the pharmaceutical composition may be 5.0 to 7.0. Specifically, for example, the pH of the pharmaceutical composition may be pH 5 to 6.8, pH 5 to 6.5, pH 5 to 6.3, pH 5.2 to 6.3, pH 5.0 to 6.0, pH 5.2 to 6.0, pH 5.0 to 5.8, pH 5.2 to 5.8, pH 5.0 to 5.6, pH 5.2 to 5.6, pH 5.1, pH 5.2, pH 5.3, pH 5.4, pH 5.5, pH 5.7 or pH 6.0. In one embodiment, the pH of the pharmaceutical composition may be 5.7.

[0059] The pharmaceutical composition may include a buffer. The buffer may be, for example, an acetate, succinate, citrate, glutamate, glycine, lactate, maleate, phosphate, tartrate or histidine buffer or any combination of the foregoing, but is not limited thereto. The concentration of the buffer included in the pharmaceutical composition may be an amount suitable for maintaining the selected pH during the storage period of the pharmaceutical composition. For example, the buffer may be present in the pharmaceutical composition at a concentration of 10 mM to 60 mM, and the concentration of the buffer may vary depending on the specific type of buffer. In one embodiment, the buffer may be 16 mM histidine. "Resakizumab," which is the main ingredient included in the pharmaceutical composition, is currently marketed under the trade name is sold as an antibody for the treatment of autoimmune diseases, in particular psoriasis or psoriatic arthritis. Risankizumab is known to be used for the treatment of autoimmune diseases, inflammatory diseases, respiratory diseases, metabolic disorders or cancer, more specifically psoriasis, psoriatic arthritis, inflammatory bowel disease, multiple sclerosis, rheumatoid arthritis, Crohn's disease, ulcerative colitis, ankylosing spondylitis, asthma, chronic obstructive pulmonary disease (COPD) or the like. Risankizumab may also include the commercially available “Biosimilar” or “biobetter” products that are similar to the active risakizumab antibody present in

[0060] Risankizumab may be an antibody including a light chain having the amino acid sequence of SEQ ID NO:1 and a heavy chain having the amino acid sequence of SEQ ID NO:2.

[0061] As used herein, the term "antigen-binding fragment" refers to a fragment capable of binding to the target antigen IL-23p19 of an antibody (ie, risankizumab), including, for example, a Fab fragment, a F(ab')2 fragment, an Fc fragment or a scFv fragment, but is not limited thereto.

[0062] Rasanizumab can be produced by general methods known in the art. For example, WO2012 / 061448 discloses methods that those skilled in the art can use to prepare Rasanizumab. These methods are incorporated herein by reference.

[0063] The concentration of risankizumab or its antigen-binding fragment in the pharmaceutical composition can be, for example, 9 mg / mL to 170 mg / mL, 9 mg / mL to 45 mg / mL, 10 mg / mL to 40 mg / mL, 15 mg / mL to 35 mg / mL, 20 mg / mL to 30 mg / mL, 130 mg / mL to 160 mg / mL, or about 150 mg / mL.

[0064] The pharmaceutical composition may be an aqueous liquid formulation.

[0065] The pharmaceutical composition can be administered by parenteral delivery. Parenteral administration can include, for example, not only subcutaneous, intramuscular, intradermal, and intramedullary injections, but also intrathecal, direct intraventricular, intravenous, intraperitoneal, and intravitreal injections. The drug can be administered in various conventional ways, such as intraperitoneal, parenteral, intraarterial, or intravenous injections.

[0066] The pharmaceutical composition can be used for subcutaneous injection, intramuscular injection or intravenous injection. The pharmaceutical composition may also include an aqueous carrier suitable for injection. When administered to humans, the aqueous carrier may be a safe, non-toxic, pharmaceutically acceptable carrier, and examples thereof may include water, physiological saline solution, Ringer's solution, dextrose, or a mixture of the foregoing. In one embodiment, the aqueous carrier may be water.

[0067] The osmotic pressure of the pharmaceutical composition can be within a suitable range for subcutaneous or intravenous injection. The osmotic pressure can be, for example, 200mOsm / kg to 400mOsm / kg, 200mOsm / kg to 350mOsm / kg, 250mOsm / kg to 300mOsm / kg, 250mOsm / kg to 290mOsm / kg, 270mOsm / kg to 328mOsm / kg, 250mOsm / kg to 269mOsm / kg, or 328mOsm / kg to 350mOsm / kg. The osmotic pressure can be appropriately adjusted to minimize the pain that may be caused during administration.

[0068] In one embodiment, in the pharmaceutical composition, the concentration of risankizumab or its antigen-binding fragment may be 9 mg / mL to 170 mg / mL, the stabilizer may be sodium chloride, lysine, arginine, glycine, proline, histidine, or a pharmaceutically acceptable salt of the foregoing, or a mixture of the foregoing, or sodium chloride, the surfactant may be polysorbate 20, the pH may be 5.0 to 7.0, and a polyol may not be included.

[0069] In one embodiment, in the pharmaceutical composition, the concentration of risankizumab or its antigen-binding fragment may be 9 mg / mL to 170 mg / mL, the stabilizer may be sodium chloride, lysine, arginine, glycine, proline, histidine or a pharmaceutically acceptable salt thereof or a mixture of the foregoing, or sodium chloride, the pH may be 5.0 to 7.0, and polyols and surfactants may not be included.

[0070] In one embodiment, the pharmaceutical composition may include 150 mg / mL risankizumab, 16 mM histidine, 2.5 wt % proline, and 0.02 wt % polysorbate 20, has a pH of 5.7, and optionally excludes polyols and / or surfactants.

[0071] In one embodiment, the pharmaceutical composition may include 150 mg / mL risankizumab, 10 mM histidine, 2.5 wt % proline, and 0.02 wt % polysorbate 20, has a pH of 5.7, and optionally excludes polyols and / or surfactants.

[0072] In the pharmaceutical compositions of the present invention, risankizumab or its antigen-binding fragment can be stabilized. The term "stabilized" means that risankizumab or its antigen-binding fragment substantially maintains its physical stability, chemical stability and / or biological activity before and after administration, during additional manufacturing processes, preservation or storage. Physical stability, chemical stability and / or biological activity can be assessed by commonly known methods. In one embodiment, stability can be assessed by testing thermal stability, photostability, freeze-thaw stability and stirring stability, as shown in the Examples below.

[0073] Rasanizumab can be used to treat any disease known in the art to be effective. For example, WO2012 / 061448 discloses a range of indications that can be treated by administering Rasanizumab. These methods are incorporated herein by reference.

[0074] Therefore, another embodiment of the present invention provides a method for treating an autoimmune disease, such as psoriasis or psoriatic arthritis, comprising administering the aforementioned pharmaceutical composition to an individual. In addition to the aforementioned diseases, inflammatory diseases, respiratory diseases, metabolic disorders or cancers may also be treated, more specifically, inflammatory bowel disease (Crohn's disease, ulcerative colitis), multiple sclerosis, rheumatoid arthritis, ankylosing spondylitis, asthma, chronic obstructive pulmonary disease (COPD) or the like. The method of treatment may include administering a therapeutically effective amount of a pharmaceutical formulation to an individual. The individual may include a human.

