IL-23 antibody composition and method of use

Abstract

Variant IL-23 binding protein containing a modified Fc region and method of use. In one aspect, an IL-23 binding protein is provided, wherein the IL-23 binding protein specifically binds to an epitope of IL-23 and comprises an Fc region including amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS). In another aspect, a method for treating inflammatory bowel disease in a patient requiring treatment for inflammatory bowel disease is provided, the method comprising the step of subcutaneously or intravenously administering an effective amount of the IL-23 binding protein as disclosed herein to the patient.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims the benefit and priority of U.S. Provisional Patent Application No. 63 / 509,687, filed June 22, 2023, the entire contents of which are incorporated herein by reference. [Background technology]

[0002] background Interleukin-23 (IL-23) is a key inflammatory cytokine that promotes the differentiation of T helper 17 cells, leading to the production of other cytokines such as interleukin-17. IL-23 plays a role in inflammatory bowel disease because levels of IL-23 are elevated in the intestines of individuals with Crohn's disease, and disease severity in individuals with ulcerative colitis correlates with elevated levels of IL-23. Due to IL-23's importance in mediating pro-inflammatory responses, it is a therapeutic target for inflammatory bowel disease and highlights the need for the development of inhibitors that target IL-23. [Overview of the Initiative] [Means for solving the problem]

[0003] Summary of Disclosure This disclosure addresses this need with an IL-23 binding protein having a modified Fc region that extends the half-life of the IL-23 binding protein.

[0004] In one aspect, an IL-23 binding protein comprising: a) a complementarity-determining region (CDR) heavy chain variable region (VH) including (i) CDR-H1, (ii) CDR-H2, and (iii) CDR-H3; b) a light chain variable region (VL) including (i) CDR-L1, (ii) CDR-L2, and (iii) CDR-L3; and c) an Fc region including amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS), wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 are (i) SEQ ID NOs. 1, 8, 15, 22, 29, and 36, respectively; (ii) (iii) each of the amino acid sequences of SEQ ID NOs. 2, 9, 16, 23, 30, and 37, (iv) each of SEQ ID NOs. 3, 10, 17, 24, 31, and 38, (iv) each of SEQ ID NOs. 4, 11, 18, 25, 32, and 39, (v) each of SEQ ID NOs. 5, 12, 19, 26, 33, and 40, (vi) each of SEQ ID NOs. 6, 13, 20, 27, 34, and 41, or (vii) each of SEQ ID NOs. 7, 14, 21, 28, 35, and 42 are provided as IL-23 binding proteins.

[0005] In some embodiments, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs. 1, 8, 15, 22, 29, and 36, respectively. In some embodiments, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs. 2, 9, 16, 23, 30, and 37, respectively. In some embodiments, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs. 3, 10, 17, 24, 31, and 38, respectively. In some embodiments, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain amino acid sequences with sequence numbers 4, 11, 18, 25, 32, and 39, respectively. In some embodiments, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain amino acid sequences with sequence numbers 5, 12, 19, 26, 33, and 40, respectively. In some embodiments, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain amino acid sequences with sequence numbers 6, 13, 20, 27, 34, and 41, respectively. In some embodiments, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of sequence numbers 7, 14, 21, 28, 35, and 42, respectively.

[0006] In some embodiments, VH and VL each contain a sequence having at least 80% sequence identity with the amino acid sequences of (i) SEQ ID NOs: 127 and 134, (ii) SEQ ID NOs: 128 and 135, (iii) SEQ ID NOs: 129 and 136, (iv) SEQ ID NOs: 130 and 137, (v) SEQ ID NOs: 131 and 138, (vi) SEQ ID NOs: 132 and 139, or (vii) SEQ ID NOs: 133 and 140.

[0007] In some embodiments, VH includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 127, and VL includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 134. In some embodiments, VH includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 128, and VL includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 135. In some embodiments, VH includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 129, and VL includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 136. In some embodiments, VH includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 130, and VL includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 137. In some embodiments, VH includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 131, and VL includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 138. In some embodiments, VH includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 132, and VL includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 139. In some embodiments, VH includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 133, and VL includes a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 140.

