Low dose il-6 formulations and methods of use thereof

Abstract

The present disclosure provides an IL-6 formulation comprising from about 40 [mu] g / mL to about 2 mg / ml of IL-6, from about 5 mM to about 15 mM of histidine, from about 5 mg / mL to about 15 mg / mL of glycine, from about 40 mg / mL to about 60 mg / mL of trehalose, from about 0.1 mg / mL to about 0.3 mg / mL of polysorbate 20, and from about 1 [mu] M to about 1000 [mu] M of DTPA wherein the pH of the formulation is from about 6.0 to about 8.0. Such formulations may be used to treat and / or prevent a disease or condition in a subject in need thereof.

Description

[0001] Cross-reference to related applications

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 377,971, filed on September 30, 2022, which is hereby incorporated by reference in its entirety. Technical Field

[0003] The present disclosure generally provides IL-6 formulations, including stable low-dose formulations suitable for both frozen liquid and lyophilized storage, and methods of use thereof, including for treating IL-6-related diseases and / or disorders.

[0004] Sequence Listing

[0005] This application contains a sequence listing, which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. The name of the XML file is 116076_5019_PR_Sequence_Listing.xml, created on September 29, 2023, and is 4000 bytes in size. Background Art

[0006] Interleukin-6 (hereinafter referred to as "IL-6") (also known as interferon-β 2 ; B cell differentiation factor; B cell stimulating factor-2; hepatocyte stimulating factor; hybridoma growth factor; and plasmacytoma growth factor) are members of the cytokine family that promote cellular responses through a receptor complex consisting of at least one subunit of the signaling glycoprotein gp130 and the IL-6 receptor ("IL-6R"; also known as gp80). IL-6R can also exist in a soluble form ("sIL-6R"). IL-6 binds to IL-6R and then dimerizes the signaling receptor gp130. See Jones, SA, J. Immunology, 175:3463-3468 (2005).

[0007] Although IL-6 cannot bind directly to gp130, it can bind to IL-6R to generate a high-affinity ternary complex of IL-6 / IL-6R / gp130. IL-6 binds to IL-6R with low affinity, however, IL-6R does not have an intracellular signal transduction domain, so this single connection does not lead to cell activation. Similarly, cell surface expression of IL-6R does not mean that the cell responds to IL-6 stimulation. Proteolytic cleavage leads to the release of soluble IL-6R (sIL-6R; sgp80), which can bind to circulating IL-6 and prolong the half-life of IL-6. For cell activation, IL-6 first binds to either of the cell-bound IL-6R or sIL-6R; the heterodimeric IL-6 / IL-6R complex then associates with the cell surface glycoprotein gp130. The resulting tripartite heterocomplex binds another IL-6 / IL-6R / gp130 and subsequently undergoes signal transduction (Bravo and Heath 2000, EMBO J., 19, (11), 2399-2411; Boulanger et al., 2003, Science, 300, 5628, 2101-2104), so both cell-bound and soluble IL-6R contribute to cell activation. IL-6 signal transduction via cell-bound IL-6R is referred to as cis signal transduction, while cell activation via soluble IL-6R is described as trans signal transduction.

[0008] The preparations that maintain the stability of IL-6 under various conditions promote the therapeutic use of IL-6. Importantly, therapeutic preparations allow storage of active proteins without unacceptable activity loss, minimize the accumulation of undesirable products such as aggregates or degradation species (e.g., fragmentation, oxidation, deamidation or isomerization species), accommodate protein of appropriate concentration, and do not contain components incompatible with therapeutic applications. In addition, previous IL-6 preparations use regulatory agencies no longer recommend chemical substances for human use, such as chemical substances containing sucrose, or chemical substances that are unstable in liquid and lyophilized forms. Therefore, there is a need in the art for a stable aqueous pharmaceutical preparation comprising IL-6 suitable for therapeutic use. Summary of the invention

[0009] The present disclosure generally provides a unique stable and / or sucrose-free IL-6 formulation in both liquid and lyophilized form. Such formulations may include about 40 μg / mL to about 2 mg / ml (e.g., 80-240 μg / mL) 240 μg / mL IL-6, about 5 mM to about 15 mM histidine, about 5 mg / mL to about 15 mg / mL glycine, about 40 mg / mL to about 60 mg / mL trehalose, about 0.1 mg / mL to about 0.3 mg / mL polysorbate 20, and about 1 μM 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, 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), wherein the pH of the formulation is about 6.0 to about 8.0.

[0010] In some embodiments, the IL-6 is human IL-6.

[0011] In some embodiments, the IL-6 comprises the sequence shown in SEQ ID NO:1 or SEQ ID NO:2.

[0012] In some embodiments, 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, 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, 195 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. In another embodiment, the formulation comprises about 80 μg / mL to about 2 mg / ml IL-6.

[0013] In some embodiments, the formulation comprises 40 ug / ml IL-6.

[0014] In some embodiments, the formulation comprises 80 μg / mL IL-6.

[0015] In some embodiments, the formulation comprises 160 μg / mL IL-6.

[0016] In some embodiments, the formulation comprises 240 μg / mL IL-6.

[0017] In some embodiments, the formulation comprises about 10 mM histidine.

[0018] In some embodiments, the formulation comprises about 10 mg / mL glycine.

[0019] In some embodiments, the formulation comprises about 50 mg / mL trehalose.

[0020] In some embodiments, the formulation comprises about 0.2 mg / mL polysorbate 20.

[0021] In some embodiments, the formulation comprises about 50 μM DTPA.

[0022] In some embodiments, the pH of the formulation is about 7.0.

[0023] In some embodiments, the formulation is a liquid formulation.

[0024] In some embodiments, the formulation is a lyophilized formulation.

[0025] The present disclosure also provides an IL-6 formulation comprising 80 μg / mL to 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 pH of the formulation is about 7.0.

[0026] In some embodiments, 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, 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, 195 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. In another embodiment, the formulation comprises about 40 μg / mL to about 2 mg / ml IL-6.

[0027] In some embodiments, the formulation comprises 40 μg / mL IL-6.

[0028] In some embodiments, the formulation comprises 80 μg / mL IL-6.

[0029] In some embodiments, the formulation comprises 160 μg / mL IL-6.

[0030] In some embodiments, the formulation comprises 240 μg / mL IL-6.

