Low-dose IL-6 preparations and methods of use thereof

Abstract

The present disclosure provides an IL-6 formulation containing about 40 μg / mL to about 2 mg / mL of IL-6, about 5 to about 15 mM histidine, about 5 to about 15 mg / mL of glycine, about 40 to about 60 mg / mL of trehalose, about 0.1 to about 0.3 mg / mL of polysorbate 20, and about 1 to about 1000 μM of DTPA, wherein the formulation has a pH of about 6.0 to about 8.0. Such formulations may be useful for treating and / or preventing diseases or disorders in subjects in need thereof.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Patent Application No. 63 / 377,971, filed September 30, 2022, which is incorporated herein by reference in its entirety.

[0002] The present disclosure generally provides IL-6 formulations, such as stable low-dose formulations that are suitable for both frozen liquid and lyophilized storage, and methods of using the formulations, such as for treating IL-6-associated diseases and / or disorders. Sequence Listing This application contains a Sequence Listing that has been submitted electronically in XML format, which is incorporated by reference in its entirety. The XML file has the file name 116076_5019_PR_Sequence_Listing.SIP, was created on September 29, 2023, and is 4000 bytes in size. [Background technology]

[0003] Interleukin-6 (IL-6) (also known as interferon-β2, B-cell differentiation factor, B-cell stimulating factor-2, hepatocyte stimulating factor, hybridoma growth factor, and plasmacytoma growth factor) is a member of a family of cytokines that promotes cellular responses through a receptor complex consisting of the signaling glycoprotein gp130 and at least one subunit of the IL-6 receptor (IL-6R, also called gp80). IL-6R can also exist in a soluble form (sIL-6R). IL-6 binds to IL-6R and dimerizes the signaling receptor gp130. See Jones, SA, J. Immunology, 175:3463-3468 (2005).

[0004] Although IL-6 cannot directly bind to gp130, it can bind to IL-6R to form a high-affinity ternary complex of IL-6 / IL-6R / gp130. Although IL-6 binds to IL-6R with low affinity, this ligation alone does not result in cell activation because IL-6R lacks an intracellular signaling domain. Similarly, cell surface expression of IL-6R does not imply that the cell will respond to IL-6 stimulation. Proteolytic cleavage results in the release of soluble IL-6R (sIL-6R; sgp80), which allows circulating IL-6 to bind and extends its half-life. For cell activation, IL-6 first binds to either cell-bound IL-6R or sIL-6R, and the heterodimeric IL-6 / IL-6R complex associates with the cell surface glycoprotein gp130. The resulting ternary heterocomplex binds to another IL-6 / IL-6R / gp130, and signal transduction occurs (Bravo and Heath 2000, EMBO J., 19, (11), 2399-2411; Boulanger et al., 2003, Science, 300, 5628, 2101-2104). Therefore, both cell-bound and soluble IL-6R contribute to cell activation. IL-6 signaling via cell-bound IL-6R is referred to as cis signaling, while cell activation via soluble IL-6R is referred to as trans signaling.

[0005] The therapeutic use of IL-6 would be facilitated by a formulation that maintains the stability of IL-6 under various conditions. Important factors for a therapeutic formulation include the ability to be stored without unacceptable loss of activity of the active protein, minimizing the accumulation of undesirable products such as aggregates or degraded species (e.g., fragmented, oxidized, deamidated, or isomerized species), allowing for an appropriate concentration of protein, and being free of components incompatible with therapeutic use. Furthermore, conventional formulations of IL-6 use chemicals that are no longer recommended by regulatory authorities for human use, such as those containing sucrose, or use chemicals that are unstable in both liquid and lyophilized forms. Therefore, there is a need in the art for stable aqueous pharmaceutical formulations containing IL-6 that are suitable for therapeutic use. Summary of the Invention

[0006] The present disclosure generally provides unique IL-6 formulations that are stable in both liquid and lyophilized form and / or do not contain sucrose. The formulations contain about 40 μg / mL to about 2 mg / mL (e.g., 80 to 240 μg / mL) IL-6, about 5 to about 15 mM histidine, about 5 to about 15 mg / mL glycine, about 40 to about 60 mg / mL trehalose, about 0.1 to about 0.3 mg / mL polysorbate 20, and about 1 to about 1000 μM DTPA (e.g., 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95). The composition may contain 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, 425, 450, 475, 500, 525, 550, 575, 600, 625, 650, 675, 700, 725, 750, 775, 800, 825, 850, 875, 900, 925, 950, 975, or 1000 μM) and have a pH of about 6.0 to about 8.0.

[0007] In some embodiments, the IL-6 is human IL-6.

[0008] In some embodiments, the IL-6 comprises the sequence shown in SEQ ID NO:1 or SEQ ID NO:2.

[0009] In some embodiments, the formulation contains 40 μg / mL IL-6, 45 μg / mL IL-6, 50 μg / mL IL-6, 55 μg / mL IL-6, 60 μg / mL IL-6, 65 μg / mL IL-6, 70 μg / mL IL-6, 75 μg / mL IL-6, 80 μg / mL IL-6, 85 μg / mL IL-6, 90 μg / mL IL-6, 95 μg / mL IL-6, 100 μg / mL IL-6, 105 μg / mL IL-6, 110 μg / mL IL-6, 115 μg / mL IL-6, 120 μg / mL IL-6, 125 μg / mL IL-6, 130 μg / mL IL-6, 135 μg / mL IL-6, 140 μg / mL IL-6, 145μg / mL IL-6, 150μg / mL IL-6, 155μg / mL IL-6, 160μg / mL IL-6, 165μg / mL IL-6, 170μg / mL IL-6, 175μg / mL IL-6, 180μg / mL IL-6, 185μg / mL IL-6, 190μg / mL IL-6, In other embodiments, the formulation contains from about 80 μg / mL to about 2 mg / mL of IL-6.

[0010] In some embodiments, the formulation comprises 40 ug / ml of IL-6.

[0011] In some embodiments, the formulation comprises 80 μg / mL of IL-6.

[0012] In some embodiments, the formulation comprises 160 μg / mL of IL-6.

[0013] In some embodiments, the formulation comprises 240 μg / mL of IL-6.

[0014] In some embodiments, the formulation comprises about 10 mM histidine.

[0015] In some embodiments, the formulation comprises about 10 mg / mL of glycine.

[0016] In some embodiments, the formulation comprises about 50 mg / mL of trehalose.

[0017] In some embodiments, the formulation comprises about 0.2 mg / mL of polysorbate 20.

[0018] In some embodiments, the formulation comprises about 50 μM DTPA.

[0019] In some embodiments, the formulation has a pH of about 7.0.

[0020] In some embodiments, the formulation is a liquid formulation.

[0021] In some embodiments, the formulation is a lyophilized formulation.

[0022] The present disclosure also provides an IL-6 formulation containing 80-240 μg / mL human IL-6, 10 mM histidine, 10 mg / mL glycine, 50 mg / mL trehalose, 0.2 mg / mL polysorbate 20, and 50 μM DTPA, wherein the formulation has a pH of about 7.0.

[0023] In some embodiments, the formulation contains 40 μg / mL IL-6, 45 μg / mL IL-6, 50 μg / mL IL-6, 55 μg / mL IL-6, 60 μg / mL IL-6, 65 μg / mL IL-6, 70 μg / mL IL-6, 75 μg / mL IL-6, 80 μg / mL IL-6, 85 μg / mL IL-6, 90 μg / mL IL-6, 95 μg / mL IL-6, 100 μg / mL IL-6, 105 μg / mL IL-6, 110 μg / mL IL-6, 115 μg / mL IL-6, 120 μg / mL IL-6, 125 μg / mL IL-6, 130 μg / mL IL-6, 135 μg / mL IL-6, 140 μg / mL IL-6, 145μg / mL IL-6, 150μg / mL IL-6, 155μg / mL IL-6, 160μg / mL IL-6, 165μg / mL IL-6, 170μg / mL IL-6, 175μg / mL IL-6, 180μg / mL IL-6, 185μg / mL IL-6, 190μg / mL IL-6, In other embodiments, the formulation contains from about 40 μg / mL to about 2 mg / mL of IL-6.

[0024] In some embodiments, the formulation comprises 40 μg / mL of IL-6.

[0025] In some embodiments, the formulation comprises 80 μg / mL of IL-6.

[0026] In some embodiments, the formulation comprises 160 μg / mL of IL-6.