[0075] Another embodiment of the present invention provides the aforementioned pharmaceutical composition for use in treating autoimmune diseases, cancer, psoriasis, psoriatic arthritis, inflammatory bowel disease, multiple sclerosis, rheumatoid arthritis, ankylosing spondylitis, asthma, chronic obstructive pulmonary disease (COPD) or the like.

[0076] Another embodiment of the present invention includes a method for preparing the aforementioned pharmaceutical composition, the method comprising: preparing a mixed solution by adding a stabilizer to an aqueous carrier; and adding risakizumab or an antigen-binding fragment thereof to the mixed solution; or

[0077] The method comprises: preparing a solution by adding risankizumab or an antigen-binding fragment thereof to an aqueous carrier; and adding a stabilizer to the solution.

[0078] The process is optionally carried out without addition of polyols.

[0079] The aqueous carrier can be an aqueous solvent (eg, water or physiological saline solution).

[0080] In this method, a buffer (or buffer solution) and / or a surfactant may be optionally added in the step of adding a stabilizer.

[0081] The details of the method of preparing the pharmaceutical composition can be directly applied from the description of the pharmaceutical composition according to the embodiments of the present invention.

[0082] Hereinafter, the technical features and effects of the present invention will be described in more detail through examples and experimental examples. However, the following examples and experimental examples are provided only for the purpose of illustration to help understand the present invention, and the scope and range of the present invention are not limited thereto.

[0083]

Abbreviation explanation

[0084] abbreviation explain SE-HPLC Size Exclusion-HPLC WCX Weak cation exchange chromatography HIC HIC HMW% High molecular weight LMW% Low molecular weight monomer% Monomer content Acidity % Acidic variant percentage Alkalinity% Basic variant percentage main% Main percentage Hydrophobicity% Percentage of hydrophobic variants Hydrophilicity% Percentage of hydrophilic variants Rpm RPM

[0085]

Analysis method

[0086] In the analytical methods of the following test examples, SE-HPLC and WCX were performed on the samples to determine thermal stability, freeze-thaw stability and stirring stability. By SE-HPLC analysis, HMW%, LMW% and monomer% were measured. By WCX analysis, acidity%, basicity% and main% were measured.

[0087] In addition, HIC as well as SE-HPLC and WCX were performed to determine photostability. From HIC, the hydrophobicity %, hydrophilicity % and primary % were measured.

[0088] In SE-HPLC analysis, a high performance liquid chromatography (HPLC) system is used, such as the Alliance HPLC system from Waters (Milford, MA, USA), which includes a size exclusion chromatography (SEC) column. Proteins separated from the SEC column can be detected by UV absorption at 280 nm, and the determination of relative amounts can be performed by calculating the area under the curve (AUC) of each separated peak. Peaks of various species attributes are determined based on the separation time corresponding to the molecular size of the species. To determine the relative HMW%, relative LMW% and relative monomer% of antibodies (especially monomeric antibodies) in the formulation, HMW and LMW species need to be separated from each other when present in the formulation. In particular, the relative content or relative amount is expressed as a percentage value, and the sum of monomer%, HMW% and LMW% is 100%.

[0089] In weak cation exchange chromatography (WCX), a high performance liquid chromatography (HPLC) system is used, such as the Alliance HPLC system from Waters (Milford, MA, USA), which includes a WCX column. The protein separated from the WCX column can be detected by UV absorption at 280nm, and the determination of relative amount can be performed by calculating the area under the curve (AUC) of each separation peak or each group of separation peaks. According to the separation conditions corresponding to the surface charge of the antibody substance type, the peaks of various substance type attributes are determined. In the case of stability analysis, it can be measured after the preparation of the formulation (T0) and then after the indicated storage time under the specified storage conditions. Acidity % or basicity % includes all peaks before or after the main % original antibody variant peak. These peaks represent antibody variants with higher acidity and / or higher basicity compared to the main % original antibody variant, and / or antibody variants with more negative or positive charges on their surface under chromatographic conditions. Specifically, relative content or relative amount is expressed as a percentage value, and the sum of the main %, acidity % and basicity % is 100%.

[0090] In HIC analysis, a high performance liquid chromatography (HPLC) system is used, such as the Alliance HPLC system from Waters (Milford, MA, USA), which includes a HIC column. Proteins separated from the HIC column can be detected by UV absorption at 280 nm, and the determination of relative amounts can be performed by calculating the area under the curve (AUC) of each separated peak or each group of separated peaks. Peaks of various species attributes are determined based on the separation time corresponding to the hydrophobicity of the species. To determine the relative hydrophobicity %, relative hydrophilicity % and relative predominant % of the antibody, hydrophobic species and hydrophilic species are separated from each other when present in the formulation. Specifically, the relative content or relative amount is expressed as a percentage value, and the sum of the hydrophobicity %, hydrophilicity % and predominant % is 100%.

[0091] [Test Example 1: Stability Analysis of Formulations Depending on pH and Buffer Solution]

[0092] Aqueous risakizumab liquid formulations of the compositions shown in Table 1 below were prepared and then analyzed for their thermal stability, photostability, freeze-thaw stability, and stirring stability.

[0093] [Table 1: Formulation composition depending on pH and buffer solution]

[0094]

[0095]

[0096] Preparation of samples: After storage under various stress conditions, the formulation samples were collected to determine their stability. Dialysis was performed using each prepared buffer solution, and the formulation samples were prepared at the desired protein concentration. The prepared formulation was sterile filtered, filled with 1 mL into a syringe and exposed to various stress conditions. The stress conditions are as follows. For thermal stability, the samples were stored and collected at a temperature of 40±2°C in the 1st, 2nd, 4th and 5th weeks. For light stability, the samples were stored and collected at an illumination of not less than 1.2 million lux hours and not less than 200 watt hours (Wh) / square meter (m 2 ) were stored and collected under the integrated near-ultraviolet conditions. For freeze-thaw stability, samples were collected after repeating freeze-thaw cycles 5 times at room temperature and -60°C or lower. For stirring stability, samples were collected after stirring at 0 rpm and 300 rpm.

[0097] Results: The results of measuring thermal stability, photostability, freeze-thaw stability and stirring stability are shown in Tables 2 to 28 below. [Table 2: SE-HPLC HMW (%) measurement results (thermal stability)]

[0098]

[0099]

[0100] [Table 3: SE-HPLC LMW (%) measurement results (thermal stability)]

[0101]

[0102] [Table 4: SE-HPLC monomer (%) measurement results (thermal stability)]

[0103]

[0104]

[0105] The results determined by SE-HPLC analysis at a storage temperature of 40°C are shown in Tables 2-4.

