Immunoproteasome inhibitors

Abstract

Provided herein are lyophilized pharmaceutical formulations comprising KZR-616 or a salt thereof and a sugar, methods for preparing the formulations, methods for treating immune-related diseases, and methods for treating inflammation.

Description

[Background technology]

[0001] The compound, (2S,3R)-N-[(2S)-3-(cyclopent-1-en-1-yl)-1-[(2R)-2-methyloxiran-2-yl]-1-oxopropan-2-yl]-3-hydroxy-3-(4-methoxyphenyl)-2-[(2S)-2-[2-(morpholin-4-yl)acetamido]propanamide]propanamide ("KZR-616"), is useful as an immunoproteasome inhibitor. [ka]

[0002] In eukaryotes, protein degradation is primarily mediated through the ubiquitin pathway, in which proteins targeted for destruction are linked to the 76-amino acid polypeptide ubiquitin. Once targeted, ubiquitinated proteins then serve as substrates for the 26S proteasome, a multicatalytic protease that cleaves proteins into short peptides through the action of its three major proteolytic activities. While having a general function in intracellular protein turnover, proteasome-mediated degradation also plays an important role in many processes, such as major histocompatibility complex (MHC) class I antigen presentation, apoptosis, cell growth regulation, NF-κB activation, antigen processing, and the transmission of proinflammatory signals.

[0003] WO 2014 / 152134 describes tripeptide epoxyketone proteasome inhibitors and methods of using these compounds to treat diseases and conditions associated with abnormal immunoproteasome activity. Because tripeptide epoxyketone proteasome inhibitors such as KZR-616 are useful in treating diseases and conditions in patients, there is a need for soluble and stable formulations of KZR-616. [Patent Document 1] International Publication No. 2014 / 152134 Summary of the Invention

[0004] Provided herein is a pharmaceutical formulation comprising KZR-616, or a salt thereof, and a sugar, wherein KZR-616 has the structure: [ka] In some embodiments, the formulation is a lyophilized formulation. In some embodiments, the KZR-616 is a maleate salt. In some cases, the formulation has a pH of 3.0 to 4.5, as measured prior to lyophilization. In some cases, the formulation has a pH of 3.8 to 4.3. In some cases, the formulation has a pH of 4.2. In some embodiments, the formulation does not include a buffer. In some embodiments, the KZR-616 or a salt thereof is present in an amount of 50 to 300 mg / mL, as measured prior to lyophilization. In some embodiments, the KZR-616 or a salt thereof is present in an amount of 100 mg / mL to 200 mg / mL. In some cases, the KZR-616 or a salt thereof is present in an amount of 125 mg / mL, based on the weight of the KZR-616 free base.

[0005] In some cases, the sugar is present in an amount of 0.1% to 5% by weight based on the total weight of the formulation. In some embodiments, the sugar is present in an amount of 1.5% to 3% by weight. In some embodiments, the sugar is present in an amount of 2% by weight. In some embodiments, the sugar comprises mannitol, trehalose, or a combination thereof. In some cases, the sugar comprises trehalose.

[0006] In an embodiment, the lyophilized formulation has a water content of less than 1%. In some cases, the formulation has a pH of 4.1 to 4.3 (measured upon reconstitution) after storage at 5° C., 25° C., or 40° C. for at least 6 months. In some cases, the formulation has a concentration of KZR-616 or a salt thereof of 140 mg / mL to 150 mg / mL (measured upon reconstitution) after storage at 5° C., 25° C., or 40° C. for at least 6 months. In some cases, the formulation has a reconstitution time of less than 4.5 minutes after storage at 5° C., 25° C., or 40° C. for at least 6 months. In some cases, the KZR-616 or a salt thereof is at least 93% pure after storage at 5° C., 25° C., or 40° C. for at least 6 months.

[0007] Also provided herein is a reconstituted formulation comprising the formulation disclosed herein and a reconstitution solvent. In an embodiment, the reconstitution solvent comprises water for injection (WFI) or 5% aqueous dextrose. In an embodiment, the reconstituted formulation has an osmolality of 275-325 mOsm. In an embodiment, the reconstituted formulation is for subcutaneous injection.

[0008] In some cases, the reconstituted formulation is 1 / 2 In an embodiment, the reconstituted formulation has a C of at least 150% that of the formulation comprising KZR-616 or a salt thereof and polysorbate-80, as measured by max has.

[0009] Also provided herein is a method for preparing the formulation described herein, comprising mixing KZR-616 or a salt thereof with a sugar to form an aqueous solution, and lyophilizing the aqueous solution to form a lyophilized formulation. Optionally, the method further comprises reconstituting the lyophilized formulation with a solvent to form a reconstituted formulation. In an embodiment, the solvent is water for injection or 5% aqueous dextrose. Further provided herein is a method for treating immune-related disease in a subject, comprising administering the reconstituted formulation described herein. Optionally, the subject suffers from lupus nephritis or systemic lupus erythematosus (SLE). Also provided herein is a method for treating inflammation in a subject by administering the reconstituted formulation described herein. [Brief description of the drawings]

[0010] [Figure 1A] Figure 2 shows RP-HPLC analysis of various formulations after 6 months storage at 5°C. [Figure 1B] Figure 2 shows RP-HPLC analysis of various formulations after 6 months storage at 25°C. [Figure 1C] Figure 2 shows RP-HPLC analysis of various formulations after 6 months storage at 40°C. [Figure 2A] Figure 1 shows the RP-HPLC analysis trends for various formulations after 6 months storage at 5°C. [Figure 2B] Showing the trends of RP-HPLC analysis of various formulations after 6 months storage at 25°C [Figure 2C] Figure 2 shows the RP-HPLC analysis trends for various formulations after 6 months storage at 40°C. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0011] Provided herein is a formulation of KZR-616 or its salt that is lyophilized and stable.Lyophilized formulations provide advantages including one or more of longer shelf life, easier storage at refrigerated temperature, and reduced weight and / or volume of formulation for transportation and handling.Reconstituted formulations of lyophilized formulations, methods of preparing formulations, and methods of using disclosed formulations to treat immune-related disease and / or inflammation are also disclosed.

[0012] As further described below, the pharmaceutical formulations of KZR-616 or a salt thereof herein can have several advantages, for example, the formulations disclosed herein can be resistant to changes in pH, concentration, reconstitution time, moisture content, and / or purity after storage at various temperatures and / or times, such as after storage at 5°C, 25°C, or 40°C for at least 6 months.

[0013] The formulations, dosage indications, and methods are intended to include embodiments that include one or more of any combination of the additional optional elements, features, and steps further described below (including those shown in the Tables), unless otherwise indicated.

[0014] As used herein, the term "treating" or "treatment" includes reversing, alleviating, or arresting the symptoms, clinical signs, and underlying pathology of the condition in a manner that improves or stabilizes the patient's condition.