[0031] In some embodiments, the formulations are suitable for parenteral administration.

[0032] In some embodiments, the formulations are suitable for intravenous administration.

[0033] In some embodiments, the formulation is suitable for subcutaneous administration.

[0034] The present disclosure also provides methods of treating an IL-6-related disease or disorder in a subject by administering to a subject in need thereof a formulation as disclosed herein. DETAILED DESCRIPTION

[0035] The present disclosure provides IL-6 (e.g., h-rIL-6) preparations, including stable low-dose preparations suitable for both frozen liquid and lyophilized storage, and methods of use thereof, including for treating IL-6 related diseases and / or disorders. Advantageously, the formulated IL-6 needs to be stored in a stable presentation (e.g., not degraded for a long time at different temperatures, withstanding multiple freezing and thawing cycles, and conveniently used for injection after thawing or reconstitution with water for injection), and can be used for diabetic patients whose diet does not allow for the intake of sugars.

[0036] 1.1 Definition

[0037] The following definitions are included to provide a clear and consistent understanding of the specification and claims. As used herein, the terms described have the following meanings. All other terms and phrases used in this specification have the common meanings as will be understood by those skilled in the art. Such common meanings can be obtained by reference to technical dictionaries, such as Hawley's "Condensed Chemical Dictionary (Condensed Chemical Dictionary)" 14th edition, author RJ Lewis, John Wiley & Sons, New York, NY, 2001.

[0038] References in this specification to "one embodiment," "an embodiment," etc. indicate that the described embodiment may include a particular aspect, feature, structure, part, or characteristic, but not every embodiment must include that aspect, feature, structure, part, or characteristic. In addition, such phrases may, but do not necessarily, refer to the same embodiment referenced in other parts of this specification. Further, when particular aspects, features, structures, parts, or characteristics are described in conjunction with an embodiment, it is within the knowledge of those skilled in the art to affect such aspects, features, structures, parts, or characteristics or to combine them with other embodiments, whether or not explicitly described.

[0039] Unless the context clearly dictates otherwise, the singular forms "a," "an," and "the" include plural referents. Thus, for example, reference to "a formulation" includes a plurality of such formulations, and thus formulation X includes a plurality of formulations X. It should be further noted that the claims may be drafted to exclude any optional elements. Therefore, this statement is intended to serve as antecedent basis for use of exclusive terminology such as "solely," "only," or use of a "negative" limitation in connection with any element described herein and / or the recitation of a claim element.

[0040] The term "and / or" means any one, any combination of, or all of the items associated with the term. The phrases "one or more" and "at least one" are readily understood by those skilled in the art, especially when read in the context in which they are used. For example, the phrases may mean one, two, three, four, five, six, ten, 100, or any upper limit.

[0041] A "stable" IL-6 formulation is one in which the IL-6 substantially maintains its physical stability and / or chemical stability and / or its biological activity during storage. The storage period is usually selected based on the expected shelf life of the formulation evaluated at different temperatures (from -80°C to +40°C). Various analytical techniques for measuring protein stability can be used in the art. Examples of analytical techniques are described below. "Substantially maintained" is intended to maintain 85% or more, such as at least 90% or at least 95%, depending on the analytical technique and specification used.

[0042] An IL-6 protein "maintains its physical stability" in a pharmaceutical formulation if it exhibits no significant physical changes, such as aggregation, precipitation, and / or denaturation, when visually inspected for color and / or clarity or when measured by UV light scattering or size exclusion chromatography.

[0043] If no significant chemical changes of the protein are shown, the IL-6 protein "maintains its chemical stability" in the pharmaceutical formulation. Chemical stability can be assessed by detecting and quantifying chemically altered forms of the protein. The chemical changes can involve size modifications (e.g., shearing), which can be assessed using, for example, 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., due to deamidation, oxidation, and / or isomerization), which can be assessed by, for example, ion exchange chromatography.

[0044] An IL-6 protein "retains its biological activity" in a pharmaceutical formulation if the biological activity of the protein at a given time is not significantly altered from the biological activity exhibited when the pharmaceutical formulation is 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.

[0045] "Histidine buffer" is a buffer comprising histidine ions. Examples of histidine buffers include histidine chloride, histidine acetate, histidine phosphate, and histidine sulfate solutions. The pH of a histidine buffer or a histidine-HCl buffer is 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.

[0046] "IL-6" or "interleukin 6 (IL-6)" or "IL-6 polypeptide" means a polypeptide or fragment thereof having at least about 85% or greater amino acid identity to the amino acid sequence provided at NCBI Accession No. NP_000591 and having IL-6 biological activity. IL-6 is a pleiotropic cytokine with multiple biological functions. Exemplary IL-6 biological activities include immunostimulatory and pro-inflammatory activities. "IL-6" or "interleukin 6 (IL-6) nucleic acid" means a polynucleotide encoding an interleukin 6 (IL-6) polypeptide. Exemplary interleukin 6 (IL-6) nucleic acid sequences are provided at NCBI Accession No. NM_000600.

[0047] The "identity" percentage between a polypeptide sequence and a 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 the alignment sequence is aligned and spaces are introduced (if necessary) to achieve the maximum sequence identity percentage. The comparison for the purpose of determining the amino acid sequence identity percentage can be achieved in various ways within 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 aligning sequences, including any algorithm required for achieving maximum alignment over the full length of the compared sequence. Unless otherwise indicated, the BLAST algorithm is used to determine the sequence identity percentage using default parameters.

[0048] "Subject" means a human or non-human mammalian (including but not limited to bovine, equine, canine, ovine, feline, and rodent (including mouse and rat)) subject. A "patient" is a human subject.

[0049] As used herein, the term "treat", "treating", "treatment" refers to reducing or improving a disease and / or signs or symptoms associated therewith, or slowing or stopping its progress. It should be understood that, although not excluded, treating a condition or disorder does not require complete elimination of the condition, disorder or symptoms associated therewith.

[0050] In this disclosure, "comprises," "comprising," "containing," "having," "includes," "including," and language variations thereof have the meanings ascribed to them by U.S. patent law, allowing for the presence of additional components other than those explicitly listed.