[0106] During the 5 week storage period, HMW% increased by 2% to 5%. The maximum increase was measured to be +4.10%, +4.51%, +4.33% and +4.99% for Formulation 3, Formulation 6, Formulation 9 and Formulation 17, respectively, and the increase for the remaining formulations, except for the formulation with the maximum increase, was measured to be an average of +2.17%.

[0107] During the 5-week storage period, the LMW% increased by 2% to 14%. The maximum increase was measured to be +8.50%, +14.25%, +9.50% and +4.15% for Formulation 4, Formulation 6, Formulation 9 and Formulation 17, respectively, and the increase of the remaining formulations, except for the formulation with the largest increase, was measured to be an average of +2.35%.

[0108] During the 5 week storage period, the monomer % decreased by 3% to 19%. The maximum decreases were measured to be -12.59%, -18.77%, -13.85% and -9.14% for Formulation 3, Formulation 6, Formulation 9 and Formulation 17, respectively, and the decreases for the remaining formulations, except for the formulation with the maximum decrease, were measured to be an average of -4.52%.

[0109] [Table 5: WCX acidity (%) measurement results (thermal stability)]

[0110]

[0111] [Table 6: WCX alkalinity (%) measurement results (thermal stability)]

[0112]

[0113]

[0114] [Table 7: Main (%) measurement results of WCX (thermal stability)]

[0115]

[0116] The results determined by WCX analysis at a storage temperature of 40°C are shown in Tables 5-7.

[0117] During the 5 week storage period, the acidity % increased by 0% to 36%. The minimum increase was measured to be +0.14%, +5.75% and +4.76% for Formulation 3, Formulation 6 and Formulation 9 respectively, and the increase for the remaining formulations except for the formulation with the minimum increase was measured to be an average of +24.78%.

[0118] During the 5 week storage period, the % alkalinity increased by 1% to 56%. The maximum increase was measured to be +48.50%, +55.72%, +54.58% and +15.45% for Formulation 3, Formulation 6, Formulation 9 and Formulation 17, respectively, and the increase for the remaining formulations, except for the formulation with the maximum increase, was measured to be an average of +8.26%.

[0119] During the 5 week storage period, the major % reductions ranged from 24% to 62%. The maximum reductions were measured to be -48.65%, -61.47%, -59.35% and -51.71% for Formulation 3, Formulation 6, Formulation 9 and Formulation 17, respectively, and the reductions for the remaining formulations, except for the formulation with the maximum reduction, were measured to be an average of -36.96%.

[0120] [Table 8: SE-HPLC HMW (%) measurement results (photostability)]

[0121]

[0122] [Table 9: SE-HPLC LMW (%) measurement results (photostability)]

[0123]

[0124]

[0125] [Table 10: SE-HPLC monomer (%) measurement results (photostability)]

[0126]

[0127] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of SE-HPLC analysis under integrated near-ultraviolet conditions are shown in Tables 8 to 10.

[0128] After exposure, the HMW% increased by 1% to 14%. The maximum increase was measured to be +11.38% and +13.67% for Formulations 16 and 17 respectively, and the increase for the remaining formulations except for the formulation with the largest increase was measured to be an average of +4.08%.

[0129] After light exposure, LMW% did not show a significant increase and was measured to be +0.29% on average.

[0130] After exposure, the monomer % decreased by 2% to 14%. The maximum decrease was measured to be -11.60% and -13.82% for Formulations 16 and 17 respectively, and the decrease for the remaining formulations except for the formulation with the maximum decrease was measured to be an average of -4.38%.

[0131] [Table 11: WCX acidity (%) measurement results (photostability)]

[0132]

[0133]

[0134] [Table 12: WCX alkalinity (%) measurement results (photostability)]

[0135]

[0136] [Table 13: WCX main (%) measurement results (photostability)]

[0137]

[0138]

[0139] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of WCX analysis under integrated near-ultraviolet conditions are shown in Tables 11 to 13.

[0140] After exposure, the % Acidity showed no significant increase and was measured to be +4.96% on average.

[0141] After exposure, the % Basicity showed no significant increase and was measured to be +16.81% on average.

[0142] After light exposure, the primary % did not show a significant decrease and was measured to be an average of -21.77%.

[0143] [Table 14: HIC hydrophobicity (%) measurement results (photostability)]

[0144]

[0145]

[0146] [Table 15: HIC hydrophilicity (%) measurement results (photostability)]

[0147]

[0148] [Table 16: HIC main (%) measurement results (photostability)]

[0149]

[0150]

[0151] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of HIC analysis under integrated near-ultraviolet conditions are shown in Tables 14 to 16.

[0152] After exposure, the % hydrophobicity increased by 8% to 27%. The minimum increase was measured to be +12.69%, +9.63%, +12.79%, +11.36% and +8.25% for Formulations 13, 14, 15, 16 and 17, respectively, and the increase for the remaining formulations, except for the formulation with the minimum increase, was measured to be an average of +22.21%.

[0153] After exposure, the % Hydrophilicity showed no significant increase and was measured to be +2.96% on average.

[0154] After exposure, the major % reduction was 15% to 28%. The minimum reduction was measured to be -14.73%, -12.52%, -17.18%, -18.48% and -14.83% for Formulations 13, 14, 15, 16 and 17 respectively, and the reduction for the remaining formulations except for the formulation with the minimum reduction was measured to be an average of -24.43%.

[0155] [Table 17: SE-HPLC HMW (%) measurement results (freeze-thaw stability)]

[0156]

[0157] [Table 18: SE-HPLC LMW (%) measurement results (freeze-thaw stability)]

[0158]

[0159]

[0160] [Table 19: SE-HPLC monomer (%) measurement results (freeze-thaw stability)]

[0161]

[0162] The results determined by SE-HPLC analysis after repeating freeze-thaw cycles 5 times under the conditions of room temperature and -60°C or lower are shown in Tables 17 to 19.

[0163] After 5 freeze-thaw cycles, HMW% did not show a significant increase and was measured to be an average of -0.23%. After 5 freeze-thaw cycles, LMW% did not show a significant increase and was measured to be an average of -0.36%. After 5 freeze-thaw cycles, monomer% did not show a significant decrease and was measured to be an average of +0.59%. [Table 20: WCX acidity (%) measurement results (freeze-thaw stability)]

[0164]

[0165] [Table 21: WCX alkalinity (%) measurement results (freeze-thaw stability)]

[0166]

[0167]

[0168] [Table 22: Main (%) measurement results of WCX (freeze-thaw stability)]

[0169]

[0170] The results determined by WCX analysis after repeating freeze-thaw cycles 5 times at room temperature and -60°C or lower are shown in Tables 20 to 22.

[0171] After 5 freeze-thaw cycles, the % Acidity did not show a significant increase and was measured to be +0.71% on average.

[0172] After 5 freeze-thaw cycles, the % Alkalinity did not show a significant increase and was measured to be an average of -0.72%.