[0015] The use of the terms "a", "an", "the" and similar referents in the context of this disclosure (especially in the context of the claims) herein should be construed to encompass both the singular and the plural unless otherwise indicated. The recitation of ranges of values ​​herein is merely intended to serve as a shorthand way of individually referring to each individual value falling within the range, unless otherwise indicated herein, and each individual value is incorporated herein as if it were individually recited herein. Any and all examples provided herein, or the use of illustrative language (e.g., "etc."), are intended to better illustrate the invention, and are not a limitation of the scope of the invention unless otherwise indicated. No language in this specification should be construed as indicating any element not recited in the claims as essential to the practice of the disclosure.

[0016] As used herein, the terms "about" and "approximately" generally mean plus or minus 10% of the stated value. For example, about 0.5 includes 0.45 and 0.55, about 10 includes 9-11, and about 1000 includes 900-1100.

[0017] Those skilled in the art of organic chemistry will understand that many organic compounds can form complexes with the solvents in which they react or in which they are precipitated or crystallized. These complexes are known as "solvates." For example, complexes with water are known as "hydrates." Solvates of KZR-616 used in the formulations herein are within the scope of the present invention.

[0018] Lyophilized preparations Provided herein is a pharmaceutical formulation comprising KZR-616, or a salt thereof, and a sugar, wherein KZR-616 has the structure: [ka] Here, the formulation is a lyophilized formulation.

[0019] The pharmaceutical formulations disclosed herein may include any suitable form of KZR-616. In embodiments, KZR-616 is present as a free base. In embodiments, KZR-616 is present as a salt. In some cases, KZR-616 is present as a salt form selected from maleate, fumarate, oxalate, malate, sulfate, methanesulfonate, 2-naphthalenesulfonate, phosphate, halide, tartrate, citrate, tosylate, propionate, benzoate, and combinations thereof. In some cases, KZR-616 is present as a solvate, such as a hydrate. In embodiments, KZR-616 is present as a maleate, e.g., monomaleate. In some embodiments, KZR-616 is present as a dimer salt, e.g., bis(KZR-616) monomaleate.

[0020] The formulations described herein can contain any concentration of KZR-616 or a salt thereof, as measured prior to lyophilization. Specific contemplated concentrations of KZR-616 or a salt thereof include 10 mg / mL to about 500 mg / mL, or about 25 mg / mL to about 400 mg / mL, or about 50 mg / mL to about 400 mg / mL, or about 50 mg / mL to about 300 mg / mL, or about 60 mg / mL to about 275 mg / mL, or about 70 mg / mL to about 250 mg / mL, or about 80 mg / mL to about 225 mg / mL, or about 90 mg / mL to about 225 mg / mL, or about 100 mg / mL to about 200 mg / mL, or about 110 mg / mL to about 200 mg / mL, or about 120 mg / mL to about 200 mg / mL, or about 120 mg / mL to about 220 mg / mL, or about 120 mg / mL to about 190 mg / mL, or about 120 mg / mL to about 180 mg / mL, or is about 120 mg / mL to about 170 mg / mL, or about 120 mg / mL to about 160 mg / mL, or about 120 mg / mL to about 150 mg / mL, or about 130 mg / mL to about 150 mg / mL, or about 140 mg / mL to about 150 mg / mL, or about 100 mg / mL to about 150 mg / mL, or about 100 mg / mL to about 140 mg / mL, or about 100 mg / mL to about 130 mg / mL, or about 100 mg / mL to about 120 mg / mL, or about 50 mg / mL to about 100 mg / mL, or about 60 mg / mL to about 90 mg / mL, or about 90 mg / mL to about 120 mg / mL, for example, about 150 mg / mL based on the weight of KZR-616 maleate or about 125 mg / mL based on the weight of KZR-616 free base. In embodiments, the concentration of KZR-616 is measured upon reconstitution of the lyophilized formulation after the lyophilized formulation has been stored, for example, at 5°C, 25°C, or 40°C for at least 6 months.

[0021] The pharmaceutical formulations disclosed herein can include a sugar, such as a reducing or non-reducing sugar, or a sugar alcohol. The formulation of KZR-616 or its salt with a sugar is stable when stored at 5°C, 25°C, or 40°C for at least 6 months. In embodiments, the sugar can include mannitol, trehalose, dextrose, glucose, fructose, mannose, galactose, xylose, sucrose, lactose, sorbitol, dextran, erythritol, or a combination thereof. In embodiments, the sugar can include mannitol and / or trehalose. In embodiments, the sugar includes trehalose.

[0022] In embodiments, the formulation contains a sugar component in an amount, for example, from about 0.01% to about 10% by weight, or from about 0.1% to about 10% by weight, or from about 0.1% to about 9% by weight, or from about 0.1% to about 8% by weight, or from about 0.1% to about 7% by weight, or from about 0.1% to about 6% by weight, or from about 0.1% to about 5% by weight, or from 0.1% to about 4% by weight, or from about 0.5% to about 5% by weight, or from about 0.5% to about 4% by weight, or about 0.5% by weight In an embodiment, the water content is measured after the lyophilized formulation has been stored for at least 6 months, e.g., at 5°C, 25°C, or 40°C. In an embodiment, the formulation described herein can have a water content of up to 5% by weight, e.g., up to 4%, 3%, 2%, 1.5%, 1%, or 0.5% by weight. In embodiments, the formulations described herein can have a moisture content of up to 5%, up to 4%, 3%, 2%, 1.5%, 1%, or 0.5% after storage at 5° C., 25° C., or 40° C. for a period of time, e.g., at least 6 months, and can have a moisture content of less than 1% after storage at 5° C., 25° C., or 40° C. for at least 6 months.

[0023] In some cases, the formulations described herein may optionally include a buffer. In some cases, the formulations do not include a buffer. As used herein, the term "buffer" refers to a buffer that is added to any salt form of KZR-616. In embodiments, the secondary buffer may include succinate, citrate, phosphate, histidine, acetate, maleate, gluconate, or a combination thereof. In embodiments, the buffer includes a succinate buffer. In embodiments, the buffer, such as succinate buffer, can have a concentration, as measured prior to lyophilization, of about 1 mM to about 50 mM, or about 1 mM to about 40 mM, or about 1 mM to about 30 mM, or about 1 mM to about 25 mM, or about 1 mM to about 20 mM, or about 1 mM to about 15 mM, or about 1 mM to about 10 mM, or about 2 mM, or about 3 mM, or about 4 mM, or about 5 mM, or about 6 mM, or about 7 mM, or about 8 mM, or about 9 mM, or about 10 mM, or about 11 mM, or about 12 mM, or about 13 mM, or about 14 mM, or about 15 mM, e.g., 10 mM.