[0027] In some embodiments, the formulation comprises 240 μg / mL of IL-6.

[0028] In some embodiments, the formulation is suitable for parenteral administration.

[0029] In some embodiments, the formulation is suitable for intravenous administration.

[0030] In some embodiments, the formulation is suitable for subcutaneous administration.

[0031] The present disclosure also provides methods of treating an IL-6-related disease or disorder in a subject in need thereof by administering to the subject the formulations disclosed herein. DETAILED DESCRIPTION OF THE INVENTION

[0032] The present disclosure provides stable, low-dose IL-6 (e.g., h-rIL-6) formulations suitable for both frozen liquid and lyophilized storage, as well as methods of using the formulations, such as for treating IL-6-related diseases and / or disorders. Advantageously, the formulated IL-6 needs remain stable (e.g., does not deteriorate over time at various temperatures, tolerates multiple freeze-thaw cycles, and is readily available for injection upon thawing or reconstitution with water for injection), allowing use in diabetic patients who limit sugar consumption in their diet.

[0033] 1.1 Definition The following definitions are provided to provide a clear and consistent understanding of the specification and claims. As used herein, the described terms have the following meanings. All other terms and phrases used herein have their ordinary meanings as understood by those skilled in the art. Such ordinary meanings can be obtained by consulting a technical dictionary such as Hawley's Condensed Chemical Dictionary 14th Edition (R.J. Lewis, John Wiley & Sons, New York, NY, 2001).

[0034] References herein to "one embodiment," "an embodiment," and the like indicate that the described embodiment may include a particular aspect, configuration, structure, moiety, or characteristic, but not all embodiments necessarily include that aspect, configuration, structure, moiety, or characteristic. Moreover, such phrases may, but do not necessarily, refer to the same embodiment that is also referenced elsewhere in this specification. Furthermore, when a particular aspect, configuration, structure, moiety, or characteristic is described in connection with an embodiment, it is within the knowledge of one of ordinary skill in the art to modify or combine such aspect, configuration, structure, moiety, or characteristic with other embodiments, whether or not explicitly described.

[0035] Singular forms such as "a," "an," and "the" are intended to include plural references unless the context clearly dictates otherwise. Thus, for example, reference to a formulation with "an" also includes a plurality of such formulations, where formulation X contains a plurality of formulations X. It is further noted that the claims may be worded in a manner that excludes any optional element. As such, this definition is intended to serve as antecedent (defining prior) to the use of exclusive terms such as "solely," "only," and the like in connection with the description of any element described herein and / or claim element, or the use of a "negative" limitation.

[0036] The term "and / or" means any one thereof, any combination thereof, or all of those with which the term is associated. The phrases "one or more" and "at least one" will be readily understood by those of ordinary skill in the art, particularly when read in the context of their use. For example, the phrases can mean 1, 2, 3, 4, 5, 6, 10, 100, or any upper limit.

[0037] A "stable" IL-6 formulation is one in which IL-6 substantially retains its physical and / or chemical stability and / or its biological activity upon storage. The storage period is generally selected based on the intended shelf life of the formulation evaluated at various temperatures (-80°C to +40°C). Various analytical techniques for measuring protein stability are available in the art. Examples of analytical techniques are described below. By "substantially retained" is intended 85% or greater retention, such as at least 90% retention or at least 95% retention, depending on the analytical technique and specifications applied.

[0038] An IL-6 protein is said to "retain physical stability" in a pharmaceutical formulation if it does not exhibit significant physical changes, such as aggregation, precipitation, and / or denaturation, upon visual inspection of color and / or clarity or as measured by UV light scattering or size exclusion chromatography.

[0039] An IL-6 protein is said to "retain chemical stability" in a pharmaceutical formulation if the protein does not exhibit significant chemical changes. Chemical stability can be evaluated by detecting and quantifying chemically modified forms of the protein. Chemical changes may include, for example, size modifications (e.g., clipping), which can be evaluated using size exclusion chromatography, SDS-PAGE, and / or matrix-assisted laser desorption / ionization / time-of-flight mass spectrometry (MALDI / TOF MS). Other types of chemical changes include charge changes (e.g., resulting from deamidation, oxidation, and / or isomerization), which can be evaluated, for example, by ion exchange chromatography.

[0040] An IL-6 protein is said to "retain its biological activity" in a pharmaceutical formulation if the biological activity of the IL-6 protein at a given time does not change significantly from the biological activity exhibited at the time the pharmaceutical formulation was prepared. The "biological activity" of an IL-6 protein refers to the ability of the protein to produce a measurable biological response that can be measured in vitro or in vivo.

[0041] A "histidine buffer" is a buffer containing histidine ions. Examples of histidine buffers include solutions of histidine chloride, histidine acetate, histidine phosphate, and histidine sulfate. Histidine buffers or histidine-HCl buffers have a pH of about 5.5 to about 6.5, about 5.6 to about 6.4, about 5.7 to about 6.3, about 5.8 to about 6.2, about 5.9 to about 6.1, about 6.0 to about 7.0, about 6.5 to about 7.0, about 7.0 to about 7.5, or about 7.0 to about 8.0.

[0042] "IL-6" or "interleukin 6 (IL-6)" or "IL-6 polypeptide" refers to a polypeptide or fragment thereof having at least about 85% or more amino acid identity with the amino acid sequence provided in NCBI Accession No. NP_000591 and having IL-6 biological activity. IL-6 is a multifunctional cytokine with multiple biological functions. Exemplary IL-6 biological activities include immunostimulatory activity and pro-inflammatory activity. "IL-6" or "interleukin 6 (IL-6) nucleic acid" refers to a polynucleotide encoding an interleukin 6 (IL-6) polypeptide. An exemplary interleukin 6 (IL-6) nucleic acid sequence is provided in NCBI Accession No. NM_000600.

[0043] The "identity" percentage between any polypeptide sequence and any reference sequence is defined as the percentage of amino acid residues in the polypeptide sequence that are identical to the amino acid residues in the reference sequence after aligning the sequences and introducing gaps as necessary to achieve the maximum sequence identity percentage.Alignment for determining amino acid sequence identity percentage can be achieved in various ways within the skill of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, MEGALIGN (DNASTAR), CLUSTALW, CLUSTAL OMEGA, or MUSCLE software.Those skilled in the art can determine the appropriate parameters for sequence alignment, such as any algorithm required to achieve maximum alignment over the entire length of the sequences being compared.Unless otherwise specified, sequence identity percentage is determined using the BLAST algorithm with default parameters.

[0044] "Subject" means a human or non-human mammalian subject, including, but not limited to, bovine, equine, canine, equine, feline, and rodent animals, including mice and rats. A "patient" is a human subject.

[0045] As used herein, the terms "treat," "treating," "treatment," and the like refer to reducing or ameliorating a disorder and / or its associated signs or symptoms, or slowing or halting its progression. It will be understood that treating a disorder or condition does not necessarily mean completely eliminating the disorder, condition, or symptoms associated therewith, although it does not eliminate them.

[0046] In this disclosure, the words "comprises," "comprising," "containing," "having," "includes," "including," and linguistic variations thereof have the meaning ascribed to them in U.S. patent law and permit the presence of additional elements other than those expressly recited.

[0047] As will be understood by those of ordinary skill in the art, all numbers, including those expressing quantities of ingredients and properties such as molecular weight, reaction conditions, and the like, are approximate and will be understood in all instances as optionally modified by the term "about." These values ​​may vary depending on the desired properties one of ordinary skill in the art seeks to obtain using the teachings set forth herein. It will also be understood that such values ​​inherently contain variability necessarily resulting from the standard deviation found in their respective testing measurements. Even when values ​​are expressed as approximations by use of "about" as an antecedent, it will be understood that any value without the modifier "about" can form a further aspect.

[0048] The terms "about" and "approximately" can be used interchangeably. Both terms can refer to a variation of ±1%, ±5%, or ±10% of the specified value. For example, in some embodiments, "about 50" percent can have a variation of 45 to 55 percent, or as otherwise defined in a particular claim. For integer ranges, the term "about" can include one or two integers greater than and / or less than the integers recited as the upper and lower limits of the range. Unless otherwise indicated herein, the terms "about" and "approximately" are intended to include values, such as weight percent, that are close to the stated range and are equivalent with respect to the functionality of the individual component, composition, or embodiment. Additionally, the terms "about" and "approximately" can modify the upper and lower limits of the stated range, as described above in this paragraph.