[0173] After 5 freeze-thaw cycles, the prime % did not show a significant decrease and was measured to be +0.01% on average.

[0174] [Table 23: SE-HPLC HMW (%) measurement results (stirring stability)]

[0175]

[0176]

[0177] [Table 24: SE-HPLC LMW (%) measurement results (stirring stability)]

[0178]

[0179] [Table 25: SE-HPLC monomer (%) measurement results (stirring stability)]

[0180]

[0181]

[0182] The results of SE-HPLC analysis after stirring at 300 rpm are shown in Tables 23 to 25.

[0183] After stirring, HMW% did not show a significant increase and was measured to be +0.09% on average.

[0184] After stirring, the LMW% did not show a significant increase and was measured to be -0.79% on average.

[0185] After stirring, the monomer % did not show a significant decrease and was measured to be +0.70% on average.

[0186] [Table 26: WCX acidity (%) measurement results (stirring stability)]

[0187]

[0188] [Table 27: WCX alkalinity (%) measurement results (stirring stability)]

[0189]

[0190]

[0191] [Table 28: WCX main (%) measurement results (stirring stability)]

[0192]

[0193] The results determined by WCX analysis after stirring at 300 rpm are shown in Tables 26 to 28. After stirring, the acidity % did not show a significant increase and was measured to be +0.04% on average.

[0194] After stirring, the % Alkalinity did not show a significant increase and was measured to be -0.32% on average.

[0195] After stirring, the Primary % did not show a significant decrease and was measured to be +0.30% on average.

[0196] Discussion of Results: According to the results in Tables 2 to 28, formulations including various types of buffers at a pH of 5.0 to 7.0 were determined to be stable under various stress conditions (heat, light, freeze-thaw and agitation stress).

[0197] [Test Example 2-1: Stability Analysis of Formulations Dependent on Surfactants

[0198] Aqueous risakizumab liquid formulations having the compositions shown in Table 29 below were prepared and subsequently analyzed for thermal stability, photostability, freeze-thaw stability, and stirring stability.

[0199] [Table 29: Formulation composition depending on surfactant]

[0200]

[0201] Preparation of samples: After storage under various stress conditions, the formulation samples were collected to determine their stability. Dialysis was performed using each prepared buffer solution, and the formulation samples were prepared at the desired protein concentration. The prepared formulation was sterile filtered, filled with 1 mL into a syringe and exposed to various stress conditions. The stress conditions are as follows. For thermal stability, the samples were stored and collected at a temperature of 40±2°C at the 1st, 2nd, 4th and 6th weeks. For light stability, the samples were stored and collected at an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 For freeze-thaw stability, samples were collected after five freeze-thaw cycles at room temperature and -60°C or lower. For stirring stability, samples were collected after stirring at 0 rpm and 300 rpm.

[0202] Results: The results of measuring thermal stability, photostability, stirring stability and freeze-thaw stability are shown in Tables 30 to 56 below. [Table 30: SE-HPLC HMW (%) measurement results (thermal stability)]

[0203]

[0204] [Table 31: SE-HPLC LMW (%) measurement results (thermal stability)]

[0205]

[0206] [Table 32: SE-HPLC monomer (%) measurement results (thermal stability)]

[0207]

[0208]

[0209] The results determined by SE-HPLC analysis at a storage temperature of 40°C are shown in Tables 30-32.

[0210] During the 6-week storage period, HMW% did not show a significant increase and was measured to be an average of +2.26%.

[0211] During the 6-week storage period, LMW% did not show a significant increase and was measured to be an average of +2.04%.

[0212] During the 6-week storage period, the % monomer did not show a significant decrease and was measured to be an average of -4.30%.

[0213] [Table 33: WCX acidity (%) measurement results (thermal stability)]

[0214]

[0215] [Table 34: WCX alkalinity (%) measurement results (thermal stability)]

[0216]

[0217] [Table 35: WCX main (%) measurement results (thermal stability)]

[0218]

[0219] The results determined by WCX analysis at a storage temperature of 40°C are shown in Tables 33-35. During the 6-week storage period, the acidic % did not show a significant increase and was measured to be an average of +25.59%. During the 6-week storage period, the basic % did not show a significant increase and was measured to be an average of +1.76%. During the 6-week storage period, the main % did not show a significant decrease and was measured to be an average of -27.35%.

[0220] [Table 36: SE-HPLC HMW (%) measurement results (photostability)]

[0221]

[0222]

[0223] [Table 37: SE-HPLC LMW (%) measurement results (photostability)]

[0224]

[0225] [Table 38: SE-HPLC monomer (%) measurement results (photostability)]

[0226]

[0227] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of SE-HPLC analysis under integrated near-ultraviolet conditions are shown in Tables 36 to 38.

[0228] After light exposure, HMW% did not show a significant increase and was measured to be +6.65% on average.

[0229] After light exposure, LMW% did not show a significant increase and was measured to be +0.00% on average.

[0230] After stirring, the monomer % did not show a significant decrease and was measured to be an average of -6.65%.

[0231] [Table 39: WCX acidity (%) measurement results (photostability)]

[0232]

[0233] [Table 40: WCX alkalinity (%) measurement results (photostability)]

[0234]

[0235] [Table 41: WCX main (%) measurement results (photostability)]

[0236]

[0237]

[0238] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of WCX analysis under integrated near-ultraviolet conditions are shown in Tables 39 to 41.

[0239] After exposure, the % Acidity showed no significant increase and was measured to be +6.73% on average.

[0240] After exposure, the % Alkalinity showed no significant increase and was measured to be +14.19% on average.

[0241] After light exposure, the primary % did not show a significant decrease and was measured to be an average of -20.92%.

[0242] [Table 42: HIC hydrophobicity (%) measurement results (photostability)]

[0243]

[0244] [Table 43: HIC hydrophilicity (%) measurement results (photostability)]

[0245]

[0246] [Table 44: HIC main (%) measurement results (photostability)]

[0247]

[0248] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of HIC analysis under integrated near-ultraviolet conditions are shown in Tables 42 to 44.

[0249] After exposure, the % hydrophobicity did not show a significant increase and was measured to be +15.46% on average.

[0250] After exposure, the % Hydrophilicity showed no significant increase and was measured to be +4.84% on average.

[0251] After light exposure, the primary % did not show a significant decrease and was measured to be an average of -20.31%.

[0252] [Table 45: SE-HPLC HMW (%) measurement results (freeze-thaw stability)]

[0253]

[0254] [Table 46: SE-HPLC LMW (%) measurement results (freeze-thaw stability)]

[0255]

[0256] [Table 47: SE-HPLC monomer (%) measurement results (freeze-thaw stability)]

[0257]

[0258] The results determined by SE-HPLC analysis after repeating freeze-thaw cycles 5 times under the conditions of room temperature and -60°C or lower are shown in Tables 45 to 47.

[0259] After 5 freeze-thaw cycles, HMW% did not show a significant increase and was measured to be +0.03% on average.