[0024] The formulations described herein can optionally further comprise a bulking agent in addition to or in place of the sugar. Contemplated additives such as bulking agents include glycine, serine, cysteine, or combinations thereof.

[0025] In one aspect, the formulations described herein can have the following advantages: (1) the lyophilisate exhibits a visually clear and / or particle-free cake upon reconstitution; (2) the moisture content of the lyophilisate is 1% (wt % or mol %) or less; (3) the lyophilisate has a fast lyophilisation and / or reconstitution time compared to other pharmaceutical formulations, as detailed in the Examples below; and / or (4) the formulation has a desirable stability, e.g., less than 0.5% (wt % or mol %) degradation when measured at 5° C. for at least 6 months, less than 7% (wt % or mol %) degradation when measured at 25° C. for at least 6 months, and less than 20% (wt % or mol %) degradation when measured at 40° C. for at least 6 months.

[0026] The formulations disclosed herein can have a pH of 3.0 to 4.5, e.g., 3.8 to 4.3, when measured prior to lyophilization. In some cases, the pH prior to lyophilization is 4.2. In embodiments, the formulations described herein can have a pH of about 2 to about 8 or about 3 to about 5, when measured upon reconstitution, at a temperature of about 25° C. In embodiments, the formulations can have a pH of about 3.1 to about 4.9, or about 3.2 to about 4.8, or about 3.3 to about 4.7, or about 3.4 to about 4.6, or about 3.5 to about 4.5, or about 3.6 to about 4.4, or about 3.7 to about 4.3, or about 3.7 to about 4.3, or about 3.8 to about 4.3, or about 3.9 to about 4.2, when measured upon reconstitution, at a temperature of about 25° C., e.g., a pH of about 4.2, at a temperature of about 25° C. In embodiments, the pH is measured upon reconstitution of the lyophilized formulation after the lyophilized formulation has been stored, for example, at 5°C, 25°C, or 40°C for at least 6 months.

[0027] In embodiments, the formulation may have a purity of greater than 70% API (KZR-616 or a salt thereof) after storage for at least 6 months at 5° C., 25° C., or 40° C., for example, greater than 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% after storage for at least 6 months at 5° C., 25° C., or 40° C., for example, greater than 93% purity after storage for at least 6 months at 5° C., 25° C., or 40° C. Purity of greater than 70% means that the amount of API in the formulation after the indicated storage time and temperature is greater than 70% (wt % or molar %) of the amount of API in the formulation before storage.

[0028] In embodiments, the formulations disclosed herein can have a reconstitution time of 10 minutes or less, e.g., 9 minutes or less, 8 minutes or less, 7 minutes or less, or 6 minutes or less, 5 minutes or less, 4.5 minutes or less, 4 minutes or less, 3.5 minutes or less, or 3 minutes or less. In embodiments, the formulations disclosed herein can have a reconstitution time of 10 minutes or less, e.g., 9 minutes or less, 8 minutes or less, 7 minutes or less, or 6 minutes or less, 5 minutes or less, 4.5 minutes or less, 4 minutes or less, 3.5 minutes or less, or 3 minutes or less, after storage at 5°C, 25°C, or 40°C for at least 6 months, e.g., the formulations disclosed herein can have a reconstitution time of 4.5 minutes or less after storage at 5°C, 25°C, or 40°C for at least 6 months. As used herein, the term "reconstitution time" refers to the time required to completely dissolve a lyophilized formulation according to the reconstitution method detailed below.

[0029] Reconstituted Formulation The formulations disclosed herein can be lyophilized into cake form, powder form, or the cake and / or powder can be further processed into gel, foam, aerosol, film, or other formulations.In embodiments, the formulations can be reconstituted with a solvent for administration to a subject in need thereof.Therefore, a kit is provided herein that includes the formulation, such as a lyophilized composition, and a reconstitution solution.

[0030] Provided herein is a reconstituted formulation of KZR-616 or a salt thereof, comprising a lyophilized pharmaceutical formulation as described herein and a reconstitution solvent. The reconstitution solvent can comprise any one or more solvents suitable for those skilled in the art. In embodiments, the reconstitution solvent can comprise water for injection (WFI), saline, or 5% aqueous dextrose. In some specific embodiments, the reconstitution solvent comprises water for injection. In embodiments, the reconstitution solvent does not contain chloride ions (e.g., is not saline). In embodiments, the reconstituted formulation can be used for subcutaneous injection.

[0031] In embodiments, the osmolarity of the reconstituted formulation may be between 200 mOsm and 400 mOsm, such as between 250 mOsm and 400 mOsm, or between 275 mOsm and 350 mOsm, or between 275 mOsm and 325 mOsm, e.g., about 300 mOsm, which is an approximately isotonic osmolarity.

[0032] In embodiments, the reconstituted formulations herein include 1 / 2 When measured by the absorption rate, the composition may have an absorption rate that is at least 10% faster than that of a formulation containing KZR-616 or a salt thereof and polysorbate-80 ("PS-80") ("PS-80 formulation"), for example, at least 20% faster, at least 30% faster, at least 40% faster, at least 50% faster, or at least 60% faster, e.g., at least 50% faster, than that of a PS-80 formulation.

[0033] In embodiments, the reconstituted formulations described herein have a C of at least 50% of that of the PS-80 formulation. max For example, the PS-80 formulation may have a C of at least 100%, at least 110%, at least 120%, at least 130%, at least 140%, at least 150%, at least 160%, at least 170%, or at least 180%, e.g., at least 150%. max may have the following structure:

[0034] Preparation method The pharmaceutical formulation disclosed herein can be prepared, for example, by mixing KZR-616 or its salt with a sugar to form a solution. In an embodiment, the solution is an aqueous solution. In an embodiment, the solution can be lyophilized to form a lyophilized pharmaceutical formulation. The method of preparing the pharmaceutical formulation can further include reconstituting the lyophilized formulation with a solvent to form a reconstituted formulation as described herein. The solvent used to form the reconstituted formulation can be any solvent suitable for a person skilled in the art to reconstitute a lyophilized formulation, for example, the solvent can include water or 5% aqueous dextrose.