[0051] As will be appreciated by those skilled in the art, all numbers, including those expressing the amount of components, properties (such as molecular weight), reaction conditions, etc., are approximate values ​​and are understood to be optionally modified by the term "about" in all cases. These values ​​can vary according to the desired properties sought to be obtained by those skilled in the art using the teachings described herein. It should also be understood that such values ​​inherently contain variability caused by the standard deviations found in their corresponding test measurements. When a value is expressed as an approximation by using the aforementioned word "about", it will be understood that the specific value without the modifier "about" also forms an additional aspect.

[0052] The terms "about" and "approximately" are used interchangeably. Both terms can refer to a change of ±1%, ±5%, or ±10% of a specified value. For example, in some embodiments, a "about 50" percentage may have a change from 45 percentages to 55 percentages, or as otherwise defined by a particular claim. For integer ranges, the term "about" may include one or two integers greater than and / or less than the stated integer at each end of the range. Unless otherwise indicated herein, the terms "about" and "approximately" are intended to include values ​​that are equal in functionality to the individual ingredients, compositions, or embodiments that are close to the stated range, for example, weight percentages. The terms "about" and "approximately" may also modify the endpoints of the stated range as discussed above in this paragraph.

[0053] As will be understood by those skilled in the art, for any and all purposes, particularly in providing a written description, all ranges described herein also encompass any and all possible sub-ranges and combinations of sub-ranges thereof, as well as individual values, particularly integer values, constituting the ranges. Therefore, it will be understood that each unit between two specific units is also disclosed. For example, if 10 to 15 are disclosed, 11, 12, 13, and 14 are also disclosed individually and as part of a range. The described range (e.g., weight percentage or carbon group) includes each specific value, integer, decimal, or identity within the described range. Any listed range can be easily recognized as fully describing and implementing the same range that is decomposed into at least two, three, four, five, or ten equal parts. As a non-limiting example, each range discussed herein can be easily decomposed into a lower third, a middle third, and an upper third, etc. As will also be understood by those skilled in the art, all languages ​​such as "at most," "at least," "greater than," "less than 1," "more than," "or more" include the described numbers, and such terms refer to the ranges that can be subsequently decomposed into sub-ranges as discussed above. In the same manner, all ratios recited herein also include all subratios falling within the broader ratios. Thus, specific values ​​recited for radicals, substituents, and ranges are for illustration only; they do not exclude other defined values ​​for radicals and substituents or other values ​​within defined ranges. It will be further understood that the endpoints of each of the ranges are significant relative to the other endpoint and independently of the other endpoint.

[0054] The present disclosure provides ranges, limits and deviations for variables such as volume, mass, percentage, ratio, etc. It will be understood by those of ordinary skill in the art that ranges (such as "number 1" to "number 2") are meant to include a continuous range of integers and fractions. For example, 1 to 10 means 1, 2, 3, 4, 5, ... 9, 10. It also means 1.0, 1.1, 1.2, 1.3, ..., 9.8, 9.9, 10.0, and also means 1.01, 1.02, 1.03, etc. If the variable disclosed is a number less than "number 10", it means a continuous range of integers and fractions less than number 10, as discussed above.

[0055] Similarly, if a variable is disclosed as a number greater than "the number 10," it is meant to include a continuous range of integers and fractions greater than the number 10.

[0056] These ranges may be modified by the term "about", the meaning of which has been described above.

[0057] Those skilled in the art will also readily recognize that where members are grouped together in a common manner, such as in a Markush group, the present invention encompasses not only the entire group listed as a whole, but also each member of the group and all possible subgroups of the main group individually. In addition, for all purposes, the present invention encompasses not only the main group, but also the main group lacking one or more group members. Thus, the present invention contemplates the explicit exclusion of any one or more of the members of the recited group. Thus, a proviso may apply to any disclosed class or embodiment, thereby excluding any one or more of the recited elements, species, or embodiments from such class or embodiment, for example, for an explicit negative limitation.

[0058] 1.2. Preparation

[0059] In the first aspect, an aqueous or lyophilized preparation (pharmaceutical composition) comprising IL-6 protein is provided herein. In an embodiment, the preparation is sterile. In another embodiment, the preparation is stable. In some embodiments, the preparation further comprises at least one free amino acid. In some embodiments, the preparation further comprises at least one surfactant. In some embodiments, the preparation further comprises at least one buffer component. In some embodiments, the preparation further comprises at least one chelating agent. In some embodiments, the preparation comprises at least one carbohydrate. In some embodiments, the preparation comprises IL-6 (e.g., human IL-6). In certain embodiments, the preparation comprises at least one free amino acid, at least one surfactant, at least one buffer component, at least one chelating agent, at least one carbohydrate, and IL-6.

[0060] In some embodiments, the formulation comprises about 40 μg / mL to about 240 μg / mL IL-6, about 5 mM to about 15 mM histidine, about 5 mg / mL to about 15 mg / mL glycine, about 40 mg / mL to about 60 mg / mL trehalose, about 0.1 mg / mL to about 0.3 mg / mL polysorbate 20, and about 5 μM to about 100 μM DTPA, wherein the pH of the formulation is about 6.0 to about 8.0. In other embodiments, the formulation comprises 40 μg / mL 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, and 50 μM DTPA, wherein the pH of the formulation is about 7.0.

[0061] IL-6 concentration

[0062] 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 concentration of IL-6 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 220 250 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.

[0063] In some embodiments, the IL-6 concentration in the formulation is about 40 μg / mL to about 280 μg / mL. In other embodiments, the IL-6 concentration in the formulation is about 100 μg / mL to about 120 μg / mL, about 120 μg / mL to about 140 μg / mL, about 140 μg / mL to about 160 μg / mL, about 160 μg / mL to about 180 μg / mL, about 180 μg / mL to about 200 μg / mL, about 200 μg / mL to about 220 μg / mL, about 220 μg / mL to about 240 μg / mL, about 240 μg / mL to about 260 μg / mL, or about 260 μg / mL to about 280 μg / mL.

[0064] 1.2.2. Chelating agents

[0065] In some embodiments, the formulation comprises a chelating agent.

[0066] In various embodiments, the concentration of the chelator is about 1 μM to about 1000 μM, such as about 25 μM to about 75 μM, preferably about 50 μM. In various embodiments, the concentration of the chelator is 1 μM to 100 μM, such as 25 μM to 75 μM, preferably 50 μM.