[0008] In some embodiments, the Fc region is an IgG1, IgG2, or IgG4 immunoglobulin Fc region. In some embodiments, the Fc region is an IgG1 immunoglobulin region. In some embodiments, the Fc region is an IgG2 immunoglobulin region. In some embodiments, the Fc region is an IgG4 immunoglobulin region.

[0009] In one aspect, a) a heavy chain variable region (VH) comprising complementarity-determining regions (CDRs) (i) CDR-H1, (ii) CDR-H2, and (iii) CDR-H3; b) a light chain variable region (VL) comprising CDRs (i) CDR-L1, (ii) CDR-L2, and (iii) CDR-L3; and c) an IL-23 binding protein comprising a modified Fc region that extends the half-life of the IL-23 binding protein compared to an IL-23 binding protein that does not contain the modified Fc region, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 are (i) SEQ ID NOs. 1, 8, 15, 22, 29, and 36, respectively, (ii (iii) each contains the amino acid sequence of SEQ ID NOs: 2, 9, 16, 23, 30, and 37, (iv) each contains the amino acid sequence of SEQ ID NOs: 3, 10, 17, 24, 31, and 38, (iv) each contains the amino acid sequence of SEQ ID NOs: 4, 11, 18, 25, 32, and 39, (v) each contains the amino acid sequence of SEQ ID NOs: 5, 12, 19, 26, 33, and 40, (vi) each contains the amino acid sequence of SEQ ID NOs: 6, 13, 20, 27, 34, and 41, or (vii) each contains the amino acid sequence of SEQ ID NOs: 7, 14, 21, 28, 35, and 42.

[0010] In one aspect, an IL-23 binding protein is provided, wherein the IL-23 binding protein specifically binds to an epitope of IL-23 and includes an Fc region comprising amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS).

[0011] In one aspect, a method is provided for treating inflammatory bowel disease in a patient requiring treatment for inflammatory bowel disease, the method comprising the step of subcutaneously or intravenously administering an effective amount of IL-23 binding protein as disclosed herein to the patient.

[0012] In some embodiments, the inflammatory bowel disease is Crohn's disease or ulcerative colitis. In some embodiments, the inflammatory bowel disease is ulcerative colitis. In some embodiments, the inflammatory bowel disease is Crohn's disease.

[0013] In some embodiments, the step of administering comprises subcutaneous administration. In some embodiments, the step of administering comprises intravenous administration.

[0014] In one aspect, there is provided a pharmaceutical composition comprising a therapeutically effective amount of an IL-23 binding protein as disclosed herein and a pharmaceutically acceptable additive.

[0015] In one aspect, there is provided a method of treating an inflammatory disease in a patient who needs treatment for the inflammatory disease, the method comprising administering subcutaneously or intravenously to the patient an effective amount of an IL-23 binding protein or pharmaceutical composition as disclosed herein.

[0016] In some embodiments, the inflammatory disease is psoriasis. In some embodiments, the inflammatory disease is psoriatic arthritis. In some embodiments, the inflammatory disease is hidradenitis suppurativa.

[0017] In some embodiments, the step of administering comprises subcutaneous administration. In some embodiments, the step of administering comprises intravenous administration. Brief Description of the Drawings

Brief Description of the Drawings

[0018] [Figure 1] FIG. 1 is a graph of normalized serum concentration (y-axis) plotted against days post-injection (x-axis) for cynomolgus monkeys administered a single bolus dose (5 mg / kg) of an IL-23p19 binding protein with (「+YTE」) or without (「reference antibody」) an Fc YTE mutation by intravenous injection. See Example 4.

[0019] [Figure 2] Figure 2 is a graph of serum concentration in μg / mL (y-axis) plotted against the number of days after injection (x-axis) of cynomolgus monkeys administered a single bolus dose (5 mg / kg) of an IL-23p19 binding protein with or without the Fc YTE mutation (“+YTE” or “reference antibody”). See Example 4.

Mode for Carrying Out the Invention

[0020] Detailed Description Definitions To facilitate understanding of the present disclosure, many terms and phrases are defined below.