[0049] As will be understood by those skilled in the art, for all purposes, and particularly where a clear description is provided, all ranges described herein encompass subranges and combinations of subranges, as well as the individual values, particularly integers, that make up the range. Thus, it is understood that every value between two specific values ​​is also disclosed. For example, if 10 to 15 is disclosed, 11, 12, 13, and 14 are also disclosed individually and as part of a range. Any range described (e.g., weight percent or carbon group) includes each specific value, integer, decimal, or identity within the range. Any recited range will be readily recognized as fully describing and allowing for the same range to be divided into at least two, three, four, five, or ten equal parts. By way of non-limiting example, each range described herein can be readily broken down into a lower third, a middle third, an upper third, etc. Also, as will be understood by those skilled in the art, all terms such as "at most," "at least," "greater than," "less than one," "greater than," "greater than or equal to," etc., are inclusive of the recited numbers, and such terms refer to ranges that can be broken down into subranges as described above. Similarly, all ratios recited herein also encompass all subratios that fall within the broader ratio. Thus, specific values ​​recited for radicals, substituents, and ranges are for illustrative purposes only and do not exclude other defined or other values ​​within the defined ranges for radicals and substituents. It will be further understood that the upper and lower limits of each range are significant both in relation to its opposite limit and independently of its opposite limit.

[0050] The present disclosure provides ranges, upper limits, and deviations for variables such as volume, mass, percentages, and ratios. Those skilled in the art will understand that a range, such as "number 1" to "number 2," refers to a continuous range of numbers, including integers and fractions. For example, 1 to 10 refers to 1, 2, 3, 4, 5, ..., 9, 10. It also refers to 1.0, 1.1, 1.2, 1.3, ..., 9.8, 9.9, 10.0, etc., as well as 1.01, 1.02, 1.03, etc. When a disclosed variable is a number less than "the number 10," this refers to a continuous range, including integers and fractions less than the number 10, as described above.

[0051] Similarly, when a disclosed variable is a number greater than "the number 10," a continuous range including integers and fractions greater than the number 10 is meant.

[0052] These ranges may be modified by the term "about," the meaning of which is explained above.

[0053] Additionally, those skilled in the art will readily recognize that when multiple entities are grouped together in a common manner, such as a Markush group, the invention encompasses not only the entire group as recited as a whole, but also the individual members comprising the group and all possible subgroups of the main group. Furthermore, for all purposes, the invention encompasses not only the main group, but also those main groups in which one or more group members are absent. Thus, the invention contemplates configurations in which any one or more of the recited group members is explicitly excluded. Thus, while this may apply to any of the disclosed categories or embodiments, any one or more of the described elements, species, or embodiments may be excluded from such categories or embodiments, such as when used in an explicit negative limitation.

[0054] 1.2. Formulation In a first aspect, provided herein is an aqueous or lyophilized formulation (pharmaceutical composition) comprising IL-6 protein. In one embodiment, the formulation is sterile. In other embodiments, the formulation is stable. In some embodiments, the formulation further comprises at least one free amino acid. In some embodiments, the formulation further comprises at least one surfactant. In some embodiments, the formulation further comprises at least one buffer component. In some embodiments, the formulation further comprises at least one chelating agent. In some embodiments, the formulation comprises at least one saccharide. In some embodiments, the formulation comprises IL-6 (e.g., human IL-6). In certain embodiments, the formulation comprises at least one free amino acid, at least one surfactant, at least one buffer component, at least one chelating agent, at least one saccharide, and IL-6.

[0055] In some embodiments, the formulation contains about 40 to about 240 μg / mL of IL-6, about 5 to about 15 mM of histidine, about 5 to about 15 mg / mL of glycine, about 40 to about 60 mg / mL of trehalose, about 0.1 to about 0.3 mg / mL of polysorbate 20, and about 5 to about 100 μM of DTPA, and has a pH of about 6.0 to about 8.0. In other embodiments, the formulation contains 40 to about 240 μg / mL of human IL-6, 10 mM of histidine, 10 mg / mL of glycine, 50 mg / mL of trehalose, 0.2 mg / mL of polysorbate 20, and 50 μM of DTPA, and has a pH of about 7.0.

[0056] 1.2.1. IL-6 concentration In some embodiments, the concentration of IL-6 in the formulation is at least about 80 μg / mL to about 2 mg / mL. In some embodiments, the IL-6 concentration in the formulation is at least about 40 μg / mL, at least about 45 μg / mL, at least about 50 μg / mL, at least about 55 μg / mL, at least about 60 μg / mL, at least about 65 μg / mL, at least about 70 μg / mL, at least about 75 μg / mL, at least about 80 μg / mL, at least about 85 μg / mL, at least about 90 μg / mL, at least about 95 μg / mL, at least about 100 μg / mL, at least about 105 μg / mL, at least about 110 μg / mL, at least about 115 μg / mL, at least about 120 μg / mL, at least about 125 μg / mL, at least about 130 μg / mL, at least about 135 μg / mL, at least about 140 μg / mL, at least about 145 μg / mL, at least about 150 μg / mL, at least about 155 μg / mL, at least about at least about 160 μg / mL, at least about 165 μg / mL, at least about 170 μg / mL, at least about 175 μg / mL, at least about 180 μg / mL, at least about 185 μg / mL, at least about 190 μg / mL, at least about 195 μg / mL, at least about 200 μg / mL, at least about 205 μg / mL, at least about 210 μg / mL, at least about 215 μg / mL, at least about 220 μg / mL mL, at least about 225 μg / mL, at least about 230 μg / mL, at least about 235 μg / mL, at least about 240 μg / mL, at least about 245 μg / mL, at least about 250 μg / mL, at least about 255 μg / mL, at least about 260 μg / mL, at least about 265 μg / mL, at least about 270 μg / mL, at least about 275 μg / mL, or at least about 280 μg / mL.

[0057] In some embodiments, the IL-6 concentration in the formulation is about 40 μg / mL to about 280 μg / mL, or in other embodiments, the IL-6 concentration in the formulation is about 100 to about 120 μg / mL, about 120 to about 140 μg / mL, about 140 to about 160 μg / mL, about 160 to about 180 μg / mL, about 180 to about 200 μg / mL, about 200 to about 220 μg / mL, about 220 to about 240 μg / mL, about 240 to about 260 μg / mL, or about 260 to about 280 μg / mL.

[0058] 1.2.2. Chelating Agents In some embodiments, the formulation comprises a chelating agent.

[0059] In various embodiments, the chelating agent is at a concentration of about 1 μM to about 1000 μM, e.g., about 25 μM to about 75 μM, preferably about 50 μM. In various embodiments, the chelating agent is at a concentration of 1 μM to 100 μM, e.g., 25 μM to 75 μM, preferably 50 μM.

[0060] In certain embodiments, the chelating agent is a naturally occurring compound. In certain other embodiments, the chelating agent is a synthetic compound. In some embodiments, the chelating agent is diethylenetriaminepentaacetic acid (DTPA).

[0061] 1.2.3. Surfactants In some embodiments, the formulation comprises a surfactant.

[0062] Surfactants can reduce the surface tension of liquids. In some embodiments, the surfactant is a non-ionic surfactant. Examples of surfactants include polysorbates (polyoxyethylene sorbitan monolaurates such as polysorbate 20 and polysorbate 80); TRITON (t-octylphenoxypolyethoxyethanol); sodium dodecyl sulfate (SDS); sodium laurel sulfate, sodium octyl glycoside, lauryl, myristyl, linoleyl, or stearyl sulfobetaine, lauryl, myristyl, linoleyl, or stearyl sarcosine; linoleyl, myristyl, or cetyl betaine; lauroamidopropyl, cocamidopropyl, linoleamidopropyl, myristamidopropyl, palmito ... Examples of surfactants include propyl- or isostearamidopropyl-betaine (e.g., lauramidopropyl); myristamidopropyl-, palmidopropyl-, or isostearamidopropyl-dimethylamine; sodium cocoyl methyl taurate or disodium oleyl methyl taurate; sorbitan monopalmitate; MONAQUAT series (Mona Instant, Inc., Paterson, New Jersey); polyethyl glycol (PEG), polypropylene glycol (PPG), and copolymers of poloxyethylene and poloxypropylene glycol (e.g., Pluronics / Poloxamer, PF68, etc.). In some of these embodiments, the surfactant is a polysorbate. In certain embodiments, the polysorbate is polysorbate 20, polysorbate 40, polysorbate 60, or polysorbate 80.