[0067] 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).

[0068] 1.2.3. Surfactants

[0069] In some embodiments, the formulation comprises a surfactant.

[0070] Surfactants can reduce the surface tension of a liquid. In some embodiments, the surfactant is a nonionic surfactant. Examples of surfactants include: polysorbates (polyoxyethylene sorbitan monolaurate, e.g., polysorbate 20 and polysorbate 80); TRITON (tert-octylphenoxypolyethoxyethanol); sodium dodecyl sulfate (SDS); sodium lauryl sulfate; sodium octyl glucoside; lauryl-sulfobetaine, myristyl-sulfobetaine, linoleyl-sulfobetaine, or stearyl-sulfobetaine; lauryl-sarcosine, myristyl-sarcosine, linoleyl-sarcosine, or stearyl-sarcosine; linoleyl-betaine, myristyl-sultaine In some embodiments, the surfactant is a polysorbate. In certain embodiments, the polysorbate is polysorbate 20, polysorbate 40, polysorbate 60, or polysorbate 80.

[0071] In various embodiments, the composition comprises a surfactant at a concentration of about 0.001% to about 1%, such as 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%, such as 0.001% to 0.1%, 0.005% to 0.2%, 0.01% to 0.2%, or 0.05% to 0.1% (w / v).

[0072] 1.2.4. Polysorbates

[0073] In some embodiments, the formulation comprises polysorbate.

[0074] 1.2.4.1. Polysorbate 80

[0075] In some embodiments, the formulation comprises polysorbate 80 (PS80).

[0076] 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 about 0.001% to about 1%, about 0.002% to about 0.5%, about 0.005% to about 0.2%, or about 0.01% to 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 specific embodiments, the formulation comprises about 0.07% (w / v) polysorbate 80.

[0077] 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 a specific embodiment, the formulation comprises 0.07% (w / v) polysorbate 80.

[0078] 1.2.4.2. Polysorbate 60

[0079] In some embodiments, the formulation comprises polysorbate 60 (PS60).

[0080] 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 about 0.001% to about 1%, about 0.002% to about 0.5%, about 0.005% to about 0.2%, or about 0.01% to 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 specific embodiments, the formulation comprises about 0.07% (w / v) polysorbate 60.

[0081] 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% to 0.1% (w / v) polysorbate 60. In certain embodiments, the formulation comprises 0.05% to 0.1% (w / v) polysorbate 60. In a specific embodiment, the formulation comprises 0.07% (w / v) polysorbate 60.

[0082] 1.2.4.3. Polysorbate 40

[0083] In some embodiments, the formulation comprises polysorbate 40 (PS40).

[0084] 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 about 0.001% to about 1%, about 0.002% to about 0.5%, about 0.005% to about 0.2%, or about 0.01% to 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 specific embodiments, the formulation comprises about 0.07% (w / v) polysorbate 40.

[0085] 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 a specific embodiment, the formulation comprises 0.07% (w / v) polysorbate 40.

[0086] 1.2.4.4. Polysorbate 20

[0087] In some embodiments, the formulation comprises polysorbate 20 (PS20).

[0088] 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 about 0.001% to about 1%, about 0.002% to about 0.5%, about 0.005% to about 0.2%, or about 0.01% to 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 specific embodiments, the formulation comprises about 0.07% (w / v) polysorbate 20.

[0089] 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 a specific embodiment, the formulation comprises 0.07% (w / v) polysorbate 20.

[0090] 1.2.5. Sugar

[0091] In certain embodiments, compositions comprises carbohydrate and derivatives thereof, comprises monosaccharide, disaccharide, trisaccharide, polysaccharide, sugar alcohol, reducing sugar, non-reducing sugar etc.The example of carbohydrate comprises glucose, mannose, trehalose, lactose, fructose, maltose, dextran, dextrin, erythritol, glycerol, arabitol, xylitol, sorbitol, mannitol, melibiose, melezitose, raffinose, mannotriose, stachyose, maltose, lactulose, maltulose, glucitol, maltitol, lactitol, isomaltulose etc.In certain embodiments, carbohydrate is disaccharide.In certain embodiments, disaccharide is trehalose.In various embodiments, the concentration of disaccharide is about 1mg / mL to about 100mg / mL.

[0092] 1.2.5.1. Trehalose

[0093] In some embodiments, the formulation comprises trehalose.

[0094] In various embodiments, the formulation comprises about 1 mg / mL to about 100 mg / mL trehalose. In some embodiments, the formulation comprises 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 trehalose. In a specific embodiment, the formulation comprises about 50 mg / mL trehalose.

[0095] 1.2.6. Free amino acids

[0096] 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 can be in L-type, D-type, or a mixture of these forms.

[0097] In certain embodiments, the at least one free amino acid is glycine, glutamine, asparagine, histidine, arginine or lysine. In various embodiments, the concentration of the free amino acid is about 1mM to about 150mM, such as about 2mM, 3mM, 4mM, 5mM, 6mM, 7mM, 8mM, 9mM, 10mM, 11mM, 12mM, 13mM, 15mM, 16mM, 17mM, 18mM, 19mM or 20mM.

[0098] 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.

[0099] 1.2.6.1. Glycine

[0100] In some embodiments, the formulation comprises glycine. In various embodiments, the formulation comprises about 5 mg / mL to about 15 mg / mL 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 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 a specific embodiment, the formulation comprises about 10 mg / mL glycine.

[0101] 1.2.7. Buffer

[0102] In some embodiments, the formulation comprises at least one buffer (buffer component). Typically, the buffer, when present, 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.

[0103] In various embodiments, the at least one buffer 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), glycine and other organic acid buffers. In some of these embodiments, the buffer is histidine, citrate, phosphate, glycine or acetate. In various embodiments, the concentration of the buffer component is 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 concentration of the buffer component is 1 mM to 200 mM, 1 mM to 50 mM or 5 mM to 20 mM. In certain embodiments, the concentration of the buffer component is about 5 mM, about 10 mM, about 15 mM, about 20 mM, or about 25 mM. In certain embodiments, the concentration of the buffer component is 10 mM.

[0104] In preferred embodiments, the buffer is histidine. In some embodiments, the formulation comprises histidine. In some embodiments, the formulation comprises L-histidine.