[0035] treatment method Another aspect of the present disclosure provides a method of treating an immune-related disease in a subject by administering a reconstituted formulation described herein. In some cases, the disease is psoriasis, dermatitis, systemic scleroderma, sclerosis, Crohn's disease, ulcerative colitis; respiratory distress syndrome, meningitis, encephalitis, uveitis, colitis, glomerulonephritis, eczema, asthma, chronic inflammation, atherosclerosis, leukocyte adhesion deficiency, rheumatoid arthritis, systemic lupus erythematosus (SLE), diabetes mellitus, multiple sclerosis, Raynaud's syndrome, autoimmune thyroiditis, allergic encephalomyelitis, Sjogren's syndrome, juvenile onset diabetes, tuberculosis, sarcoidosis, polymyositis, granulomatosis, vasculitis, pernicious anemia (Addison's disease), white blood cells, and the like. The disorder is a disease with cytopenia; a central nervous system (CNS) inflammatory disease; multiple organ injury syndrome; hemolytic anemia; myasthenia gravis; an antigen-antibody complex-mediated disease; antiglomerular basement membrane disease; antiphospholipid syndrome; allergic neuritis; Graves' disease; Lambert-Eaton myasthenic syndrome; pemphigoid; pemphigus; autoimmune polyendocrinopathy; Reiter's disease; stiff-body syndrome; Behçet's disease; giant cell arteritis; immune complex nephritis; IgA nephropathy; IgM polyneuropathy; immune thrombocytopenic purpura (ITP) or autoimmune thrombocytopenia. In various cases, the disorder is lupus, lupus nephritis, rheumatoid arthritis, diabetes, scleroderma, ankylosing spondylitis, psoriasis, multiple sclerosis, Hashimoto's disease, meningitis, or inflammatory bowel disease. In an embodiment, the disease is lupus nephritis or SLE.

[0036] Another aspect of the disclosure provides a method of treating inflammation in a subject by administering a reconstituted formulation described herein.

[0037] Reconstruction method As described herein, the lyophilized pharmaceutical formulation is placed in a vial. A certain amount of filtered Milli-Q® ("MQ") water, such as milliliters, is added to the vial. Upon addition of the MQ water, a timer is started to measure the time until the lyophilized pharmaceutical formulation is reconstituted and / or completely dissolved. Every 30 seconds, the vial containing the MQ water and the lyophilized formulation is gently swirled until reconstitution is complete. EXAMPLES

[0038] Example 1 The stability of KZR-616 under various formulation conditions was investigated in this lyophilized formulation development study. Conditions investigated in this study included buffered (10 mM succinate) and unbuffered formulations with various excipients (L-glycine, mannitol, and trehalose) at various pH values ​​(3.9-4.2). The stability of KZR-616 at 150 mg / mL in the various formulations was examined under static storage conditions for up to 6 months at refrigerated (5°C), ambient (25°C), and accelerated (40°C) temperatures. During the course of this 6-month study, reversed-phase HPLC (RP-HPLC) analysis was the primary analytical method used to monitor the stability of KZR-616.

[0039] The lyophilized cakes showed a consistent appearance over 6 months of storage at all storage temperatures. Some increase in yellow coloration of the lyophilisates was observed over time, especially at 25°C and 40°C. This yellow coloration became more pronounced after reconstitution. After reconstitution, all formulations were free of visible particles. Over the 6-month storage period, the pH and KZR-616 concentration values ​​of the formulations remained comparable to those observed at time zero. Reconstitution times also remained consistent over the 6-month storage period at less than 4 minutes and 30 seconds.

[0040] The stability of KZR-616 was evaluated using RP-HPLC analysis. After 6 months of storage at 5°C, all formulations showed a slight decrease in the ratio of the main peak than that observed at time zero. After storage at 25°C, the formulations showed a decrease in the ratio of the main peak, with the non-buffered trehalose-containing formulation showing a higher ratio of the main peak than the other formulations. After 6 months of storage at 40°C, all formulations showed a significant decrease in the ratio of the main peak, mostly due to an increase in the degradant peaks of unknown RRT=0.1, KZR-59587, unknown RRT=0.5, KZR-0214143 and unknown RRT=2.2. The non-buffered trehalose-containing formulations with or without mannitol showed the highest ratio of the main peak among the formulations tested.

[0041] Since no degradation was expected at 5°C, a lyophilization cycle optimization study was performed to determine whether optimization of the lyophilization cycle could further reduce the residual moisture content and thereby improve the stability of the formulation. This moisture optimization study was performed by lyophilizing the lead formulation at various secondary shelf temperatures of 10°C, 25°C, 35°C, and 50°C. However, after 4 weeks of storage at 40°C, no differences were observed in visual appearance, pH, concentration, reconstitution time, and peak percentage by RP-HPLC, regardless of secondary drying shelf temperature.

[0042] Based on the results of this study, the primary formulation of lyophilized 150 mg / mL KZR-616 was identified as a formulation containing 2% trehalose at pH 4.2 with no additional buffer.

[0043] material The active pharmaceutical ingredient (API) examined in this study was KZR-616. The materials used in this study consisted of the following: (1) Drug Substance (DS): KZR-616 Maleate (Lot A) (2) Drug Substance (DS): KZR-616 Maleate (Lot B) (3) Reference Standard (RS): KZR-616 Maleate Material

[0044] The drug substance was stored at 2-8 °C prior to use. Formulation parameters

[0045] In the accelerated stability study, the following parameters were kept constant: (1) Filling amount: 1.0mL (2) Buffer concentration: 10 mM or 0 mM The following formulation (F1-F9) parameters were investigated: (1)API concentration: 150mg / mL (2) Buffer: Succinate (3) pH: 3.9 to 4.2 (4) Tonicity adjuster: 0.2% glycine, 1% trehalose, 0.4% mannitol, 0.4% glycine, 1% mannitol, 2% trehalose, 0.3% glycine, 0.8% mannitol [Table 1]

[0046] Preparation of formulations Freeze Drying Cycle Development #1 To determine the storage cycle parameters, sub-ambient differential scanning calorimetry (DSC) (PerkinElmer Pyris Diamond) was performed on three representative formulations, F3, F4, and F5, each containing glycine, mannitol, or trehalose. The method program used was as follows: isothermal hold at -60°C for 1 min, heat from -60°C to 25°C at 5°C / min. F3, F4, and F5 have glass transition temperatures (T) of -16°C, -15°C, and -33°C, respectively. g ) (Table 1). For F4, the devitrification temperature (T d ), an annealing step was added prior to primary drying to obtain more complete crystallization. This data was implemented in the first freeze-drying cycle (Table 3).

[0047] Based on the subatmospheric DSC results, a conservative lyophilization cycle was designed to produce samples with a residual moisture concentration of <1%. Table 3 displays the parameters of the first conservative cycle.