[0063] In various embodiments, the composition comprises a surfactant at a concentration of about 0.001% to about 1%, e.g., about 0.001% to about 0.1%, about 0.005% to about 0.2%, about 0.01% to about 0.2%, or about 0.05% to about 0.1% (w / v). In various embodiments, the composition comprises a surfactant at a concentration of 0.001% to 1%, e.g., 0.001% to 0.1%, 0.005% to 0.2%, 0.01% to 0.2%, or 0.05% to 0.1% (w / v).

[0064] 1.2.4. Polysorbates In some embodiments, the formulation comprises a polysorbate.

[0065] Polysorbate 80 In some embodiments, the formulation comprises polysorbate 80 (PS80).

[0066] In some embodiments, the formulation comprises at least about 0.001%, at least about 0.002%, at least about 0.003%, at least about 0.005%, at least about 0.01%, at least about 0.02%, at least about 0.05%, at least about 0.1%, at least about 0.2%, at least about 0.5%, or at least about 1% (w / v) polysorbate 80. In some embodiments, the formulation comprises between about 0.001% and about 1%, between about 0.002% and about 0.5%, between about 0.005% and about 0.2%, or between about 0.01% and about 0.1% (w / v) polysorbate 80. In some embodiments, the formulation comprises about 0.001%, about 0.002%, about 0.003%, about 0.005%, about 0.01%, about 0.02%, about 0.05%, about 0.07%, about 0.1%, about 0.2%, about 0.5%, or about 1% (w / v) polysorbate 80. In certain embodiments, the formulation comprises about 0.005% to about 0.5% (w / v) polysorbate 80. In certain embodiments, the formulation comprises about 0.03% to about 0.1% (w / v) polysorbate 80. In certain embodiments, the formulation comprises about 0.05% to about 0.1% (w / v) polysorbate 80. In certain embodiments, the formulation comprises about 0.07% (w / v) polysorbate 80.

[0067] In some embodiments, the formulation comprises at least 0.001%, at least 0.002%, at least 0.003%, at least 0.005%, at least 0.01%, at least 0.02%, at least 0.05%, at least 0.1%, at least 0.2%, at least 0.5%, or at least 1% (w / v) polysorbate 80. In some embodiments, the formulation comprises 0.001% to 1%, 0.002% to 0.5%, 0.005% to 0.2%, or 0.01% to 0.1% (w / v) polysorbate 80. In some embodiments, the formulation comprises 0.001%, 0.002%, 0.003%, 0.005%, 0.01%, 0.02%, 0.05%, 0.07%, 0.1%, 0.2%, 0.5%, or 1% (w / v) polysorbate 80. In certain embodiments, the formulation comprises 0.005% to 0.5% (w / v) polysorbate 80. In certain embodiments, the formulation comprises 0.03% to 0.1% (w / v) polysorbate 80. In certain embodiments, the formulation comprises 0.05% to 0.1% (w / v) polysorbate 80. In certain embodiments, the formulation comprises 0.07% (w / v) polysorbate 80.

[0068] Polysorbate 60 In some embodiments, the formulation comprises polysorbate 60 (PS60).

[0069] In some embodiments, the formulation comprises at least about 0.001%, at least about 0.002%, at least about 0.003%, at least about 0.005%, at least about 0.01%, at least about 0.02%, at least about 0.05%, at least about 0.1%, at least about 0.2%, at least about 0.5%, or at least about 1% (w / v) polysorbate 60. In some embodiments, the formulation comprises between about 0.001% and about 1%, between about 0.002% and about 0.5%, between about 0.005% and about 0.2%, or between about 0.01% and about 0.1% (w / v) polysorbate 60. In some embodiments, the formulation comprises about 0.001%, about 0.002%, about 0.003%, about 0.005%, about 0.01%, about 0.02%, about 0.05%, about 0.07%, about 0.1%, about 0.2%, about 0.5%, or about 1% (w / v) polysorbate 60. In certain embodiments, the formulation comprises about 0.03% to about 0.1% (w / v) polysorbate 60. In certain embodiments, the formulation comprises about 0.05% to about 0.1% (w / v) polysorbate 60. In certain embodiments, the formulation comprises about 0.07% (w / v) polysorbate 60.

[0070] In some embodiments, the formulation comprises at least 0.001%, at least 0.002%, at least 0.003%, at least 0.005%, at least 0.01%, at least 0.02%, at least 0.05%, at least 0.1%, at least 0.2%, at least 0.5%, or at least 1% (w / v) polysorbate 60. In some embodiments, the formulation comprises 0.001% to 1%, 0.002% to 0.5%, 0.005% to 0.2%, or 0.01% to 0.1% (w / v) polysorbate 60. In some embodiments, the formulation comprises 0.001%, 0.002%, 0.003%, 0.005%, 0.01%, 0.02%, 0.05%, 0.07%, 0.1%, 0.2%, 0.5%, or 1% (w / v) polysorbate 60. In certain embodiments, the formulation comprises 0.03%-0.1% (w / v) polysorbate 60. In certain embodiments, the formulation comprises 0.05%-0.1% (w / v) polysorbate 60. In certain embodiments, the formulation comprises 0.07% (w / v) polysorbate 60.

[0071] Polysorbate 40 In some embodiments, the formulation comprises polysorbate 40 (PS40).

[0072] In some embodiments, the formulation comprises at least about 0.001%, at least about 0.002%, at least about 0.003%, at least about 0.005%, at least about 0.01%, at least about 0.02%, at least about 0.05%, at least about 0.1%, at least about 0.2%, at least about 0.5%, or at least about 1% (w / v) polysorbate 40. In some embodiments, the formulation comprises between about 0.001% and about 1%, between about 0.002% and about 0.5%, between about 0.005% and about 0.2%, or between about 0.01% and about 0.1% (w / v) polysorbate 40. In some embodiments, the formulation comprises about 0.001%, about 0.002%, about 0.003%, about 0.005%, about 0.01%, about 0.02%, about 0.05%, about 0.07%, about 0.1%, about 0.2%, about 0.5%, or about 1% (w / v) polysorbate 40. In certain embodiments, the formulation comprises about 0.03% to about 0.1% (w / v) polysorbate 40. In certain embodiments, the formulation comprises about 0.05% to about 0.1% (w / v) polysorbate 40. In certain embodiments, the formulation comprises about 0.07% (w / v) polysorbate 40.

[0073] In some embodiments, the formulation comprises at least 0.001%, at least 0.002%, at least 0.003%, at least 0.005%, at least 0.01%, at least 0.02%, at least 0.05%, at least 0.1%, at least 0.2%, at least 0.5%, or at least 1% (w / v) polysorbate 40. In some embodiments, the formulation comprises 0.001% to 1%, 0.002% to 0.5%, 0.005% to 0.2%, or 0.01% to 0.1% (w / v) polysorbate 40. In some embodiments, the formulation comprises 0.001%, 0.002%, 0.003%, 0.005%, 0.01%, 0.02%, 0.05%, 0.07%, 0.1%, 0.2%, 0.5%, or 1% (w / v) polysorbate 40. In certain embodiments, the formulation comprises 0.03% to 0.1% (w / v) polysorbate 40. In certain embodiments, the formulation comprises 0.05% to 0.1% (w / v) polysorbate 40. In certain embodiments, the formulation comprises 0.07% (w / v) polysorbate 40.

[0074] Polysorbate 20 In some embodiments, the formulation comprises polysorbate 20 (PS20).

[0075] In some embodiments, the formulation comprises at least about 0.001%, at least about 0.002%, at least about 0.003%, at least about 0.005%, at least about 0.01%, at least about 0.02%, at least about 0.05%, at least about 0.1%, at least about 0.2%, at least about 0.5%, or at least about 1% (w / v) polysorbate 20. In some embodiments, the formulation comprises between about 0.001% and about 1%, between about 0.002% and about 0.5%, between about 0.005% and about 0.2%, or between about 0.01% and about 0.1% (w / v) polysorbate 20. In some embodiments, the formulation comprises about 0.001%, about 0.002%, about 0.003%, about 0.005%, about 0.01%, about 0.02%, about 0.05%, about 0.07%, about 0.1%, about 0.2%, about 0.5%, or about 1% (w / v) polysorbate 20. In certain embodiments, the formulation comprises about 0.03% to about 0.1% (w / v) polysorbate 20. In certain embodiments, the formulation comprises about 0.05% to about 0.1% (w / v) polysorbate 20. In certain embodiments, the formulation comprises about 0.07% (w / v) polysorbate 20.