[0105] In some embodiments, the composition comprises at least about 1mM, at least about 5mM, at least about 10mM, at least about 15mM, at least about 20mM, at least about 30mM, at least about 50mM, at least about 100mM, at least about 150mM or at least about 200mM histidine. In various embodiments, the composition comprises about 1mM to about 200mM, about 5mM to about 150mM, about 10mM to about 100mM, about 15mM to about 50mM or about 20mM to about 30mM histidine. In certain embodiments, the composition comprises about 1mM, about 5mM, about 10mM, about 15mM, about 20mM, about 30mM, about 50mM, about 100mM, about 150mM or about 200mM histidine. In specific embodiments, the formulation comprises about 10mM to about 100mM histidine. In specific embodiments, the formulation comprises about 10 mM histidine.

[0106] In some embodiments, the composition comprises at least 1mM, at least 5mM, at least 10mM, at least 15mM, at least 20mM, at least 30mM, at least 50mM, at least 100mM, at least 150mM or at least 200mM histidine. In various embodiments, the composition comprises 1mM to 200mM, 5mM to 150mM, 10mM to 100mM, 15mM to 50mM or 20mM to 30mM histidine. In certain embodiments, the composition comprises 1mM, 5mM, 10mM, 15mM, 20mM, 30mM, 50mM, 100mM, 150mM or 200mM histidine. In a particular embodiment, the preparation comprises 10mM to 100mM histidine. In a specific embodiment, the preparation comprises 10mM histidine.

[0107] 1.2.7.1.pH

[0108] In some embodiments, the pH of the formulation is about 4.0 to about 8.0, such as 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 pH of the formulation is 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 specific embodiments, the pH of the formulation is about 5.0 to about 7.0. In specific embodiments, the pH of the formulation is about 5.0 to about 7.0. In certain embodiments, the pH of the formulation is about 5.5 to about 6.5. In certain embodiments, the pH of the formulation is about 5.7 to about 6.3. In specific embodiments, the pH of the formulation is about 6.0.

[0109] In some embodiments, the pH of the formulation is 4.0 to 8.0, such as 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 pH of the formulation is 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 specific embodiments, the pH of the formulation is 5.0 to 7.0. In certain embodiments, the pH of the formulation is 5.5 to 6.5. In certain embodiments, the pH of the formulation is 5.7 to 6.3. In a specific embodiment, the pH of the formulation is 6.0.

[0110] 1.2.8. Preservatives

[0111] In some embodiments, the formulation further comprises at least one preservative. In various embodiments, the at least one preservative is selected from octadecyl dimethyl benzyl ammonium chloride, hexamethonium chloride, benzalkonium chloride (a mixture of alkyl benzyl dimethyl ammonium chloride, wherein the alkyl group is a long-chain compound) and benzethonium chloride. Other types of preservatives include aromatic alcohols (such as phenol, butanol and benzyl alcohol), alkyl parabens (such as methyl paraben or propyl paraben), catechol, resorcinol, cyclohexanol, 3-pentanol and meta-cresol.

[0112] 1.2.9. Viscosity

[0113] In some embodiments, the viscosity of the formulation is less than 50 cP at 25° C., such as 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 specific 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.

[0114] 1.2.10. Aggregation and post-peak percentage

[0115] In some embodiments, the preparation reduces the aggregation of IL-6 protein. In certain embodiments, the preparation reduces the formation of soluble aggregates. In certain embodiments, the preparation reduces the formation of insoluble aggregates. In certain embodiments, compared with preparations with less polysorbate, the preparation reduces the formation of soluble aggregates, as measured by size exclusion high performance liquid chromatography (SEC-HPLC). In certain embodiments, compared with preparations with less polysorbate, the preparation reduces the formation of insoluble aggregates, as measured by visual appearance inspection. In certain embodiments, compared with preparations without histidine, glycine or DTPA, the preparation reduces the formation of soluble aggregates, as measured by RP-HPLC. In certain embodiments, the preparation has less than 15% soluble aggregates after stirring at 300rpm for 20 hours, as measured by RP-HPLC. In certain embodiments, the preparation has less than 10% soluble aggregates after stirring at 300rpm for 20 hours, as measured by RP-HPLC. In some embodiments, the formulation has less than 5% soluble aggregates after stirring at 300 rpm for 20 hours, as measured by RP-HPLC. In some embodiments, the formulation has less than 2% soluble aggregates after stirring at 300 rpm for 20 hours, as measured by RP-HPLC.

[0116] In some embodiments, after 14 days at -80°C, the post-peak % measured by reverse phase high performance liquid chromatography (RP-HPLC) is not significantly increased. In certain embodiments, after 14 days at -80°C, the post-peak percentage measured by RP-HPLC is about 3. In other embodiments, after 14 days at 2-8°C, the post-peak % measured by RP-HPLC data is not significantly increased. In certain embodiments, after 14 days at 2-8°C, the post-peak percentage measured by RP-HPLC is about 3.

[0117] 1.2.11. Effectiveness

[0118] 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.

[0119] In some embodiments, the biological activity (e.g., relative potency) of the preparation is reduced by less than 50% after storage at 5±3°C for 12 months, as measured by a reporter gene assay. In various embodiments, the biological activity of the preparation is reduced by less than 50%, less than 40%, less than 30%, less than 20%, or less than 10% after storage at 5±3°C for 12 months, as measured by RP-HPLC. In certain embodiments, the biological activity of the preparation is reduced by less than 30% after storage at 5±3°C for 12 months, as measured by RP-HPLC. In certain embodiments, the biological activity of the preparation is reduced by less than 20% after storage at 5±3°C for 12 months, as measured by RP-HPLC. In certain embodiments, the biological activity of the preparation is reduced by less than 10% after storage at 5±3°C for 12 months, as measured by RP-HPLC.

[0120] 1.3. Administration of the preparation

[0121] Suitable routes of administration of the IL-6 formulations described herein include, but are not limited to, parenteral (e.g., by 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.

[0122] In some embodiments, the formulation is suitable for administration in a single dose or in multiple doses.

[0123] 1.4. Dosage form

[0124] In another aspect, provided herein is a dosage form containing one or more unit doses of a pharmaceutical composition comprising IL-6.