[0048] KZR-616 maleate was dissolved in filtered Milli-Q® (MQ) water at a target concentration of 170 mg / mL, and the KZR-616 in MQ was heated at 60° C. for approximately 2 minutes to dissolve the drug substance into solution. Formulations were prepared as 10x stocks and added to aliquots of the DS / MQ solution at a ratio of 1:9 (buffer:DS). The pH of each formulation was titrated to the target pH with NaOH. Under aseptic conditions in a BSC, all samples were sterile filtered through a 0.2 μm filter and filled into 3 cc vials with a fill volume of 1.0 mL. Samples were partially stoppered with sterile stoppers and placed in the lyophilizer surrounded by empty vials. Samples were lyophilized using the parameters detailed in Table 3. Following the first lyophilization cycle, samples were reconstituted with 1 mL of filtered MQ water and analyzed. Lyophilization cycle development formulations were prepared per Table 2. [Table 2] [Table 3]

[0049] Freeze Drying Cycle Development #2 KZR-616 was dissolved in filtered MQ water at a target concentration of 170 mg / mL. The KZR-616 in MQ was then heated at 60°C for approximately 2 minutes to dissolve the drug substance into solution. Formulations were prepared as 10x stocks and added to aliquots of the DS / MQ solution at a ratio of 1:9 (buffer:DS). The pH of each formulation was titrated to the target pH with NaOH. All samples were sterile filtered through a 0.2 μm filter and filled into 3 cc vials with a fill volume of 1.0 mL under aseptic conditions in a BSC. A summary of the formulation matrix (buffer, bulking agent concentration, pH) is provided in Table 1. Samples were partially stoppered with sterile stoppers and placed in the lyophilizer surrounded by empty vials. Samples were lyophilized using the parameters detailed in Table 4. After lyophilization, samples were reconstituted in 1 mL of filtered MQ water and analyzed according to the methods outlined in Table 8. [Table 4]

[0050] Accelerated Stability Testing Once IBI identified a freeze-drying cycle that produced an elegant cake with less than 1% moisture content, the experiment was repeated to confirm the cycle results and to prepare samples for accelerated stability studies.

[0051] KZR-616 was dissolved in filtered MQ water at a concentration of 170 mg / mL as a stock solution. The KZR-616 stock solution was heated at 60°C for approximately 2 minutes to completely dissolve the drug substance. Formulation buffers were prepared as 10x stock solutions. Each formulation was prepared using 10x buffer stock, DS stock (170 mg / mL), and MQ to target a final concentration of 150 mg / mL. The pH of each formulation was adjusted to the target pH using NaOH. Under sterile conditions in a BSC, all samples were sterile filtered through a 0.2 μm filter and filled into 3 cc vials with a fill volume of 1.0 mL. Samples were partially stoppered with sterile stoppers and placed in the lyophilizer surrounded by empty vials. Samples were lyophilized using the parameters detailed in Table 4.

[0052] After lyophilization, the product quality and integrity were assessed by comparing the pre-lyophilized liquid sample, the lyophilized cake, and the reconstituted sample using the analytical methods listed in Table 9.

[0053] Moisture Optimization Two different drug substance lots of KZR-616 (KZR-0214142 and C15072369-FF1600) were individually dissolved in MQ at a concentration of 170 mg / mL and heated at 60 °C for approximately 2 minutes to completely dissolve the drug substance. Both 170 mg / mL preparations were then diluted to 150 mg / mL with 10x the target buffer (F5, 2% trehalose at pH 4.2). The pH of each formulation was titrated to the target pH using NaOH. Under sterile conditions in a BSC, all samples were sterile filtered through a 0.2 μm filter and filled into 3 cc vials with a fill volume of 1.0 mL. Samples were partially stoppered with sterile stoppers and placed in the lyophilizer, surrounded by empty vials to prevent migration. Samples were lyophilized using the parameters detailed in Table 5.

[0054] The lyophilization cycle was preprogrammed with the first secondary drying temperature (10°C). Following the first secondary drying temperature, all subsequent operations of the lyophilizer were performed manually to accommodate all shelf temperatures. Each secondary drying temperature was run for at least 8 hours minimum. At the end of each secondary drying temperature, the vials at that temperature were stoppered, the lyophilizer was manually stopped, and the vacuum was purged. The stoppered tray was then removed and the lyophilizer was manually restarted for the next secondary drying temperature. The last step was repeated for each secondary drying temperature. After lyophilization, the samples were placed in stable storage conditions and the T=0 sample was reconstituted in 1 mL of filtered MQ water and analyzed. The analytical method is outlined in Table 8. [Table 5]

[0055] Stress conditions acceleration stability The stability of the lyophilized formulations was tested under the following conditions as listed in Table 6. [Table 6]

[0056] Moisture Optimization The stability of the lyophilized formulations was tested under the following conditions as listed in Table 7. [Table 7]

[0057] Analysis method Freeze-drying cycle development (1&2) Various analyses were performed on the samples following lyophilization cycles 1 and 2. Table 8 summarizes these analyses for the stability studies. [Table 8]

[0058] Accelerate your stability testing Various analyses were performed on the lyophilized samples after incubation at each temperature and time point. At each of the following times, samples were analyzed for appearance, pH, concentration, reconstitution time, RP-HPLC, and moisture content. Table 9 summarizes the various analyses of the accelerated stability study. [Table 9] [Table 10]

[0059] Analytical methods for all the aforementioned studies are detailed below. (1) Visual Inspection: Visual inspection was performed under a white light source (13 W fluorescent lamp) against a black and white background. Digital photographs were taken of the pre-lyophilized, post-lyophilized, and reconstituted formulations. (2) Osmolality: Osmolality was measured at time zero for each objective test. (3) pH: pH analysis was performed using a SympHony® pH meter (VWR Scientific, Cat. No. SB70P) and calibrated with three pH standard solutions (pH 4, 7, and 10) with a calibration slope of ≥95%. Samples were equilibrated to ambient temperature prior to measurement. (4) Absorbance spectrophotometry: The concentration of KZR-616 was determined via a Beckman Coulter spectrophotometer. An extinction coefficient of 1.8495 (mg / mL)-1cm-1 was determined by constructing a standard curve (1mg / mL, 0.75mg / mL, 0.5mg / mL, and 0.25mg / mL, serially diluted with MQ). KZR-616 was analyzed with a full wavelength scan and showed maximum absorbance at 272nm. This wavelength was used to measure the concentration. (5) FTIR: Pre- and post-lyophilized formulations were analyzed to determine the fingerprint of the formulation containing bulking agents. (Post-lyophilized development cycles #1 and #2 were performed) (6) Reconstitution Time: The lyophilized formulation was reconstituted with 1 milliliter of filtered water. Upon addition of water, a timer was started to measure the time it took for the KZR-616 drug formulation to reconstitute. Every 30 seconds, the reconstituted vial was gently swirled. This procedure was repeated until reconstitution was complete. (7) Karl Fischer titration: The formulations after lyophilization were analyzed to determine the moisture content of the lyophilized product. (8) DSC: Subatmospheric DSC analysis was performed on each formulation from -60°C to 25°C at 5°C / min to determine the recrystallization, Tg, and other important thermal transition data of the filler in the frozen state.