[0260] After 5 freeze-thaw cycles, LMW% did not show a significant increase and was measured to be +0.03% on average.

[0261] After 5 freeze-thaw cycles, the % monomer did not show a significant decrease and was measured to be -0.06% on average.

[0262] [Table 48: WCX acidity (%) measurement results (freeze-thaw stability)]

[0263]

[0264] [Table 49: WCX alkalinity (%) measurement results (freeze-thaw stability)]

[0265]

[0266] [Table 50: Main (%) measurement results of WCX (freeze-thaw stability)]

[0267]

[0268]

[0269] The results determined by WCX analysis after repeating freeze-thaw cycles 5 times under room temperature and -60°C or lower temperature conditions are shown in Tables 48 to 50.

[0270] After 5 freeze-thaw cycles, the % Acidity did not show a significant increase and was measured to be +0.73% on average.

[0271] After 5 freeze-thaw cycles, the % Alkalinity did not show a significant increase and was measured to be an average of -0.09%.

[0272] After 5 freeze-thaw cycles, the prime % did not show a significant decrease and was measured to be an average of -0.64%.

[0273] [Table 51: SE-HPLC HMW (%) measurement results (stirring stability)]

[0274]

[0275] [Table 52: SE-HPLC LMW (%) measurement results (stirring stability)]

[0276]

[0277] [Table 53: SE-HPLC monomer (%) measurement results (stirring stability)]

[0278]

[0279] The results of SE-HPLC analysis after stirring at 300 rpm are shown in Tables 51 to 53.

[0280] After stirring, HMW% did not show a significant increase and was measured to be -0.02% on average.

[0281] After stirring, the LMW% did not show a significant increase and was measured to be -0.14% on average.

[0282] After stirring, the monomer % did not show a significant decrease and was measured to be +0.17% on average.

[0283] [Table 54: WCX acidity (%) measurement results (stirring stability)]

[0284]

[0285] [Table 55: WCX alkalinity (%) measurement results (stirring stability)]

[0286]

[0287] [Table 56: WCX main (%) measurement results (stirring stability)]

[0288]

[0289] The results of the WCX analysis after stirring at 300 rpm are shown in Tables 54 to 56.

[0290] After stirring, the % acidity did not show a significant increase and was measured to be -0.04% on average.

[0291] After stirring, the % Alkalinity did not show a significant increase and was measured to be +0.02% on average.

[0292] After stirring, the Primary % did not show a significant decrease and was measured to be +0.02% on average.

[0293] Discussion of Results: According to the results in Tables 30 to 56, formulations with or without surfactants showed stability under various stress conditions (heat, light, freeze-thaw and stirring stress).

[0294] [Test Example 2-2: Stability Analysis of Formulations Dependent on Surfactants

[0295] Aqueous risakizumab liquid formulations having the compositions shown in Table 57 below were prepared and then analyzed for their thermal stability.

[0296] [Table 57: Formulation composition depending on surfactant]

[0297]

[0298] Preparation of samples: After storage under various stress conditions, the formulation samples were collected to determine their stability. Dialysis was performed using each prepared buffer solution, and the formulation samples were prepared at the desired protein concentration. The prepared formulations were sterile filtered and 1 mL of the formulations were filled into syringes to confirm their thermal stability. For thermal stability, samples were stored and collected at a temperature of 40 ± 2 ° C at 1, 2, and 4 weeks.

[0299] Results: The results of measuring thermal stability are shown in Tables 58 to 63 below.

[0300] [Table 58: SE-HPLC HMW (%) measurement results (thermal stability)]

[0301]

[0302]

[0303] [Table 59: SE-HPLC LMW (%) measurement results (thermal stability)]

[0304]

[0305] [Table 60: SE-HPLC monomer (%) measurement results (thermal stability)]

[0306]

[0307] The results determined by SE-HPLC analysis at a storage temperature of 40°C are shown in Tables 58 to 60. During the 4-week storage period, HMW% did not show a significant increase and was measured to be an average of +1.14%. During the 4-week storage period, LMW% did not show a significant increase and was measured to be an average of +1.86%. During the 4-week storage period, Monomer% did not show a significant decrease and was measured to be an average of -3.00%.

[0308] [Table 61: WCX acidity (%) measurement results (thermal stability)]

[0309]

[0310] [Table 62: WCX alkalinity (%) measurement results (thermal stability)]

[0311]

[0312] [Table 63: WCX main (%) measurement results (thermal stability)]

[0313]

[0314]

[0315] The results determined by WCX analysis at a storage temperature of 40°C are shown in Tables 61-63. During the 4-week storage period, the acidic % did not show a significant increase and was measured to be an average of +13.78%. During the 4-week storage period, the basic % did not show a significant increase and was measured to be an average of +8.02%. During the 4-week storage period, the main % did not show a significant decrease and was measured to be an average of -21.79%.

[0316] Discussion of Results: According to the results in Tables 58 to 63, formulations with or without surfactants showed stability under hot temperature conditions.

[0317] [Test Example 3: Stability Analysis of Formulations Dependent on Stabilizers]

[0318] Aqueous risakizumab liquid formulations having the compositions shown in Table 64 below were prepared and subsequently analyzed for thermal stability, photostability, freeze-thaw stability, and stirring stability.

[0319] [Table 64: Formulation composition depending on stabilizer]

[0320]

[0321]

[0322] Preparation of samples: After storage under various stress conditions, the formulation samples were collected to determine their stability. Dialysis was performed using each prepared buffer solution, and the formulation samples were prepared at the desired protein concentration. The prepared formulation was sterile filtered, filled with 1 mL into a syringe and exposed to various stress conditions. The stress conditions are as follows. For thermal stability, the samples were stored and collected at a temperature of 40±2°C at 2 weeks, 4 weeks and 6 weeks. For light stability, the samples were stored and collected at an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 For freeze-thaw stability, samples were collected after five freeze-thaw cycles at room temperature and -60°C or lower. For stirring stability, samples were collected after stirring at 0 rpm and 300 rpm.

[0323] Results: The results of measuring stability are shown in Tables 65-91 below.

[0324] [Table 65: SE-HPLC HMW (%) measurement results (thermal stability)]

[0325]

[0326]

[0327] [Table 66: SE-HPLC LMW (%) measurement results (thermal stability)]

[0328]

[0329] [Table 67: SE-HPLC monomer (%) measurement results (thermal stability)]

[0330]

[0331]

[0332] The results determined by SE-HPLC analysis at a storage temperature of 40°C are shown in Tables 65 to 67. During the 6-week storage period, HMW% increased by 0.8% to 2.0%, with an average increase of 1.75% measured for formulations including surfactants and polyols or salts (Formulations 1 to 5), an average increase of 1.27% measured for formulations including amino acids and surfactants (Formulations 6 to 9 and 20), an average increase of 1.78% measured for formulations including polyols or salts (Formulations 10 to 14), and an average increase of 1.26% measured for formulations including amino acids (Formulations 15 to 18 and 22).