[0125] In various embodiments, the dosage form is a prefilled syringe. In various embodiments, the dosage form is an automatic injection pen. In some embodiments, the dosage form is a device that uses automatic administration.

[0126] In various embodiments, the dosage form comprises one or more unit doses of the preparation as described above. In typical embodiments, the preparation is a liquid preparation. In other embodiments, the preparation is a dry preparation, including but not limited to lyophilized material. In a specific embodiment, the dosage form comprises a dry preparation and a measured amount of an aqueous diluent.

[0127] In some embodiments, the unit dosage form comprises a formulation comprising about 40 μg / mL to about 240 μg / mL IL-6, about 5 mM to about 15 mM histidine, about 5 mg / mL to about 15 mg / mL glycine, about 40 mg / mL to about 60 mg / mL trehalose, about 0.1 mg / mL to about 0.3 mg / mL polysorbate 20, and about 5 μM to about 100 μM DTPA, wherein the pH of the formulation is about 6.0 to about 8.0.

[0128] 1.5. Treatment

[0129] On the other hand, provided herein is a method for treating a disease or condition in a patient, the method comprising administering to the patient an IL-6 formulation as described herein. In some embodiments, the patient suffers from a disease or condition that can be treated or alleviated by administering IL-6, including, for example, chemotherapy-induced peripheral neuropathy (CIPN), weakness, or diabetic peripheral neuropathy (DPN).

[0130] Below is the example of the specific embodiment of 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 the accuracy of the numerals (e.g., amount, temperature, etc.) used, but of course some experimental errors and deviations should be allowed.

[0131] Example

[0132] Example 1 :Experimental Design Studies for Drug Substances

[0133] The IL-6 formulation (Drug Substance "DS" formulation) had 10 mM histidine, 10 mg / mL glycine, 50 mg / mL trehalose, 0.2 mg / mL polysorbate 20, and 50 μM DTPA at pH 7.0. A Design of Experiments (DOE) study was completed using the final DS formulation. A two-factor-two-level DOE study was performed; Factor 1 was pH 7.0 ± 0.5 range, and Factor 2 was protein concentration 5 ± 3 mg / mL. The details of the DOE combinations are mentioned in Table 1.

[0134] Table 1: DSDOE Combination

[0135]

[0136] Six different buffers of selected DS preparations were prepared. All buffers were prepared and the composition was recorded in subsequent Tables 3 to 7. The IB022 standard provided by Sonnet BioTherapeutics has a buffer component as 40mM phosphate and an excipient of 10mM NaCl as pH 7.0. In order to carry out DOE research, IB022 was buffer exchanged in six prepared buffers. After buffer exchange, the protein content and pH of the retentate sample were analyzed. The protein content and pH of the retentate sample were recorded in Table 8 (Sample ID). If necessary, the retentate was diluted to the target protein concentration based on the DOE factor (as mentioned in Table 2). All six preparations were filtered and filled in 5mL PETG bottles with a filling volume of 1.5mL. These six preparations were loaded to carry out a 14-day stability study at a temperature of 2-8°C (pressure) and -80°C (real time). The IV phase excipient screening plan is mentioned in Table 2 below.

[0137] Table 2: Phase IV DSDOE Plan

[0138]

[0139] Table 1: Composition of SON080F14a and SON080F14d

[0140]

[0141] Table 4: Composition of SON080F14b and SON080F14c

[0142]

[0143] Table 2: Composition of SON080F14e

[0144]

[0145] Table 3: Composition of SON080F14f

[0146]

[0147]

[0148] Table 4: Composition of reference buffer (40 mM phosphate + 10 mM NaCl, pH 7.0)

[0149]

[0150] Table 5: Retentate sample pH and protein content results

[0151]

[0152] Phase IV DSDOE samples were processed and diluted to their target DOE protein concentrations by the corresponding pH buffers. Filtering and DS filling were performed in 5 mL PETG, then loaded under pressure (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 osmotic pressure.

[0153] The purity of the DOE samples loaded for 14-day stability at -80°C and 2-8°C was checked by SEC-HPLC and is reported in Tables 9 and 10, respectively. The -80°C and 2-8°C data for all formulations were consistent up to 7 days. A 14-day analysis was performed and the results are not reported because the trend of the purity obtained by SEC-HPLC was not in line with expectations or deviated from the trend.

[0154] Table 9: Phase IV SEC-HPLC data for 14 days at -80°C

[0155]

[0156] Table 10: Phase IV SEC-HPLC data for 14 days at 2-8°C

[0157]

[0158]

[0159] The purity of the excipient screening samples charged for 14-day stability at -80°C and 2-8°C were checked by RP-HPLC and are recorded in Tables 11 and 12, respectively. The -80°C data for all formulations were consistent over 7 days, with only an increase in the peak % after 7 days of storage in formulation SON080F14d compared to IB022.

[0160] In SON080F14a, storage at 2-8°C was consistent up to 7 days, but in the other samples, an increase in the post-peak % and a decrease in the main peak % were observed when compared to IB022. Results for 14 days are not reported because the trend in purity obtained by SEC-HPLC did not meet expectations or deviate from the trend.

[0161] Table 11: Phase IV RP HPLC data at -80°C for 14 days

[0162]

[0163]

[0164] Table 12: Phase IV RP HPLC data for 14 days at 2-8°C

[0165]

[0166]

[0167] A DOE study was performed to investigate different pH and concentrations. Based on the 7-day data, no changes in quality attributes were observed over the pH range of 6.5 to 7.5, with a concentration range of 2 mg / mL to 8 mg / mL for a target pH of 7.0. The 14-day results were not as expected or out of trend and are therefore not considered for this report.

[0168] Embodiment 2: Drug Product (DP) Stability Studies

[0169] Three drug products of liquid and lyophilized IL-6 were manufactured to present 40 μg / mL, 120 μg / mL and 240 μg / mL and were packaged for 14-day pressure and real-time stability. Results for liquid and lyophilized DP stability up to 14 days were recorded for all three DP strengths. Due to some unexpected results, the 120 μg / mL liquid DP data was discontinued from 3 days and therefore not reported here.

[0170] For DP samples loaded for 14-day stability at 25°C (pressure, ST) and -20°C (real time, RT) for liquid formulations and at 40°C (ST) and 5°C (RT) for lyophilized formulations, protein concentrations of 40 μg / mL, 120 μg / mL and 240 μg / mL were measured by the RP-UPLC method. The results are recorded in Tables 13, 14 and 15, respectively.