[0060] (9) RP-HPLC: The RP-HPLC method used in this study was transferred from Kezar and the parameters were as follows: Mobile phase A: 75% 5 mM ammonium acetate, 25% acetonitrile Mobile phase B: 65% 5 mM ammonium acetate, 35% acetonitrile Mobile phase C: 15% 5 mM ammonium acetate, 85% acetonitrile Column: Agilent Poroshell 120 EC-C18, 4.6 x 75 mm, 2.7 μm (P / N:697975-902) Equipment: Agilent 1100 HPLC Column temperature: 40℃ Autosampler temperature: 5℃ Flow rate: 1.5mL / min Duration: 38 minutes Detection: 210nm Sample preparation: Dilute to 1.8mg / mL in 70% ACN Filling volume: 18μg (10μL at 1.8mg / mL)

[0061] Moisture Optimization Test Various analyses were performed on the lyophilized samples after incubation at each temperature and time point. At each of the following times, samples were analyzed for appearance, pH, concentration, reconstitution time, RP-HPLC, and moisture content. Table 11 summarizes these analyses for the stability study. [Table 11]

[0062] Results and Discussion Freeze-drying cycle development test Multiple freeze-drying cycles were performed to develop one that produced a cake with the following attributes: ·Elegant cake Moisture content within specification range (<1.0%) Reduced freeze-drying and reconstitution times for lyophilized cakes Good product stability

[0063] FT-IR (Fourier Transform Infrared Spectroscopy) was performed before and after lyophilization development cycles #1 and #2. FT-IR showed good structural stability before lyophilization, during lyophilization, and upon reconstitution. The stresses (pressure and temperature) caused by lyophilization did not destroy the structural integrity of KZR-616 in the various formulations.

[0064] Visual Analysis At time zero, both development cycles showed an elegant cake. Development cycles #1 and #2 showed samples that were visually clear, particle free, and slightly yellow in color.

[0065] Formulation osmolality Osmolality of the nine formulations was measured after sample preparation and prior to any stress exposure. A summary of the formulations and specific reagents for lyophilization cycle #1 is shown in Table 2. After confirming the osmolality of cycle #1 formulations (Table 12), the bulking agent concentration was reduced in cycle #2 to target an isotonic osmolality (approximately 300 Osmo). Lyophilization cycle #2 demonstrated isotonic osmolality measurements before lyophilization and after reconstitution (Table 12). [Table 12]

[0066] pH Measurement Lyophilization cycles 1 and 2 showed comparable pH values ​​before lyophilization and after reconstitution (Table 13). However, after reconstitution, lyophilization cycle #2 showed a pH value closer to the target. [Table 13]

[0067] Reconstruction Time Measurements Following lyophilization cycles 1 and 2, the formulations were reconstituted with 1 milliliter of filtered MQ. Upon addition of the MQ, a timer was started to measure the time required for each formulation to homogenize. Lyophilization cycle #1 exhibited a reconstitution time of 4:31 minutes or less, while cycle #2 exhibited a reconstitution time of 4:12 minutes or less (Table 14). [Table 14]

[0068] Moisture content measurement (Karl Fischer) Following lyophilization cycles 1 and 2, the formulations exhibited moisture values ​​comparable to each other (Table 15). All lyophilized formulations exhibited moisture values ​​below 1%. Lyophilization cycle #1 exhibited moisture values ​​ranging from 0.28% to 0.60%. Lyophilization cycle #2 exhibited moisture values ​​ranging from 0.21% to 0.39%. [Table 15]

[0069] RP-HPLC analysis After 4 weeks of incubation at 40°C, all reconstituted formulations showed a decrease in the percentage of the main peak (90.3%-91.7%) compared to that observed at time zero (97.7%-98.6%). RP-HPLC identified the major degradant species of the KZR-616 molecule. Peaks RRT=0.1 displayed KZR-59587, KZR-0214143, and RRT=2.2 showed the greatest increase in peak percentage when stressed. Variation in secondary drying temperature showed little difference in the stability of the main peak at time zero. After 4 weeks at 40°C, all formulated samples showed comparable stability (Tables 16-19). [Table 16] [Table 17] [Table 18] [Table 19]

[0070] Accelerated Stability Testing For accelerated stability studies, nine buffered (succinate) and non-buffered formulations containing various concentrations of bulking agent at pH 3.9 and pH 4.2 were prepared at 150 mg / mL.

[0071] Visual Analysis Following 6 months of temperature-related stress, the appearance of the lyophilized formulation exhibited an elegant cake. The lyophilized cake exhibited a slight yellowish tint that increased with increasing temperature. After reconstitution, the formulation was clear, particle-free, and also exhibited a yellowish color. After 6 months of incubation, the formulation at 5° C. and 25° C. exhibited coloration comparable to time zero. At 40° C., the formulation exhibited a darker yellowish tint compared to time zero.

[0072] Formulation osmolality The osmolality of the nine formulations was measured after sample preparation and prior to any stress exposure, as shown in Table 20. Bulking agent concentrations were based on the results of lyophilization cycle Development #2, targeting an isotonic formulation at approximately 300 mOsm. After six months, formulation F5 demonstrated osmolality values ​​equivalent to those observed at time zero. [Table 20]

[0073] pH Measurement The pH of the nine formulations was measured at all time points following temperature-related stress. After six months, the formulations exhibited pH values ​​equivalent to time zero, as shown in Table 21. [Table 21]

[0074] Concentration measurements After six months, the formulations demonstrated concentration measurements equivalent to those observed at time zero (Table 22). [Table 22]

[0075] Reconstruction Time Measurements At each time point after lyophilization, the formulations were reconstituted with 1 milliliter of filtered MQ. Upon addition of the MQ, a timer began measuring the time required for each formulation to homogenize. After 6 months, the formulations showed reconstitution times equivalent to time zero. After 6 months, all formulations showed reconstitution times less than 3 minutes 30 seconds (Table 23). [Table 23]

[0076] Moisture content measurement (Karl Fischer) After 6 months of incubation, the formulations showed moisture content values ​​comparable to time zero (Table 24). All lyophilized formulations continued to show moisture values ​​below 1%. [Table 24]

[0077] RP-HPLC analysis After 6 months of incubation at 5°C, all reconstituted formulations showed a decrease in the percentage of the main peak (97.9%-98.1%) compared to that observed at time zero (98.2%-98.4%), as seen in Figure 1A. After 6 months of incubation at 25°C, all reconstituted formulations showed an increase in the degradant peaks compared to those stored at 5°C. Formulations F2 and F5 showed the highest percentage of KZR-616 (93.4% and 93.1%, respectively), as seen in Figure 1B. After 6 months of incubation at 40°C, all reconstituted formulations showed an increase in unknown RRT=0.1 (≦12.5%), KZR-59587 (≦16.1%), unknown RRT=0.5 (≦1.4%), KZR-0214143 (≦1.9%), and unknown RRT=2.2 (≦2.4%), as seen in Figure 1C, compared to those stored at 5°C. After storage at 25°C and 40°C, formulations F2 (unbuffered formulation containing 0.4% mannitol and 1% trehalose at pH 4.2) and F5 (unbuffered formulation containing 2% trehalose at pH 4.2) exhibited the highest main peak ratios among all candidate formulations. This data suggested that KZR-616 was most stable in the unbuffered formulation at pH 4.2 in the presence of sugars (Table 27). This is further supported by the trend for formulations F3, F6, and F8 without sugar to show the poorest stability results. The RP-HPLC data results support formulation F5 as the lead formulation and best candidate formulation for KZR-616. [Table 25] [Table 26] [Table 27]