[0333] During the 6-week storage period, LMW% did not show a significant increase and was measured to be an average of +2.8%.

[0334] During the 6 week storage period, the % monomer decreased by 3.74% to 4.97%, with an average decrease of 4.41% measured for formulations including surfactants and polyols or salts (Formulations 1-5), an average decrease of 4.19% measured for formulations including amino acids and surfactants (Formulations 6-9 and 20), an average decrease of 4.40% measured for formulations including polyols or salts (Formulations 10-14), and an average decrease of 4.16% measured for formulations including amino acids (Formulations 15-18 and 22).

[0335] [Table 68: WCX acidity (%) measurement results (thermal stability)]

[0336]

[0337]

[0338] [Table 69: WCX alkalinity (%) measurement results (thermal stability)]

[0339]

[0340] [Table 70: WCX main (%) measurement results (thermal stability)]

[0341]

[0342] The results determined by WCX analysis at a storage temperature of 40°C are shown in Tables 68-70. During the 6-week storage period, the % Acidity did not show a significant increase and was measured to be an average of +17.73%.

[0343] During the 6 week storage period, the % Alkalinity did not show a significant increase and was measured to be an average of +6.41%.

[0344] During the 6 week storage period, the Primary % did not show a significant decrease, and an average decrease of 24.14% was measured.

[0345] According to the results in Tables 65 to 70, it was confirmed that the formulation including amino acids and optional surfactants had better stability under stress conditions compared to the formulation including polyols and optional surfactants.

[0346] [Table 71: SE-HPLC HMW (%) measurement results (photostability)]

[0347]

[0348]

[0349] [Table 72: SE-HPLC LMW (%) measurement results (photostability)]

[0350]

[0351]

[0352] [Table 73: SE-HPLC monomer (%) measurement results (photostability)]

[0353]

[0354] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of SE-HPLC analysis under integrated near-ultraviolet conditions are shown in Tables 71 to 73.

[0355] After exposure, the HMW% increased by 0.5% to 5.2%, with an average increase of 4.27% measured for formulations including surfactants and polyols or salts (Formulations 1-5), an average increase of 3.24% measured for formulations including amino acids and surfactants (Formulations 6-9 and 20), an average increase of 4.05% measured for formulations including polyols or salts (Formulations 10-14), and an average increase of 2.80% measured for formulations including amino acids (Formulations 15-18 and 22).

[0356] After light exposure, LMW% did not show a significant increase and was measured to be +0.07% on average.

[0357] After exposure, the monomer % decreased by 0.6% to 5.2%, with an average decrease of 4.39% measured for formulations including surfactants and polyols or salts (Formulations 1-5), an average decrease of 3.37% measured for formulations including amino acids and surfactants (Formulations 6-9 and 20), an average decrease of 4.13% measured for formulations including polyols or salts (Formulations 10-14), and an average increase of 2.82% measured for formulations including amino acids (Formulations 15-18 and 22).

[0358] [Table 74: WCX acidity (%) measurement results (photostability)]

[0359]

[0360] [Table 75: WCX alkalinity (%) measurement results (photostability)]

[0361]

[0362]

[0363] [Table 76: WCX main (%) measurement results (photostability)]

[0364]

[0365] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of WCX analysis under integrated near-ultraviolet conditions are shown in Tables 74 to 76.

[0366] After exposure, the acidity % increased by 2.4% to 9.9%, with an average increase of 7.48% measured for formulations comprising surfactants and polyols or salts (Formulations 1 to 5), an average increase of 6.87% measured for formulations comprising amino acids and surfactants (Formulations 6 to 9 and Formulation 20), an average increase of 6.85% measured for formulations comprising polyols or salts (Formulations 10 to 14), and an average increase of 5.41% measured for formulations comprising amino acids (Formulations 15 to 18 and Formulation 22).

[0367] After exposure, the % alkalinity increased by 3.3% to 16.0%, with an average increase of 14.70% measured for formulations comprising surfactants and polyols or salts (Formulations 1 to 5), an average increase of 12.83% measured for formulations comprising amino acids and surfactants (Formulations 6 to 9 and Formulation 20), an average increase of 13.96% measured for formulations comprising polyols or salts (Formulations 10 to 14), and an average increase of 10.82% measured for formulations comprising amino acids (Formulations 15 to 18 and Formulation 22).

[0368] After exposure, the major % decrease ranged from 5.7% to 25.0%, with an average decrease of 22.18% measured for formulations comprising surfactants and polyols or salts (Formulations 1-5), an average decrease of 19.71% measured for formulations comprising amino acids and surfactants (Formulations 6-9 and 20), an average decrease of 20.81% measured for formulations comprising polyols or salts (Formulations 10-14), and an average increase of 16.23% measured for formulations comprising amino acids (Formulations 15-18 and 22).

[0369] [Table 77: HIC hydrophobicity (%) measurement results (photostability)]

[0370]

[0371]

[0372] [Table 78: HIC hydrophilicity (%) measurement results (photostability)]

[0373]

[0374] [Table 79: HIC main (%) measurement results (photostability)]

[0375]

[0376]

[0377] At an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 The results of HIC analysis under integrated near-ultraviolet conditions are shown in Tables 77 to 79.

[0378] After exposure, the % hydrophobicity increased by 5% to 17%. The minimum increase was measured to be +9.68%, +9.33%, +5.89% and +4.74% for Formulations 15, 18, 20 and 22 respectively, and the increase for the remaining formulations except for the formulation with the minimum increase was measured to be an average of +12.70%.

[0379] After exposure, the % hydrophilicity increased by 0.1% to 3.0%, with an average increase of 2.28% measured for formulations comprising surfactants and polyols or salts (Formulations 1-5), an average increase of 1.65% measured for formulations comprising amino acids and surfactants (Formulations 6-9 and 20), an average increase of 2.23% measured for formulations comprising polyols or salts (Formulations 10-14), and an average increase of 1.42% measured for formulations comprising amino acids (Formulations 15-18 and 22).

[0380] After exposure, the major % reduction was 5% to 19%. The minimum reductions were measured to be -11.09%, -11.34%, -6.08% and -4.85% for Formulations 15, 18, 20 and 22 respectively, and the reductions for the remaining formulations, except for the formulation with the minimum reduction, were measured to be an average of -14.76%.

[0381] According to the results in Tables 71 to 79, it was confirmed that the formulation including the amino acid and the optional surfactant had better photostability under stress conditions compared with the formulation including the polyol and the optional surfactant.

[0382] [Table 80: SE-HPLC HMW (%) measurement results (freeze-thaw stability)]

[0383]

[0384]

[0385] [Table 81: SE-HPLC LMW (%) measurement results (freeze-thaw stability)]

[0386]

[0387] [Table 82: SE-HPLC monomer (%) measurement results (freeze-thaw stability)]

[0388]

[0389] The results determined by SE-HPLC analysis after repeating freeze-thaw cycles 5 times under the temperature conditions of room temperature and -60°C or lower are shown in Tables 80 to 82.