[0171] The protein concentration of 40 μg / mL was consistent for up to 14 days for both formulations and was approximately 40 μg / mL at both RT and ST conditions. The protein concentration of 120 μg / mL was consistent for up to 14 days for both formulations and was approximately 120 μg / mL at both RT and ST conditions. ST did not report 120 μg / mL liquid DP data. For the liquid formulation, the protein concentration of 240 μg / mL remained consistent up to 14 days compared to day 0 at both RT and ST; and for the lyophilized formulation, the protein concentration remained consistent up to 14 days compared to day 0 at both RT and ST.

[0172] Table 13: Protein concentration (μg / mL) data for 40 μg / mL DP up to 14 days

[0173]

[0174] Table 14: Protein concentration (μg / mL) data for 120 μg / mL DP up to 14 days

[0175]

[0176] Table 15: Protein concentration (μg / mL) data for up to 14 days at 240 μg / mL DP

[0177]

[0178] The pH of 40 μg / mL, 120 μg / mL and 240 μg / mL DP samples loaded for 14-day stability at temperatures of 25°C (ST) and -20°C (RT) for liquid formulations and at temperatures of 40°C (ST) and 5°C (RT) for lyophilized formulations are recorded in Tables 16, 17 and 18, respectively. The target pH of the formulation was initially 7.0, and thus the DP stability of 40 μg / mL and 120 μg / mL was presented at pH 7.0. For 240 μg / mL, the pH was updated to 7.2, and thus a formulation of pH 7.2 was prepared.

[0179] For both conditions (RT and ST) and both formulations (liquid and lyophilized), the pH at 40 μg / mL and 120 μg / mL was consistent for up to 14 days and was approximately in the range of 6.8-7.3. The pH at 240 μg / mL was consistent for up to 14 days and was approximately 7.0-7.1 for both liquid and lyophilized formulations under both RT and ST conditions.

[0180] Table 16: pH data for 40 μg / mL DP up to 14 days

[0181]

[0182] Table 17: pH data for 120 μg / mL DP up to 14 days

[0183]

[0184] Table 18: Protein concentration (μg / mL) data for up to 14 days at 240 μg / mL DP

[0185]

[0186] When 40 μ g / mL, 120 μ g / mL and 240 μ g / mL of DP samples were loaded for 14-day stability at 25° C. (ST) and -20° C. (RT) for liquid formulations and at 40° C. (ST) and 5° C. (RT) for lyophilized formulations, the purity of the samples was measured by SEC-HPLC. The results are recorded in Tables 19, 20 and 21, respectively.

[0187] The purity of 40 μg / mL was consistent up to 14 days for both conditions (RT and ST) and both formulations (liquid and lyophilized), except for the increase in HMW% observed at 14 days under the pressure conditions of the lyophilized formulation. The purity of 120 μg / mL of the liquid formulation under RT conditions was consistent up to 14 days, while ST data were not reported. For the lyophilized formulation of 120 μg / mL up to 7 days, the purity data was consistent, but at day 14, both RT and ST data showed an increase in HMW%. The purity of 240 μg / mL was consistent up to 14 days for both conditions (RT and ST) and both formulations (liquid and lyophilized), and no changes were observed.

[0188] Table 19: SEC-HPLC data for 40 μg / mL DP up to 14 days

[0189]

[0190] (1)

[0192] Table 20: SEC-HPLC data for 120 μg / mL DP up to 14 days

[0193]

[0194] Table 21: SEC-HPLC data for 240 μg / mL DP up to 14 days

[0195]

[0196] When 40 μ g / mL, 120 μ g / mL and 240 μ g / mL of DP samples were loaded for 14-day stability at 25° C. (ST) and -20° C. (RT) for liquid formulations and at 40° C. (ST) and 5° C. (RT) for lyophilized formulations, the purity of the samples was measured by RP-HPLC. The results are recorded in Table 22, Table 23 and Table 2824, respectively.

[0197] For both conditions (RT and ST), for lyophilized preparations, the purity of 40 μg / mL was consistent for up to 14 days. In contrast, for both conditions (RT and ST) of 40 μg / mL, the pre-peak% and post-peak% were observed to increase in the liquid preparation after day 0. Under RT conditions, the purity of 120 μg / mL of the liquid preparation was consistent for up to 3 days, with the pre-peak% being found to decrease and the main peak% increasing on the 7th and 14th days. For 120 μg / mL liquid preparations, ST data were not reported. Under RT conditions, for the lyophilized preparations of 120 μg / mL, the pre-peak% was observed to decrease on the 7th and 14th days, while for ST conditions, the data were consistent for up to 14 days. For both conditions (RT and ST) and both preparations (liquid and lyophilized), the purity of 240 μg / mL was consistent for up to 14 days, with no changes observed.

[0198] Table 22: RP HPLC data for 40 μg / mL DP up to 14 days

[0199]

[0200] Table 23: RP HPLC data for 120 μg / mL DP up to 14 days

[0201]

[0202]

[0203] Table 24: RP HPLC data for 240 μg / mL DP up to 14 days

[0204]

[0205] The drug product formulations in liquid as well as lyophilized form were found to be stable under stress conditions for up to 14 days for lower and higher concentrations (40 μg / mL and 240 μg / mL). The results apply to all intermediate strengths.

[0206] Embodiment 3: Chelating agents are added to stabilize the formulation.

[0207] Subvisible particles in formulations may consist of aggregated proteins and may directly affect the efficacy and immunogenicity of the drug product. In addition, they often act as nucleation sites for further protein aggregation and / or lead to the development of larger particles by agglomeration (see Chapter 2.9.19. Particulate contamination: Subvisible particles have been published in the European Pharmacopoeia 33.2).

[0208] To evaluate the effect of chelating agents such as EDTA or DTPA on the stability of the formulation of Example 1, subvisible particle count analysis was performed using a HIAC9703+ liquid particle counting system at 37°C. The following formulations were tested: SON080F49a (without EDTA or DTPA), SON080F49b (with DTPA), and SON080F49c (with EDTA). The SON080F49a formulation contained 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 a pH of 7.2.