[0076] In some embodiments, the formulation comprises at least 0.001%, at least 0.002%, at least 0.003%, at least 0.005%, at least 0.01%, at least 0.02%, at least 0.05%, at least 0.1%, at least 0.2%, at least 0.5%, or at least 1% (w / v) polysorbate 20. In some embodiments, the formulation comprises 0.001% to 1%, 0.002% to 0.5%, 0.005% to 0.2%, or 0.01% to 0.1% (w / v) polysorbate 20. In some embodiments, the formulation comprises 0.001%, 0.002%, 0.003%, 0.005%, 0.01%, 0.02%, 0.05%, 0.07%, 0.1%, 0.2%, 0.5%, or 1% (w / v) polysorbate 20. In certain embodiments, the formulation comprises 0.03% to 0.1% (w / v) polysorbate 20. In certain embodiments, the formulation comprises 0.05% to 0.1% (w / v) polysorbate 20. In certain embodiments, the formulation comprises 0.07% (w / v) polysorbate 20.

[0077] 1.2.5. Sugars

[0078] In some embodiments, the composition comprises sugars and their derivatives, such as monosaccharides, disaccharides, trisaccharides, polysaccharides, sugar alcohols, reducing sugars, and non-reducing sugars. Examples of sugars include glucose, mannose, trehalose, lactose, fructose, maltose, dextran, dextrin, erythritol, glycerol, arabitol, syritol, sorbitol, mannitol, melibiose, melezitose, raffinose, mannotriose, stachyose, maltose, lactulose, maltulose, glucitol, maltitol, lactitol, and isomaltulose. In some embodiments, the sugar is a disaccharide. In certain embodiments, the disaccharide is trehalose. In various embodiments, the disaccharide is at a concentration of about 1 to about 100 mg / mL.

[0079] 1.2.5.1. Trehalose In some embodiments, the formulation comprises trehalose.

[0080] In various embodiments, the formulation contains about 1 to about 100 mg / mL of trehalose. In some embodiments, the formulation contains at least about 25 mg / mL, about 30 mg / mL, about 35 mg / mL, about 40 mg / mL, about 45 mg / mL, about 50 mg / mL, about 55 mg / mL, about 60 mg / mL, about 65 mg / mL, about 70 mg / mL, or about 75 mg / mL of trehalose. In certain embodiments, the formulation contains about 50 mg / mL of trehalose.

[0081] Free amino acids In some embodiments, the formulation comprises at least one free amino acid. In some embodiments, the formulation comprises only one free amino acid. In some embodiments, the formulation comprises only two free amino acids. In some embodiments, the formulation comprises only three free amino acids. The free amino acids may be L-, D-, or a mixture thereof.

[0082] In certain embodiments, the at least one free amino acid is glycine, glutamine, asparagine, histidine, arginine, or lysine. In various embodiments, the free amino acid is at a concentration of about 1 mM to about 150 mM, e.g., about 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 11 mM, 12 mM, 13 mM, 15 mM, 16 mM, 17 mM, 18 mM, 19 mM, or 20 mM.

[0083] In some embodiments, the formulation comprises histidine and at least one additional free amino acid selected from glycine, glutamine, asparagine, histidine, arginine, and lysine. In certain embodiments, the formulation comprises histidine and glycine.

[0084] Glycine In some embodiments, the formulation comprises glycine. In various embodiments, the formulation comprises about 5 to about 15 mg / mL of glycine. In some embodiments, the formulation comprises at least about 5 mg / mL, at least about 10 mg / mL, or at least about 15 mg / mL of glycine. In various embodiments, the formulation comprises about 5 mg / mL to about 10 mg / mL, or about 10 mg / mL to about 20 mg / mL. In certain embodiments, the formulation comprises about 10 mg / mL of glycine.

[0085] 1.2.7. Buffers In some embodiments, the formulation comprises at least one buffering agent (buffering component). Typically, when present, the buffering agent is used to adjust the pH of the formulation to about 4.0 to about 8.0, about 4.5 to about 7.5, about 5.0 to about 7.0, about 5.5 to about 6.5, about 5.7 to about 6.3, about 5.9 to about 6.1, or about 6.0.

[0086] In various embodiments, at least one buffering agent is selected from acetate, succinate, gluconate, histidine, citrate, phosphate, maleate, cacodylate, 2-[N-morpholino]ethanesulfonic acid (MES), bis(2-hydroxyethyl)iminotris[hydroxymethyl]methane (bis-tris), N-[2-acetamido]-2-iminodiacetic acid (ADA), glycylglycine, and other organic acid buffers. In some of these embodiments, the buffering agent is histidine, citrate, phosphate, glycine, or acetate. In various embodiments, the buffering components are at a concentration of about 1 mM to about 200 mM, about 1 mM to about 50 mM, or about 5 mM to about 20 mM. In various embodiments, the buffering components are at a concentration of 1 mM to about 200 mM, about 1 mM to about 50 mM, or about 5 mM to about 20 mM. In certain embodiments, the buffer components are at a concentration of about 5 mM, about 10 mM, about 15 mM, about 20 mM, or about 25 mM. In certain embodiments, the buffer components are at a concentration of 10 mM.

[0087] In a preferred embodiment, the buffering agent is histidine. In some embodiments, the formulation comprises histidine. In some embodiments, the formulation comprises L-histidine.

[0088] In some embodiments, the composition comprises at least about 1 mM, at least about 5 mM, at least about 10 mM, at least about 15 mM, at least about 20 mM, at least about 30 mM, at least about 50 mM, at least about 100 mM, at least about 150 mM, or at least about 200 mM histidine. In various embodiments, the composition comprises about 1 mM to about 200 mM, about 5 mM to about 150 mM, about 10 mM to about 100 mM, about 15 mM to about 50 mM, or about 20 mM to about 30 mM histidine. In certain embodiments, the composition comprises about 1 mM, about 5 mM, about 10 mM, about 15 mM, about 20 mM, about 30 mM, about 50 mM, about 100 mM, about 150 mM, or about 200 mM histidine. In certain embodiments, the formulation comprises about 10 mM to about 100 mM histidine. In certain embodiments, the formulation comprises about 10 mM histidine.

[0089] In some embodiments, the composition comprises at least 1 mM, at least 5 mM, at least 10 mM, at least 15 mM, at least 20 mM, at least 30 mM, at least 50 mM, at least 100 mM, at least 150 mM, or at least 200 mM histidine. In various embodiments, the composition comprises 1 mM to 200 mM, 5 mM to 150 mM, 10 mM to 100 mM, 15 mM to 50 mM, or 20 mM to 30 mM histidine. In certain embodiments, the composition comprises 1 mM, 5 mM, 10 mM, 15 mM, 20 mM, 30 mM, 50 mM, 100 mM, 150 mM, or 200 mM histidine. In certain embodiments, the formulation comprises 10 mM to 100 mM histidine. In certain embodiments, the formulation comprises 10 mM histidine.

[0090] pH In some embodiments, the formulation has a pH of about 4.0 to about 8.0, e.g., about 4.5 to about 7.5, about 5.0 to about 7.0, about 5.5 to about 6.5, about 5.7 to about 6.3, or about 5.9 to about 6.1. In certain embodiments, the formulation has a pH of about 4.0, about 4.5, about 5.0, about 5.5, about 5.7, about 5.9, about 6.0, about 6.1, about 6.3, about 6.5, about 7.0, about 7.5, or about 8.0. In certain embodiments, the formulation has a pH of about 5.0 to about 7.0. In certain embodiments, the formulation has a pH of about 5.5 to about 6.5. In certain embodiments, the formulation has a pH of about 5.7 to about 6.3. In certain embodiments, the formulation has a pH of about 6.0.

[0091] In some embodiments, the formulation has a pH of 4.0 to 8.0, e.g., 4.5 to 7.5, 5.0 to 7.0, 5.5 to 6.5, 5.7 to 6.3, or 5.9 to 6.1. In certain embodiments, the formulation has a pH of 4.0, 4.5, 5.0, 5.5, 5.7, 5.9, 6.0, 6.1, 6.3, 6.5, 7.0, 7.5, or 8.0. In certain embodiments, the formulation has a pH of 5.0 to 7.0. In certain embodiments, the formulation has a pH of 5.5 to 6.5. In certain embodiments, the formulation has a pH of 5.7 to 6.3. In certain embodiments, the formulation has a pH of 6.0.