[0021] As used herein, all numerical values or numerical ranges include integers and values or fractions of integers within such ranges or encompassing such ranges, unless the context clearly indicates otherwise. Thus, for example, reference to a range of 90 to 100% includes 91%, 92%, 93%, 94%, 95%, 95%, 96%, 97%, etc., as well as 91.1%, 91.2%, 91.3%, 91.4%, 91.5%, etc., 92.1%, 92.2%, 92.3%, 92.4%, 92.5%, etc. In another example, reference to a range of​​​​​​​As used herein, the terms “about,” “approximately,” and “comparable to,” when used herein in reference to a value, mean a value similar to the value referred to in the context of that value. Generally, a person skilled in the art familiar with the context will recognize the degree of relevant variation encompassed by “about,” “approximately,” and “comparable to” in that context. For example, in some embodiments, the terms “about,” “approximately,” and “comparable to” may encompass a range of values ​​that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less than these percentages of the value referred to.

[0024] As used herein, unless otherwise indicated, the term “antibody” means an intact antibody (e.g., an intact monoclonal antibody), a fragment thereof (e.g., an Fc fragment of an antibody (e.g., an Fc fragment of a monoclonal antibody)), or an antigen-binding fragment of an antibody (e.g., an antigen-binding fragment of a monoclonal antibody), and is understood to include intact antibodies, antigen-binding fragments, or modified, manipulated, or chemically conjugated Fc fragments. Generally, antibodies are 4 It is a multimer protein containing two polypeptide chains. Two of the polypeptide chains are called immunoglobulin heavy chains (H chains), and two of the polypeptide chains are called immunoglobulin light chains (L chains). The immunoglobulin heavy and light chains are linked by interchain disulfide bonds. The immunoglobulin heavy chains are linked by interchain disulfide bonds. The light chains consist of one variable region (VL) and one constant region (CL). The heavy chains consist of one variable region (VH) and at least three constant regions (C H 1, C H 2 and C H3) consists of the above variable regions, which determine the binding specificity of the antibody. Each variable region contains three hypervariable regions known as complementarity-determining regions (CDRs), adjacent to four relatively conserved regions known as framework regions (FRs). The extents of the above FRs and CDRs are defined (Kabat, EA, et al. (1991) Sequences of Proteins of Immunological Interest, 5th edition, US Department of Health and Human Services, NIH Publication No. 91-3242; and Chothia, C. et al. (1987) J. Mol. Biol. 196:901-917). The three CDRs in each variable region (e.g., in a typical antibody form, there are light chain variable regions or heavy chain variable regions, for a total of six CDRs) are referred to as CDR1, CDR2, and CDR3, and collectively contribute to antibody binding specificity. Naturally occurring antibodies are used as starting materials for engineered antibodies (e.g., chimeric antibodies and humanized antibodies). Examples of antibody-based antigen-binding fragments include Fab, Fab', (Fab')2, Fv, single-chain antibodies (e.g., scFv), minibodies, and diabodies. Examples of modified or manipulated antibodies include chimeric antibodies, humanized antibodies, and multispecific antibodies (e.g., bispecific antibodies). An example of a chemically conjugated antibody is an antibody conjugated to a toxin moiety.

[0025] Antibody-dependent cell-mediated cytotoxicity (ADCC) refers to a form of cytotoxicity in which secreted immunoglobulins (Ig) bound to Fc receptors (FcRs) present on certain cytotoxic cells (e.g., natural killer (NK) cells, neutrophils, and macrophages) enable these cytotoxic effector cells to specifically bind to antigen-containing target cells and subsequently kill them with cytotoxins. Antibodies "arm" the cytotoxic cells and are absolutely necessary for such killing. NK cells, the main cells mediating ADCC, express only FcγRIII, while monocytes express FcγRI, FcγRII, and FcγRIII. FcR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, Annu. Rev. Immunol 9:457-92 (1991). To evaluate the ADCC activity of the target molecule, an in vitro ADCC assay (e.g., as described in U.S. Patent No. 5,500,362 or 5,821,337) may be performed. Useful effector cells for such assays include peripheral blood mononuclear cells (PBMCs) and natural killer (NK) cells. Alternatively, or in addition, the ADCC activity of the target molecule may be evaluated in vivo in animal models, such as those disclosed in Clynes et al. PNAS (USA) 95:652-656 (1998).