[0209]

[0210] The addition of chelators to the SON080F49a formulation reduced large subvisible particles of 2 μm, 5 μm, 10 μm, and 25 μM at days 1, 7, and 14. Furthermore, the formulation containing DTPA (SON080F49b) unexpectedly reduced subvisible particle counts compared to the formulation containing EDTA (SON080F49c).

[0211] Unless otherwise indicated, all numbers used in the specification and claims to indicate the amount of ingredients, properties (such as molecular weight), reaction conditions, etc. should be understood as being modified in all cases by the term "about". Therefore, unless otherwise indicated, the numerical parameters set forth in the specification and the appended claims are approximate values ​​that can be varied depending on the desired properties sought to be obtained by the present disclosure. At a minimum, and without attempting to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be interpreted in light of the number of reported significant digits and by applying ordinary rounding techniques.

[0212]

[0013] While the numerical ranges and parameters setting forth the broad scope of the present disclosure are approximations, the numerical values ​​set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0213] Unless otherwise specified herein or obviously contradictory to the context, the terms "a", "an", "the" and similar references used in the context of describing the present disclosure (particularly in the context of the following claims) should be interpreted as covering both the singular and the plural. The description of the range of values ​​herein is intended only to serve as a simplified method of referring to each individual value falling within the range individually. Unless otherwise specified herein, each individual value is incorporated into this specification as if each individual value is quoted individually in this specification. Unless otherwise specified herein or obviously contradictory to the context, all methods described herein can be performed in any suitable order. Unless otherwise stated, the use of any and all examples or exemplary language (e.g., "such as") provided herein is intended only to better illustrate the present disclosure without limiting the scope of the present disclosure. Any language in this specification should not be interpreted as indicating any unclaimed element as being essential to the practice of the present disclosure.

[0214] The grouping of alternative elements or embodiments of the present disclosure disclosed herein should not be construed as limiting. Each group member can be mentioned and claimed individually or in any combination with other members of the group or other elements found herein. For the purpose of convenience and / or patentability, it is expected that one or more members in a group may be included in the group or deleted from the group. When any such inclusion or deletion is performed, the specification is considered to contain the modified group, thereby satisfying the written description of all Markush groups used in the appended claims.

[0215] Certain embodiments of the present disclosure are described herein, including the best mode known to the inventor for implementing the present disclosure. Of course, by reading the foregoing description, the changes of these described embodiments will become clear to those of ordinary skill in the art. The inventor expects that the technician can adopt such variations when appropriate, and the inventor intends to practice the present disclosure in other ways different from the manner specifically described herein. Therefore, where applicable law permits, the present disclosure includes all modifications and equivalents of the subject matter recited in the attached claims. In addition, the present disclosure covers any combination of the above-mentioned elements in all possible variations thereof, unless otherwise indicated herein or otherwise clearly contradictory to the context.

[0216] Specific embodiments disclosed herein may be further limited in the claims using "consisting of" or "consisting essentially of" language. When used in the claims, whether as filed or added by amendment, the transitional term "consisting of" excludes any element, step, or ingredient not specified in the claim. The transitional phrase "consisting essentially of" limits the scope of the claim to the specified materials or steps and those that do not materially affect the basic and novel characteristics. The embodiments of the present disclosure so claimed are inherently or expressly described and enabled herein.

[0217] It should be understood that the embodiments of the present disclosure disclosed herein are illustrations of the principles of the present disclosure. Other modifications that may be adopted are within the scope of the present disclosure. Therefore, as an example and not limitation, alternative configurations of the present disclosure may be used according to the teachings herein. Therefore, the present disclosure is not limited to exactly as shown and described.

[0218] Although the present disclosure has been described and illustrated herein by reference to various specific materials, procedures and examples, it should be understood that the present disclosure is not limited to the specific combination of materials and procedures selected for this purpose. As will be appreciated by those skilled in the art, many variations of such details may be implicit therein. This specification and examples are intended to be considered as merely exemplary, and the actual scope and spirit of the present disclosure are 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. An IL-6 preparation comprising: - about 40 μg / mL to about 2 mg / mL IL-6; - about 5 mM to about 15 mM histidine; - from about 5 mg / mL to about 15 mg / mL glycine; - from about 40 mg / mL to about 60 mg / mL trehalose; - from about 0.1 mg / mL to about 0.3 mg / mL polysorbate 20; and - about 1 μM to about 1000 μM DTPA, wherein the pH of the formulation is from 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. The IL-6 preparation of claim 1, wherein the preparation 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, 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. IL-6, 140μg / 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, 195μg / mL IL-6, 200μg / mL IL-6, 205μg / mL IL-6, 210μg / mL IL-6, 215μg / mL IL-6, 220μg / mL IL-6, 225μg / mL IL-6, 230μg / mL IL-6, 235μg / mL IL-6 or 240 μg / mL IL-6. The IL-6 preparation according to claim 3 , wherein the preparation comprises 80 μg / mL IL-6.

5. The IL-6 preparation according to claim 3, wherein the preparation comprises 120 μg / mL IL-6. The IL-6 preparation according to claim 3 , wherein the preparation comprises 240 μg / mL IL-6.

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

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

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

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

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

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

0.

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

0.

14. The IL-6 preparation of claim 13, wherein the preparation 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, 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. IL-6, 140μg / 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 / mLIL-6, 190μg / mL IL-6, 195μg / mL IL-6, 200μg / mL IL-6, 205μg / mL IL-6, 210μg / mL IL-6, 215μg / mL IL-6, 220μg / mL IL-6, 225μg / mL IL-6, 230μg / mL IL-6, 235μg / mL IL-6 or 240 μg / mL IL-6.

15. The IL-6 preparation according to claim 14, wherein the preparation comprises 80 μg / mL IL-6.

16. The IL-6 preparation according to claim 14, wherein the preparation comprises 120 μg / mL IL-6.

17. The IL-6 preparation according to claim 14, wherein the preparation comprises 240 μg / mL IL-6.

18. The IL-6 preparation according to claim 13, wherein the preparation is suitable for parenteral administration.

19. The IL-6 preparation according to claim 13, wherein the preparation is suitable for intravenous administration.

20. The IL-6 preparation according to claim 13, wherein the preparation is suitable for subcutaneous administration.

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

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

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

Images