[0092] Preservatives In some embodiments, the formulation further comprises at least one preservative. In various embodiments, the at least one preservative is selected from octadecyldimethylbenzylammonium chloride, hexamethonium chloride, benzalkonium chloride (a mixture of alkylbenzyldimethylammonium chlorides in which the alkyl group is a long-chain compound), and benzethonium chloride. Other types of preservatives include aromatic alcohols such as phenol, butyl, and benzyl alcohol, alkyl parabens such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol, 3-pentanol, and m-cresol.

[0093] 1.2.9. Viscosity In some embodiments, the viscosity of the formulation is less than 50 cP at 25° C., e.g., less than 40 cP, less than 30 cP, less than 20 cP, less than 10 cP, or less than 5 cP at 25° C. In certain embodiments, the viscosity of the formulation is less than 10 cP at 25° C. In various embodiments, the viscosity of the formulation is 1 cP, 2 cP, 3 cP, 4 cP, 5 cP, 10 cP, 15 cP, 20 cP, 25 cP, 30 cP, 35 cP, or 40 cP at 25° C.

[0094] 1.2.10. Aggregation and Post-Peak Rates In some embodiments, the formulation reduces aggregation of IL-6 protein. In certain embodiments, the formulation reduces the formation of soluble aggregates. In certain embodiments, the formulation reduces the formation of insoluble aggregates. In certain embodiments, the formulation reduces the formation of soluble aggregates compared to formulations with less polysorbate, as measured by size-exclusion high-performance liquid chromatography (SEC-HPLC). In certain embodiments, the formulation reduces the formation of insoluble aggregates compared to formulations with less polysorbate, as measured by visual inspection. In certain embodiments, the formulation reduces the formation of soluble aggregates compared to formulations without histidine, glycine, or DTPA, as measured by RP-HPLC. In some embodiments, the formulation contains less than 15% soluble aggregates after 20 hours of agitation at 300 rpm, as measured by RP-HPLC. In some embodiments, the formulation contains less than 10% soluble aggregates after 20 hours of agitation at 300 rpm, as measured by RP-HPLC. In some embodiments, the formulation contains less than 5% soluble aggregates after 20 hours of stirring at 300 rpm as measured by RP-HPLC, hi some embodiments, the formulation contains less than 2% soluble aggregates after 20 hours of stirring at 300 rpm as measured by RP-HPLC.

[0095] In some embodiments, there is no significant increase in the % post-peak as measured by reversed-phase high performance liquid chromatography (RP-HPLC) after 14 days at -80°C. In particular embodiments, the post-peak percentage as measured by RP-HPLC after 14 days at -80°C is about 3. In other embodiments, there is no significant increase in the % post-peak as measured by RP-HPLC after 14 days at 2-8°C. In particular embodiments, the post-peak percentage as measured by RP-HPLC after 14 days at 2-8°C is about 3.

[0096] Effect In some embodiments, the formulation maintains the potency of the IL-6 protein. In various embodiments, the potency of the IL-6 protein is measured by RP-HPLC.

[0097] In some embodiments, the formulation exhibits less than a 50% loss in biological activity (e.g., relative potency) after 12 months of storage at 5±3° C. as measured by a reporter gene assay. In various embodiments, the formulation exhibits less than a 50%, less than a 40%, less than a 30%, less than a 20%, or less than a 10% loss in biological activity after 12 months of storage at 5±3° C. as measured by RP-HPLC. In certain embodiments, the formulation exhibits less than a 30% loss in biological activity after 12 months of storage at 5±3° C. as measured by RP-HPLC. In certain embodiments, the formulation exhibits less than a 20% loss in biological activity after 12 months of storage at 5±3° C. as measured by RP-HPLC. In certain embodiments, the formulation exhibits less than a 10% loss in biological activity after 12 months of storage at 5±3° C. as measured by RP-HPLC.

[0098] 1.3. Administration of the formulation Suitable routes of administration for the IL-6 formulations described herein include, but are not limited to, parenteral (e.g., subcutaneous, intravenous, intramuscular, intradermal, or intrasternal injection or infusion (e.g., as a sterile injectable aqueous or non-aqueous solution or suspension, etc.). In certain embodiments, the formulation is suitable for parenteral injection. In certain embodiments, the formulation is suitable for intravenous injection. In certain embodiments, the formulation is suitable for subcutaneous injection.

[0099] In some embodiments, the formulations are suitable for administration in a single dose or multiple doses.

[0100] 1.4. Dosage Form In other aspects, provided herein are dosage forms containing one or more unit doses of a pharmaceutical composition comprising IL-6.

[0101] In various embodiments, the dosage form is a pre-filled syringe. In various embodiments, the dosage form is an auto-injection pen. In some embodiments, the dosage form is a device with automatic administration.

[0102] In various embodiments, the dosage form comprises one or more unit doses of the formulation described above.In typical embodiments, the formulation is a liquid formulation.In other embodiments, the formulation is a dry formulation, including but not limited to, a lyophilized product.In certain embodiments, the dosage form comprises a dry formulation and a measured amount of aqueous diluent.

[0103] In some embodiments, the unit dosage form comprises a formulation containing about 40 to about 240 μg / mL IL-6, about 5 to about 15 mM histidine, about 5 to about 15 mg / mL glycine, about 40 to about 60 mg / mL trehalose, about 0.1 to about 0.3 mg / mL polysorbate 20, and about 5 to about 100 μM DTPA, wherein the formulation has a pH of about 6.0 to about 8.0.

[0104] 1.5. Treatment method In another aspect, provided herein is a method for treating a disease or disorder in a patient, comprising administering to the patient an IL-6 formulation described herein. In some embodiments, the patient has a disease or disorder that can be treated or alleviated by administering IL-6, such as chemotherapy-induced peripheral neuropathy (CIPN), weakness, or diabetic peripheral neuropathy (DPN).

[0105] The following are examples of specific embodiments for carrying out the present invention. These examples are provided for illustrative purposes only and are not intended to limit the scope of the present invention in any way. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperatures, etc.), but some experimental error and deviation should, of course, be allowed for. [Example]

[0106] Example 1 : Drug substance design for experimental testing The IL-6 formulation (drug substance "DS" formulation) contains 10 mM histidine, 10 mg / mL glycine, 50 mg / mL trehalose, 0.2 mg / mL polysorbate 20, and 50 μM DTPA at a pH of 7.0. A design of experiments (DOE) study was conducted using the finalized DS formulation. A two-way DOE study was performed with factor 1 ranging from pH 7.0 ± 0.5 and factor 2 ranging from a protein concentration of 5 ± 3 mg / mL. The details of the DOE combinations are listed in Table 1.

[0107] [Table 1]

[0108] Six different buffer solutions were prepared for the selected DS formulation. The compositions of all buffer solutions are shown in Tables 3-7 below. The IB022 standard solution provided by Sonnet BioTherapeutics contained 40 mM phosphate as the buffer component and 10 mM NaCl at pH 7.0 as the excipient. To conduct the DOE study, IB022 was buffer-exchanged into the six prepared buffer solutions. After buffer exchange, retentate samples were analyzed for protein content and pH. The protein content and pH of the retentate samples are shown in Table 8 (sample ID). The retentate was diluted as needed to the target protein concentration based on the DOE factors (listed in Table 2). All six formulations were filtered and filled into 5 mL PETG bottles in 1.5 mL fill volumes. These six formulations were subjected to stability testing at temperatures between 2 and 8°C (stress) and -80°C (real-time) for 14 days. The Phase IV excipient screening design is described in Table 2 below.

[0109] [Table 2]

[0110] [Table 3]

[0111] [Table 4]

[0112] [Table 5]

[0113] [Table 6]

[0114] [Table 7]

[0115] [Table 8]

[0116] Stage IV DS DOE samples were processed and diluted to their target DOE protein concentrations via their respective pH buffers. They were filtered, loaded onto 5 mL of PETG, and subsequently subjected to stress (2-8 °C) and real-time (-80 °C) conditions for 14 days. These samples were analyzed for protein concentration, pH, SEC-HPLC, RP-HPLC, and osmolality.