[0390] After 5 freeze-thaw cycles, HMW% did not show a significant increase and was measured to be +0.03% on average.

[0391] After 5 freeze-thaw cycles, LMW% did not show a significant increase and was measured to be +0.32% on average.

[0392] After 5 freeze-thaw cycles, the % monomer did not show a significant decrease and was measured to be an average of -0.35%.

[0393] [Table 83: WCX acidity (%) measurement results (freeze-thaw stability)]

[0394]

[0395]

[0396] [Table 84: WCX alkalinity (%) measurement results (freeze-thaw stability)]

[0397]

[0398]

[0399] [Table 85: Main (%) measurement results of WCX (freeze-thaw stability)]

[0400]

[0401] The results determined by WCX analysis after repeating freeze-thaw cycles 5 times under room temperature and -60°C or lower temperature conditions are shown in Tables 83 to 85.

[0402] After 5 freeze-thaw cycles, the % Acidity did not show a significant increase and was measured to be an average of -0.96%.

[0403] After 5 freeze-thaw cycles, the % Alkalinity did not show a significant increase and was measured to be +0.98% on average.

[0404] After 5 freeze-thaw cycles, the prime % did not show a significant decrease and was measured to be an average of -0.01%.

[0405] According to the results in Tables 80 to 85, all formulations were confirmed to be stable under freeze-thaw stress conditions.

[0406] [Table 86: SE-HPLC HMW (%) measurement results (stirring stability)]

[0407]

[0408]

[0409] [Table 87: SE-HPLC LMW (%) measurement results (stirring stability)]

[0410]

[0411]

[0412] [Table 88: SE-HPLC monomer (%) measurement results (stirring stability)]

[0413]

[0414] The results determined by SE-HPLC analysis after stirring at 300 rpm are shown in Tables 86 to 88. After stirring, HMW% did not show a significant increase, and was measured to be +0.05% on average. After stirring, LMW% did not show a significant increase, and was measured to be -0.06% on average. After stirring, monomer% did not show a significant decrease, and was measured to be 0.00% on average.

[0415] [Table 89: WCX acidity (%) measurement results (stirring stability)]

[0416]

[0417]

[0418] [Table 90: WCX alkalinity (%) measurement results (stirring stability)]

[0419]

[0420]

[0421] [Table 91: WCX main (%) measurement results (stirring stability)]

[0422]

[0423] The results determined by WCX analysis after stirring at 300 rpm are shown in Tables 89 to 91. After stirring, the acid % did not show a significant increase and was measured to be an average of -0.22%. After stirring, the basic % did not show a significant increase and was measured to be an average of +0.13%. After stirring, the main % did not show a significant increase and was measured to be an average of +0.08%.

[0424] According to the results in Tables 86 to 91, all formulations were confirmed to be stable under stirring stress conditions.

[0425] Discussion of Results: Formulations including amino acid stabilizers exhibited stability under various stress conditions (heat, light, and agitation stress) with or without surfactants. Meanwhile, formulations including polyol stabilizers tended to have lower stability compared to formulations including amino acid stabilizers.

[0426] [Test Example 4: Stability Analysis of a Formulation Including Only a Stabilizer]

[0427] Aqueous risakizumab liquid formulations having the compositions shown in Table 92 below were prepared and then analyzed for their thermal stability.

[0428] [Table 92: Composition of formulations including only stabilizers]

[0429]

[0430] Preparation of samples: After storage under thermal temperature stress conditions, samples of formulations including only stabilizers but not buffer solutions and surfactants were collected to determine their stability. Dialysis was performed using each prepared buffer solution, and formulation samples were prepared at the desired protein concentration. The prepared formulations were sterile filtered, filled with 0.3 mL into tubes and exposed to temperature stress conditions. For thermal stability, samples were stored and collected at a temperature of 40±2°C at weeks 1, 2, and 4.

[0431] Results: The results of measuring thermal stability are shown in Tables 93 to 95 below.

[0432] [Table 93: SE-HPLC HMW (%) measurement results (thermal stability)]

[0433]

[0434] [Table 94: SE-HPLC LMW (%) measurement results (thermal stability)]

[0435]

[0436]

[0437] [Table 95: SE-HPLC monomer (%) measurement results (thermal stability)]

[0438]

[0439] The results determined by SE-HPLC analysis at a storage temperature of 40°C are shown in Tables 93-95.

[0440] During the 4-week storage period, HMW% increased by 0.9% to 1.7%, with an average increase of 1.67% measured for formulations including only polyols (Formulations 1 to 4) and an average increase of 1.10% measured for formulations including only amino acids (Formulations 5 to 8). The changes in HMW% (ΔHMW%) at Weeks 1, 2, and 4 relative to the initial period are shown in Figure 3 middle.

[0441] During the 4-week storage period, LMW% did not show a significant increase and was measured to be +1.39% on average.

[0442] During the 4 week storage period, as the HMW% increased, the Monomer% decreased by 2.3% to 3.1%, with an average decrease of 2.95% measured for formulations including only polyols (Formulations 1-4) and an average decrease of 2.59% measured for formulations including only amino acids (Formulations 5-8).

[0443] According to the results in Tables 92 to 95, it was confirmed that the formulation including amino acids had better thermal temperature stability than the formulation including polyols.

[0444] Discussion of Results: According to the results in Tables 93 to 95, it was confirmed that the formulation including only the amino acid stabilizer was more stable under heat stress conditions compared to the formulation including only the polyol stabilizer.

[0445] [Test Example 5: Concentration-dependent stability analysis of formulations including proline stabilizers]

[0446] Aqueous risakizumab liquid formulations having the compositions shown in Table 96 below were prepared and subsequently analyzed for thermal stability, photostability, freeze-thaw stability, and stirring stability.

[0447] [Table 96: Composition of formulations with different stabilizer concentrations]

[0448]

[0449]

[0450] Preparation of samples: In order to determine the appropriate concentration range for stability, formulation samples were prepared by designing a total of 5 factors (protein concentration, pH, concentration of histidine buffer, concentration of proline stabilizer, and concentration of surfactant) based on experimental design (DoE), and then collected after storage. Dialysis was performed using each prepared buffer solution, and formulation samples were prepared at the desired protein concentration. The prepared formulation was sterile filtered, filled with 1 mL into a syringe and exposed to various stress conditions. The stress conditions are as follows. For thermal stability, samples were stored and collected at a temperature of 40±2°C at weeks 4 and 6. For light stability, the samples were stored and collected at an illumination of not less than 1.2 million lux hours and not less than 200Wh / m 2 For freeze-thaw stability, samples were collected after five freeze-thaw cycles at room temperature and -60°C or lower. For stirring stability, samples were collected after stirring at 0 rpm and 400 rpm.