[0078] After 6 months of incubation at 5°C, all formulations showed slight changes in the percentage of the main peak. However, all formulations still retained a main peak ratio of ≧97.9%. After 6 months of incubation at 25°C, all formulations retained a main peak ratio of ≧90.7%. Formulations F2 and F5 showed the highest main peak ratio and the slowest degradation rate. After 6 months of incubation at 40°C, all formulations except F8 retained a main peak ratio of ≧70.0%. Formulation F5 showed the highest main peak ratio and the slowest degradation rate (80.3%).

[0079] Moisture Optimization The objective of this study was to determine whether a modified lyophilization cycle could further improve the stability of KZR-616 during refrigeration. This lyophilization cycle optimization study focused on the lead formulation (2% trehalose, pH 4.2) from the previous study. Lead formulation F5 was prepared using drug substance from lot A (C15072369-FF16001) and lot B (1605R110). To achieve optimal moisture content, various drug substance lots were lyophilized using different secondary drying temperatures (Table 5).

[0080] Visual Analysis At time zero, all pre-lyophilization samples were visually clear, free of particles, and showed a yellowish color. The fresher drug substance (Lot B) showed a lighter color compared to the drug substance (Lot A). After lyophilization, all samples showed an elegant cake. After 4 weeks at 40°C, samples showed transparency and color equivalent to time zero.

[0081] pH Measurement All reconstituted samples at T=4 weeks showed comparable pH values ​​(Table 28). A slight increase was observed (≦±0.19) compared to time zero. [Table 28]

[0082] Concentration measurements After 4 weeks of incubation at 40°C, all samples showed similar concentrations compared to time zero (Ta [Table 29]

[0083] Reconstruction Time Measurements Following lyophilization at various secondary drying temperatures, the formulations were reconstituted with 1 milliliter of filtered MQ. Upon addition of the MQ, a timer was started to measure the time required for each formulation to homogenize. After 4 weeks of incubation at 40° C., all samples showed reconstitution times of <4 minutes, with 35A showing the shortest reconstitution time of 2 minutes 15 seconds and sample 10A showing the longest reconstitution time of 3 minutes 35 seconds (Table 30). [Table 30]

[0084] Moisture content measurement (Karl Fischer) After 6 months of incubation, the formulations showed moisture content values ​​comparable to time zero (Table 31). All lyophilized formulations continued to show moisture content values ​​below 1%. Differences in secondary drying temperatures showed little difference in moisture content, with higher temperatures showing the lowest moisture content. [Table 31]

[0085] RP-HPLC analysis After 4 weeks of incubation at 40°C, all reconstituted formulations showed a decrease in the percentage of the main peak (90.3%-91.7%) compared to that observed at time zero (97.7%-98.6%). RP-HPLC identified the major degradant species of the KZR-616 molecule. Peaks RRT=0.1 displayed KZR-59587, KZR-0214143, and RRT=2.2 showed the greatest increase in peak percentage when stressed. Differences in secondary drying temperatures showed little difference in the stability of the main peak at time zero. After 4 weeks at 40°C, all samples showed comparable stability.

[0086] The ratio of the main peak was significantly decreased in the samples from Lot B compared to the corresponding samples from Lot A, with the ratio of the main peak being 90.3%, due to the significant increase in KZR-0214142 observed in addition to the degradant peaks not observed in the samples from Lot A. [Table 32] [Table 33]

[0087] RP-HPLC showed an increase in degradant species after 4 weeks at 40° C. Lot B showed an additional peak (approximately 10 minutes) that was not present in the Lot A samples.

[0088] The stability of KZR-616 under various formulation conditions was investigated in this lyophilized formulation development study. The conditions investigated in this study included buffered (10 mM succinate) and non-buffered formulations with various bulking agents (L-glycine, mannitol, and trehalose) at various pH values ​​(3.9-4.2). The stability of KZR-616 in the different formulations was investigated under storage conditions up to 6 months at refrigerated (5°C), ambient (25°C), and accelerated (40°C) temperatures and analyzed by reversed-phase HPLC (RP-HPLC) to evaluate the stability of KZR-616.

[0089] Two rounds of lyophilization cycle development were performed before the accelerated stability study was initiated. Both development cycles showed elegant cakes and samples after reconstitution that were visually particle-free and slightly yellow in color. Lyophilization cycle #1 showed good stability by RP-HPLC. However, after measuring pH and osmolality, the results showed off-target pH and osmolality values. Lyophilization cycle #2 was performed with a formulation containing optimized bulking agents and achieved the target pH and osmolality. Additionally, cycle #2 reduced the fill step, increased the primary drying shelf temperature, and decreased the secondary drying shelf temperature. Samples produced in cycle #2 showed elegant cakes and good stability by RP-HPLC.

[0090] When stored for 6 months or longer, the lyophilized formulations consistently exhibited an elegant cake at all storage temperatures. However, the lyophilizates showed an increase in yellow coloration, especially when stored at 25°C and 40°C, which became even more evident after reconstitution. After reconstitution, all formulations were free of particles. No significant changes in pH or concentration were observed over time. Reconstitution times were consistently less than 4 minutes 30 seconds for all formulations over 6 months of storage, regardless of storage temperature.

[0091] Chemical stability evaluation of all formulations at all storage temperatures was performed using RP-HPLC. After 6 months of storage at 5°C, the formulations showed a similar main peak ratio (98.2%-98.4%) as at time zero and maintained a main peak ratio of ≦97.9%. After 6 months of storage at 25°C, the formulations showed a decreased main peak ratio (98.2%-98.4%) as compared to time zero and maintained a main peak (KZR-616) ratio (90.7%-93.4%). Trehalose-containing formulations F1, F2, and F5 showed the highest main peak ratios (92.8%, 93.4%, and 93.1%) after 6 months at 25°C. After storage at 40°C for 6 months, the formulations showed a significantly decreased proportion of the main peak (98.2%-98.4%) compared with time zero and maintained the proportion of the main peak (KZR-616) (63.8%-80.3%).