[0117] DOE samples subjected to 14-day stability testing at -80°C and 2-8°C were confirmed for purity by SEC-HPLC, as shown in Tables 9 and 10, respectively. Data for all formulations at -80°C and 2-8°C were consistent for up to 7 days. 14-day analyses were performed, but the purity trends by SEC-HPLC were not as expected or consistent, so the results are not reported.

[0118] [Table 9]

[0119] [Table 10]

[0120] The purity by RP-HPLC was confirmed for excipient screening samples subjected to stability testing at temperatures of -80°C and 2-8°C for 14 days and is shown in Tables 11 and 12, respectively. The -80°C data for all formulations was consistent over 7 days when compared to IB022, with only formulation SON080F14d showing an increase in % post-peak over 7 days of storage.

[0121] For SON080F14a, storage at 2-8°C was consistent out to 7 days, but an increase in post-peak % and a decrease in main peak % were observed for other samples compared to IB022. Results at 14 days were omitted from reporting because the purity trend by SEC-HPLC was not as expected and was out of trend.

[0122] [Table 11]

[0123] [Table 12]

[0124] DOE studies were conducted, testing various pHs and concentrations. Seven-day data showed no observed changes in quality attributes at concentrations ranging from 2 mg / mL to 8 mg / mL, with a target pH of 7.0, in the pH range of 6.5 to 7.5. The 14-day results were not as expected and were outside the trend, so were not considered in the report.

[0125] Example 2: Formulation (DP) stability test Three formulations of IL-6, 40, 120, and 240 μg / mL, liquid and lyophilized, were manufactured and subjected to 14-day stress and real-time stability studies. Results are presented for all three formulation strengths for the liquid and lyophilized stability up to 14 days. Data for the 120 μg / mL liquid formulation were discontinued on day 3 due to some unexpected results and are not reported here.

[0126] Protein concentrations were measured by RP-UPLC method at 40, 120, and 240 μg / mL for DP samples subjected to stability testing for 14 days at temperatures of 25° C. (stress, ST) and −20° C. (real-time, RT) for the liquid formulation, and 40° C. (ST) and 5° C. (RT) for the lyophilized formulation. The results are shown in Table 13, Table 14, and Table 15, respectively.

[0127] The 40 μg / mL protein concentration was consistent for both formulations up to day 14, approximately 40 μg / mL in both RT and ST conditions. The 120 μg / mL protein concentration was consistent for both formulations up to day 14, approximately 120 μg / mL in both RT and ST conditions. ST data was not reported for the 120 μg / mL liquid formulation. The 240 μg / mL protein concentration was consistent for the liquid formulation up to 14 days compared to day 0 in both RT and ST conditions, while it was consistent for the lyophilized formulation up to 14 days compared to day 0 in both RT and ST conditions.

[0128] [Table 13]

[0129] [Table 14]

[0130] [Table 15]

[0131] The pH values ​​of the 40, 120, and 240 μg / mL formulation samples, which were subjected to stability testing for 14 days at 25°C (ST) and -20°C (RT) for the liquid formulation and at 40°C (ST) and 5°C (RT) for the lyophilized formulation, are shown in Tables 16, 17, and 18, respectively. Because the initial target pH for the formulation was 7.0, stability testing of the 40 and 120 μg / mL formulations was performed at pH 7.0. For the 240 μg / mL formulation, the pH was updated to 7.2, so a formulation with a pH of 7.2 was prepared.

[0132] The pH of 40 μg / mL and 120 μg / mL for both conditions (RT and ST) and formulations (liquid and lyophilized) up to 14 days was consistent and ranged from 6.8 to 7.3. The pH of 240 μg / mL for both liquid and lyophilized formulations up to 14 days was consistent at pH 7.0 to 7.1 in both RT and ST conditions.

[0133] [Table 16]

[0134] [Table 17]

[0135] [Table 18]

[0136] Purity by SEC-HPLC was measured for formulation samples at 40, 120, and 240 μg / mL when subjected to stability testing for 14 days at temperatures of 25° C. (ST) and −20° C. (RT) for the liquid formulation, and 40° C. (ST) and 5° C. (RT) for the lyophilized formulation. The results are shown in Tables 19, 20, and 21, respectively.

[0137] Purity at 40 μg / mL was consistent for both conditions (RT and ST) and for both formulations (liquid and lyophilized) up to 14 days, except that an increase in HMW% was observed at day 14 in the stressed condition for the lyophilized formulation. Purity at 120 μg / mL for liquid formulations at RT conditions up to 14 days was consistent, but ST data was not reported. For the lyophilized formulations at 120 μg / mL up to 7 days, purity data was consistent, but at day 14, both RT and ST data show an increase in HMW%. Purity at 240 μg / mL for both conditions (RT and ST) and for both formulations (liquid and lyophilized) up to 14 days was consistent, with no change observed.

[0138] [Table 19]

[0139] [Table 20]

[0140] [Table 21]

[0141] Purity by RP-HPLC was measured for 40, 120, and 240 μg / mL DP samples subjected to stability testing for 14 days at temperatures of 25° C. (ST) and −20° C. (RT) for the liquid formulation, and 40° C. (ST) and 5° C. (RT) for the lyophilized formulation. The results are shown in Table 22, Table 23, and Table 24, respectively.

[0142] Purity at 40 μg / mL was consistent for both conditions (RT and ST) of the lyophilized formulation for up to 14 days. In contrast, an increase in the pre-peak and post-peak percentages was observed after the liquid formulation on day 0 for both conditions (RT and ST) at 40 μg / mL. Purity at 120 μg / mL for the liquid formulation at RT conditions for up to 3 days was consistent, with a decrease in the pre-peak percentage and an increase in the main peak percentage on days 7 and 14. ST data was not reported for the 120 μg / mL liquid formulation. A decrease in the pre-peak percentage was observed on days 7 and 14 for the 120 μg / mL lyophilized formulation at RT conditions, but for the ST condition, the data were consistent for up to 14 days. Purity at 240 μg / mL for both conditions (RT and ST) and both formulations (liquid and lyophilized) for up to 14 days was consistent, with no change observed.

[0143] [Table 22]

[0144] [Table 23]

[0145] [Table 24]

[0146] The formulations in liquid and lyophilized form were found to be stable under stress conditions for up to 14 days for low and high concentrations (40 μg / mL and 240 μg / mL). The results are applicable to all intermediate strengths.

[0147] Example 3: Stabilization of formulations by adding chelating agents Subvisible particles in drug products may consist of aggregated proteins and can directly affect the efficacy and immunogenicity of pharmaceutical products. Subvisible particles often act as nucleation sites for further protein aggregation and / or can lead to the generation of larger particles due to aggregation (see "Particulate contamination," Chapter 2.9.19, Subvisible Particles, published in Pharmauropa 33.2).

[0148] To evaluate the effect of chelating agents such as EDTA or DTPA on the stability of the formulations of Example 1, subvisible particle count analysis was performed at 37°C using a HIAC 9703+ liquid particle counter system. The following formulations were tested: SON080F49a (without EDTA or DTPA), SON080F49b (with DTPA), and SON080F49c (with EDTA). The SON080F49a formulation contains 2 mg / mL IL-6, 10 mM histidine buffer, 10 mg / mL glycine, 50 mg / mL trehalose, 0.2 mg / mL polysorbate 20, water for injection, and has a pH of 7.2.

[0149] [Table 25]

[0150] The addition of a chelating agent to the SON080F49a formulation reduced the number of large subvisible particles of 2, 5, 10, and 25 μm on days 1, 7, and 14. Furthermore, the formulation containing DTPA (SON080F49b) unexpectedly reduced the number of subvisible particles compared to the formulation containing EDTA (SON080F49c).

[0151] Unless otherwise expressly indicated, all numbers expressing quantities of ingredients, molecular weights, reaction conditions, and so forth, used in the specification and claims should be understood as being modified in all instances by the term "about." Accordingly, unless indicated to the contrary, the numerical parameters set forth in the specification and appended claims are approximations that may vary depending upon the desired properties sought to be obtained by the present disclosure. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques.