[0451] Analytical method: A prediction model was established by considering the main effects, interaction effects and secondary effects of a total of five factors (protein concentration, pH, histidine concentration, proline concentration and surfactant concentration), and then the thermal stability, photostability, freeze-thaw stability and stirring stability at different concentrations were determined.

[0452] Results: The results of measuring thermal stability, photostability, freeze-thaw stability and stirring stability are shown in Tables 97 to 123 below. [Table 97: SE-HPLC HMW (%) measurement results (thermal stability)]

[0453]

[0454] [Table 98: SE-HPLC LMW (%) measurement results (thermal stability)]

[0455]

[0456]

[0457] [Table 99: SE-HP LC monomer (%) measurement results (thermal stability)]

[0458]

[0459]

[0460] [Table 100: WCX acidity (%) measurement results (thermal stability)]

[0461]

[0462]

[0463] [Table 101: WCX alkalinity (%) measurement results (thermal stability)]

[0464]

[0465] [Table 102: WCX main (%) measurement results (thermal stability)]

[0466]

[0467]

[0468] [Table 103: SE-HPLC HMW (%) measurement results (photostability)]

[0469]

[0470]

[0471] [Table 104: SE-HP LC LMW (%) measurement results (photostability)]

[0472]

[0473] [Table 105: SE-HPLC monomer (%) measurement results (photostability)]

[0474]

[0475]

[0476] [Table 106: WCX acidity (%) measurement results (photostability)]

[0477]

[0478]

[0479] [Table 107: WCX alkalinity (%) measurement results (photostability)]

[0480]

[0481] [Table 108: WCX main (%) measurement results (photostability)]

[0482]

[0483] [Table 109: HIC hydrophobicity (%) measurement results (photostability)]

[0484]

[0485]

[0486] [Table 110: HIC hydrophilicity (%) measurement results (photostability)]

[0487]

[0488]

[0489] [Table 111: HIC main (%) measurement results (photostability)]

[0490]

[0491] [Table 112: SE-HPLC HMW (%) measurement results (freeze-thaw stability)]

[0492]

[0493]

[0494] [Table 113: SE-HPLC LMW (%) measurement results (freeze-thaw stability)]

[0495]

[0496]

[0497] [Table 114: SE-HPLC monomer (%) measurement results (freeze-thaw stability)]

[0498]

[0499] [Table 115: WCX acidity (%) measurement results (freeze-thaw stability)]

[0500]

[0501] [Table 116: WCX alkalinity (%) measurement results (freeze-thaw stability)]

[0502]

[0503]

[0504] [Table 117: Main (%) measurement results of WCX (freeze-thaw stability)]

[0505]

[0506]

[0507] [Table 118: SE-HPLC HMW (%) measurement results (stirring stability)]

[0508]

[0509] [Table 119: SE-HPLC LMW (%) measurement results (stirring stability)]

[0510]

[0511]

[0512] [Table 120: SE-HPLC monomer (%) measurement results (stirring stability)]

[0513]

[0514]

[0515] [Table 121: WCX acidity (%) measurement results (stirring stability)]

[0516]

[0517] [Table 122: WCX alkalinity (%) measurement results (stirring stability)]

[0518]

[0519]

[0520] [Table 123: WCX main (%) measurement results (stirring stability)]

[0521]

[0522]

[0523]

Discussion of the results

[0524] The appropriate concentration range of each component of the proline-containing formulation was confirmed through DoE statistical analysis of the test results of thermal stability, freeze-thaw stability, stirring stability and photostability.

[0525] The graphs obtained by stability modeling based on DoE statistical analysis of thermal stability test results (SE-HPLC and WCX results), stirring stability test results (SE-HPLC and WCX results), photostability test results (SE-HPLC, WCX and HIC results) and freeze-thaw stability results (SE-HPLC and WCX results) are shown in Figure 4 , Figure 5 , Figure 6 and Figure 7 middle.

[0526] according to Figure 4 , Figure 5 , Figure 6 and Figure 7 The results showed that in terms of stability, stirring stability, photostability and freeze-thaw stability, 150 mg / mL risankizumab had the best stability at pH 5.7 and in the presence of 16 mM histidine, 2.5 wt % proline stabilizer and 0.02 wt % polysorbate 20.

Claims

1. An aqueous pharmaceutical composition comprising: (a) risankizumab or an antigen-binding fragment thereof; and (b) stabilizers, wherein the aqueous pharmaceutical composition does not contain a polyol.

2. The aqueous pharmaceutical composition according to claim 1, wherein the stabilizer comprises an amino acid or a pharmaceutically acceptable salt thereof or sodium chloride.

3. An aqueous pharmaceutical composition comprising: (a) risankizumab or an antigen-binding fragment thereof; and (b) an amino acid or a pharmaceutically acceptable salt thereof, Wherein (c) the aqueous pharmaceutical composition has a pH of 5.0 to 7.

0.

4. The aqueous pharmaceutical composition according to claim 3, which does not contain a polyol.

5. The aqueous pharmaceutical composition of claim 2 or 3, wherein the amino acid comprises lysine, arginine, glycine, proline, histidine, phenylalanine, tyrosine, tryptophan, a pharmaceutically acceptable salt thereof, or a mixture thereof.

6. The aqueous pharmaceutical composition according to claim 1 or 4, wherein the polyol comprises sorbitol, sucrose, trehalose, mannose, maltose, mannitol or a mixture thereof.

7. The aqueous pharmaceutical composition according to any one of claims 1 to 4, further comprising a surfactant.

8. The aqueous pharmaceutical composition according to any one of claims 1 to 4, which does not contain a surfactant.

9. The aqueous pharmaceutical composition of claim 7, wherein the surfactant comprises polysorbate, poloxamer, sorbitan ester of another fatty acid, or a mixture thereof.

10. The aqueous pharmaceutical composition of claim 9, wherein the polysorbate comprises polysorbate 20, polysorbate 80, or a mixture thereof.

11. The aqueous pharmaceutical composition according to claim 1 or 2, which has a pH of 5.0 to 7.

0.

12. The aqueous pharmaceutical composition according to any one of claims 1 to 4, further comprising a buffer.

13. The pharmaceutical composition of claim 12, wherein the buffer comprises acetate, succinate, citrate, glutamate, glycine, lactate, maleate, phosphate, tartrate, histidine, or any combination of the foregoing.

14. The pharmaceutical composition according to any one of claims 1 to 4, wherein the concentration of risankizumab or its antigen-binding fragment is 9 mg / ml to 170 mg / ml.

15. The pharmaceutical composition according to any one of claims 1 to 4, wherein the pharmaceutical composition is for subcutaneous injection, intramuscular injection or intravenous injection.

16. The pharmaceutical composition according to any one of claims 1 to 4, wherein the pharmaceutical composition is used to treat autoimmune diseases, cancer, psoriasis, psoriatic arthritis, inflammatory bowel disease, multiple sclerosis, rheumatoid arthritis, ankylosing spondylitis, asthma or chronic obstructive pulmonary disease (COPD).

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