[0092] An additional moisture optimization study was performed on lead formulation F5 to optimize the lyophilization cycle, reduce moisture content, and significantly improve the stability of KZR-616. The moisture optimization study was performed with lyophilization cycles at various secondary shelf temperatures of 10°C, 25°C, 35°C, and 50°C.

[0093] After 4 weeks of storage at 40°C, all samples from lot C15072369-FF16001 showed no observed differences in visual appearance, pH, concentration, reconstitution time, and percentage of peaks by RP-HPLC, regardless of secondary drying shelf temperature. After 4 weeks of incubation at 40°C, all reconstituted formulations from lot C15072369-FF16001 showed a decrease in the percentage of the main peak (91.5%-91.7%) compared to that observed at time zero (97.7%-98.6%). Utilizing RP-HPLC, the main degradant species of the KZR-616 molecule were identified. Peaks RRT=0.1 displayed KZR-59587, KZR-0214143, and RRT=2.2 showed the greatest increase in peak percentage when stressed. Samples from lot 1605R110 showed a significant decrease in the main peak ratio compared to the corresponding samples from lot C15072369-FF16001, with the main peak ratio being 90.3%. This was due to a significant increase in KZR-0214142 observed in addition to the degradant peaks observed in samples from lot C15072369-FF16001. This study yielded comparable KZR-616 stability after 4 weeks, confirming low moisture content values ​​with no significant moisture difference between lyophilization cycles. Differences in secondary drying temperatures showed little difference in the main peak stability at time zero. After 4 weeks at 40°C, all formulated samples showed comparable stability, indicating that further drying of the cake did not achieve improved stability.

[0094] In conclusion, the trehalose-containing formulations, F2, and F5, showed the highest main peak ratios (78.7%, 80.3%) after 6 months at 40°C. The data suggest that KZR-616 is most stable in the unbuffered formulations at pH 4.2 in the presence of sugar. This is further supported by the trend for the sugar-free formulations, F3, F6, and F8, to show the poorest stability results. EXAMPLES

[0095] Example 2 Tables 34-36 show a comparison of the pharmacokinetics ("PK") and toxicokinetics ("TK") observed with different KZR-616 formulations in various animals. The test formulations contain various concentrations of KZR-616 as described in 2% trehalose by weight at pH 4.2 when dissolved in water for injection. The comparative formulations contain various concentrations of KZR-616 as described in 10% aqueous polysorbate-80 ("PS-80") by weight.

[0096] Table 34 shows that the test formulations exhibited faster absorption and elimination, and 2-3 times higher C max Similar AUCs were observed for both formulations. [Table 34]

[0097] Table 35 shows faster absorption and elimination, and 1.2 to 3.7 times higher C max was observed for the test formulation. A 0.7- to 1.7-fold difference in AUC was observed between the two formulations. [Table 35]

[0098] The test formulations in Table 36 were prepared as water for injection ("WFI") formulations as described above, and the comparative formulations were prepared as aqueous 10 wt% polysorbate-80 ("PS-80") formulations. Faster absorption and elimination, and 1.1-2.4 times higher C max was observed for the test formulation. A 0.6- to 1.0-fold difference in AUC was observed between the two formulations. [Table 36]

Claims

1. A pharmaceutical formulation comprising KZR-616 maleate and trehalose, KZR-616 is the structure 【Chemistry 1】 wherein the formulation has a reconstitution time of less than 4.5 minutes after storage at 5° C., 25° C. or 40° C. for at least 6 months, and wherein the formulation does not contain an added buffer and is a lyophilized formulation having a pH measured prior to lyophilization or upon reconstitution of between 3.8 and 4.

3.

2. 2. The formulation of claim 1 having a pH of 4.

2.

3. 3. The formulation of claim 2, wherein the KZR-616 maleate is present in an amount of from 100 mg / mL to 200 mg / mL as measured prior to lyophilization.

4. 4. The formulation of claim 3, wherein the KZR-616 maleate salt is present in an amount of 125 mg / mL based on the weight of the KZR-616 free base.

5. The formulation of any one of claims 1 to 4, wherein the trehalose is present in an amount of about 1.5% to 3% by weight.

6. A formulation according to any one of claims 1 to 5, wherein the trehalose is present in an amount of 2% by weight.

7. The formulation of any one of claims 1 to 6, wherein the lyophilized formulation has a moisture content of less than 1%.

8. 8. The formulation of any one of claims 1 to 7, having a concentration of KZR-616 maleate of 140 mg / mL to 150 mg / mL (measured upon reconstitution) after storage at 5°C, 25°C or 40°C for at least 6 months.

9. 9. The formulation of any one of claims 1 to 8, wherein the lyophilized formulation has a moisture content of less than 1% after storage at 5°C, 25°C, or 40°C for at least 6 months.

10. 10. The formulation of any one of claims 1 to 9, wherein the KZR-616 maleate salt is at least 93% pure after storage at 5°C, 25°C, or 40°C for at least 6 months.

11. A reconstituted formulation comprising the formulation of any one of claims 1 to 10 and a solvent for reconstitution.

12. 12. The reconstituted formulation of claim 11, wherein the solvent for reconstitution comprises water for injection (WFI) or 5% aqueous dextrose.

13. 13. The reconstituted formulation of claim 11 or 12, having an osmolality of 275 to 325 mOsm.

14. The reconstituted formulation according to any one of claims 11 to 13, which is for subcutaneous injection.

15. T １／２ 15. The reconstituted formulation of any one of claims 11 to 14, having an absorption rate at least 50% faster than that of a formulation comprising KZR-616 maleate and polysorbate-80, as measured by:

16. A C of at least 150% that of a formulation containing KZR-616 maleate and polysorbate-80 (the "PS-80 formulation"). ｍａｘ The reconstituted formulation of any one of claims 11 to 15, having the formula:

17. 11. A method of preparing the formulation of any one of claims 1 to 10, comprising mixing KZR-616 maleate and the trehalose to form an aqueous solution, and lyophilizing the aqueous solution to form the lyophilized formulation.

18. 20. The method of claim 17, further comprising reconstituting the lyophilized formulation with a solvent to form a reconstituted formulation.

19. 19. The method of claim 18, wherein the solvent is water for injection or 5% aqueous dextrose.

20. A therapeutic agent for immune-related diseases, comprising as an active ingredient the reconstituted preparation according to any one of claims 11 to 16.

21. The therapeutic agent for an immune-related disease according to claim 20, wherein the immune-related disease is lupus nephritis or systemic lupus erythematosus (SLE).

22. An inflammation treatment agent comprising the reconstituted preparation according to any one of claims 11 to 16 as an active ingredient.

23. Use of a reconstituted formulation according to any one of claims 11 to 16 in the manufacture of a medicament for treating an immune-related disease.

24. Use of a reconstituted formulation according to any one of claims 11 to 16 in the manufacture of a medicament for treating inflammation.

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