[0152] Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the disclosure are approximations, the numerical values ​​set forth in the specific examples are reported as precisely as possible. However, any numerical value inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0153] As used in the context of describing this disclosure (particularly in the context of the claims below), the terms "a," "an," "the," and similar referents should be construed to cover both the singular and the plural unless otherwise indicated herein or clearly contradicted by context. The recitation of ranges of values ​​herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each separate value is incorporated herein as if each were individually set forth herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or clearly contradicted by context. The use of any and all examples or exemplary language (e.g., "etc.") provided herein is intended merely to better clarify the disclosure and does not pose a limitation on the scope of the claims. No language in the specification should be construed as indicating any element essential to the practice of the disclosure but absent from the recitation of a claim.

[0154] Groupings of alternative elements or embodiments of the present disclosure disclosed herein are not to be construed as limiting. Each group member may be referred to or claimed individually, or may be referred to or claimed in any combination with other members of the group or other elements found herein. It will be understood that one or more members of a group may be included in, or deleted from, a group for reasons of convenience and / or patentability. When such inclusions or deletions occur, the specification is deemed to encompass the modified group, thus satisfying the written description requirement for all Markush groups used in the appended claims.

[0155] Some embodiments of the present disclosure are described herein, including the best mode known to the inventors for carrying out the disclosure. Of course, variations on these described embodiments will become apparent to those skilled in the art upon reading the foregoing description. The inventors anticipate that skilled artisans will employ such variations as appropriate, and the inventors contemplate that the present disclosure may be practiced in ways not specifically described herein. Accordingly, this disclosure includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the present disclosure unless otherwise indicated herein or clearly contradicted by context.

[0156] Certain embodiments disclosed herein may be further limited in the claims using the phrases "consisting of" or "consisting essentially of." When used in a claim as filed or added by amendment, the transitional phrase "consisting of" excludes any element, step, or ingredient not specified in the claim. The transitional phrase "consisting essentially of" limits the claim to the specified materials or steps and to materials or steps that do not materially affect the basic and novel characteristic(s). An embodiment of the present disclosure so claimed is essentially or explicitly described herein and satisfies enablement requirements.

[0157] It will be understood that the embodiments of the present disclosure disclosed herein are illustrative of the principles of the present disclosure. Other variations that may be employed are within the scope of the present disclosure. Thus, for example, but not by way of limitation, alternative configurations of the present disclosure may be utilized in accordance with the teachings herein. Thus, the present disclosure is not limited to that exact matter shown and described.

[0158] While the present disclosure has been described and illustrated herein by reference to various specific materials, procedures, and examples, it will be understood that the disclosure is not limited to the particular combination of materials and procedures selected for that purpose. Numerous variations of these details may be implied as will be understood by those skilled in the art. It is intended that the specification and examples be considered exemplary only, with the true scope and spirit of the present disclosure being indicated by the following claims. All references, patents, and patent applications mentioned in this application are incorporated herein by reference in their entirety.

Claims

1. 1. An IL-6 preparation, comprising: - about 40 μg / mL to about 2 mg / mL of IL-6; about 5 to about 15 mM histidine, and - about 5 to about 15 mg / mL glycine; about 40 to about 60 mg / mL of trehalose; and - about 0.1 to about 0.3 mg / mL of polysorbate 20, - about 1 to about 1000 μM DTPA, The IL-6 formulation, wherein the formulation has a pH of about 6.0 to about 8.

0.

2. The IL-6 preparation according to claim 1, wherein the IL-6 is human IL-6.

3. 10. The IL-6 formulation of claim 1, wherein the formulation comprises 40 μg / mL IL-6, 45 μg / mL IL-6, 50 μg / mL IL-6, 55 μg / mL IL-6, 60 μg / mL IL-6, 65 μg / mL IL-6, 70 μg / mL IL-6, 75 μg / mL IL-6, 80 μg / mL IL-6, 85 μg / mL IL-6 IL-6, 90 μg / mL IL-6, 95 μg / mL IL-6, 100 μg / mL IL-6, 105 μg / mL IL-6, 110 μg / mL IL-6, 115 μg / mL IL-6, 120 μg / mL IL-6, 125 μg / mL IL-6, 130 μg / mL IL-6, 135 μg / mL IL-6, 140 μg / mL mL IL-6, 145 μg / mL IL-6, 150 μg / mL IL-6, 155 μg / mL IL-6, 160 μg / mL IL-6, 165 μg / mL IL-6, 170 μg / mL IL-6, 175 μg / mL IL-6, 180 μg / mL IL-6, 185 μg / mL IL-6, 190 μg / mL IL-6, 19 IL-6 formulations comprising 5 μg / mL IL-6, 200 μg / mL IL-6, 205 μg / mL IL-6, 210 μg / mL IL-6, 215 μg / mL IL-6, 220 μg / mL IL-6, 225 μg / mL IL-6, 230 μg / mL IL-6, 235 μg / mL IL-6, or 240 μg / mL IL-6.

4. 4. The IL-6 formulation of claim 3, wherein the formulation comprises 80 μg / mL of IL-6.

5. 4. The IL-6 formulation of claim 3, wherein the formulation comprises 120 μg / mL of IL-6.

6. 4. The IL-6 formulation of claim 3, wherein the formulation comprises 240 μg / mL of IL-6.

7. 2. The IL-6 formulation of claim 1, wherein the formulation comprises about 10 mM histidine.

8. 2. The IL-6 formulation of claim 1, wherein the formulation comprises about 10 mg / mL of glycine.

9. 2. The IL-6 formulation of claim 1, wherein the formulation comprises about 50 mg / mL of trehalose.

10. 2. The IL-6 formulation of claim 1, wherein the formulation comprises about 0.2 mg / mL polysorbate 20.

11. 2. The IL-6 formulation of claim 1, wherein the formulation comprises about 50 μM DTPA.

12. 2. The IL-6 formulation of claim 1, wherein the formulation has a pH of about 7.

0.

13. 1. An IL-6 preparation, comprising: -40 to about 240 μg / mL human IL-6; 10 mM histidine, 10 mg / mL glycine, 50 mg / mL trehalose, - 0.2 mg / mL polysorbate 20, - 50 μM DTPA, The IL-6 formulation, wherein the formulation has a pH of about 7.

0.

14. 14. The IL-6 formulation of claim 13, wherein the formulation comprises 40 μg / mL IL-6, 45 μg / mL IL-6, 50 μg / mL IL-6, 55 μg / mL IL-6, 60 μg / mL IL-6, 65 μg / mL IL-6, 70 μg / mL IL-6, 75 μg / mL IL-6, 80 μg / mL IL-6, 85 μg / mL IL-6 IL-6, 90 μg / mL IL-6, 95 μg / mL IL-6, 100 μg / mL IL-6, 105 μg / mL IL-6, 110 μg / mL IL-6, 115 μg / mL IL-6, 120 μg / mL IL-6, 125 μg / mL IL-6, 130 μg / mL IL-6, 135 μg / mL IL-6, 140 μg / mL mL IL-6, 145 μg / mL IL-6, 150 μg / mL IL-6, 155 μg / mL IL-6, 160 μg / mL IL-6, 165 μg / mL IL-6, 170 μg / mL IL-6, 175 μg / mL IL-6, 180 μg / mL IL-6, 185 μg / mL IL-6, 190 μg / mL IL-6, 19 IL-6 formulations comprising 5 μg / mL IL-6, 200 μg / mL IL-6, 205 μg / mL IL-6, 210 μg / mL IL-6, 215 μg / mL IL-6, 220 μg / mL IL-6, 225 μg / mL IL-6, 230 μg / mL IL-6, 235 μg / mL IL-6, or 240 μg / mL IL-6.

15. 15. The IL-6 formulation of claim 14, wherein the formulation comprises 80 μg / mL of IL-6.

16. 15. The IL-6 formulation of claim 14, wherein the formulation comprises 120 μg / mL of IL-6.

17. 15. The IL-6 formulation of claim 14, wherein the formulation comprises 240 μg / mL of IL-6.

18. The IL-6 formulation of claim 13, wherein the formulation is suitable for parenteral administration.

19. The IL-6 formulation of claim 13, wherein the formulation is suitable for intravenous administration.

20. The IL-6 formulation of claim 13, wherein the formulation is suitable for subcutaneous administration.

21. 10. A method of treating a disease or disorder in a subject in need thereof, comprising administering to the subject the formulation of claim 1 or claim 6.

22. 2. The IL-6 formulation of claim 1, wherein the formulation has less than about 3% post-peak as measured by RP-HPLC after 14 days at −80° C.

23. 2. The IL-6 formulation of claim 1, wherein the formulation has less than about 3% post-peak as measured by RP-HPLC after 14 days at 2-8°C.

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