[0026] An antibody “antigen-binding fragment,” or “antibody fragment,” contains a portion of the intact antibody, a portion of which still retains the ability to bind to the antigen. In some embodiments, the antibody has a function in addition to its antigen-binding function, and the antigen-binding fragment retains that function. Typically, the antigen-binding fragment contains the variable region of the antibody. Papain digestion of the antibody yields two identical antigen-binding fragments (referred to as “Fab” fragments) and the remaining “Fc” fragment (a name reflecting its ability to readily crystallize). The Fab fragment contains a heavy chain (V HThe entire light chain along with the variable region domain of one heavy chain (C H 1) consists of each Fab fragment, which is monovalent with respect to antigen binding; that is, it has a single antigen-binding site. Pepsin treatment of the antibody produces a single large F(ab')2 fragment, which roughly corresponds to two disulfide-bonded Fab fragments having different antigen-binding activities and still cross-linking to the antigen. The Fab' fragment contains one or more cysteine ​​from the antibody hinge region described above, C H It differs from the Fab fragment by having several additional residues at the carboxyl terminus of one domain. Fab'-SH refers to Fab' in which the cysteine ​​residue of the constant domain has a free thiol group. The F(ab')2 antibody fragment was originally generated as a pair of Fab' fragments with a hinged cysteine ​​between them. Other chemical couplings of antibody fragments are also known.

[0027] As used herein, the term “chimeric antibody” refers to an antibody that has part of its heavy chain and / or light chain that is identical or homologous to a corresponding sequence in an antibody of a particular species or belonging to a particular antibody class or subclass, but the rest of its chain is identical or homologous to a corresponding sequence in an antibody of a different species or belonging to a different antibody class or subclass.

[0028] The "complementarity-determining region" (abbreviated as "CDR") is a highly variable region scattered within a more conserved region (called the "framework region" (abbreviated as "FR")). In some embodiments, the sequence of the framework region is identical to that of the framework region in the human germline sequence. In some embodiments, the sequence of the framework region is modified with respect to the human germline sequence.

[0029] As used herein, the terms “complement-dependent cell injury” or “CDC” refer to the lysis of target cells in the presence of complement. Activation of the classical complement pathway is initiated by the binding of the first component of the complement system (C1q) to antibodies (of the appropriate subclass) that bind to their congener antigens. To assess complement activation, a CDC assay (e.g., as described in Gazzano-Santoro et al., J. Immunol. Methods 202:163 (1996)) may be performed.

[0030] As used herein, antibody “effector function” refers to the biological activity that may be attributable to the Fc region of an antibody (either the native sequence Fc region or the amino acid sequence variant Fc region), which typically varies with antibody isotype. Examples of antibody effector functions include, but are not limited to, C1q binding and complement-dependent cytotoxicity, Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis, downregulation of cell surface receptors (e.g., B cell receptors), and B cell activation.

[0031] As used herein, the terms “effective dose” and “therapeutically effective dose” for a drug (e.g., IL-23 binding protein as described herein) are interchangeable and refer to an effective dose for the required duration and dosage to achieve the desired therapeutic effect. An effective dose may be administered in one or more doses, applications, or dosages and is not intended to be limited to a particular formulation or route of administration. As used herein, the term “to treat” includes any effect (e.g., to alleviate, reduce, regulate, improve, or eliminate) that results in improvement of a condition, disease, disorder, etc., or ameliorating its symptoms. An effective dose may vary depending on factors such as the type of disease of the individual (e.g., disease state), age, sex, and / or weight, as well as the ability of the binding protein (or its pharmaceutical composition) to induce the desired response in that individual. An effective dose may also be the amount in which the therapeutically beneficial effect outweighs any toxic or adverse effects of the binding protein or its pharmaceutical composition.

[0032] As used herein, the term “epitope” refers to an antigenic determinant that interacts with a specific antigen-binding site (known as a paratope) in the variable region of an antibody molecule (or binding protein) and is composed of six complementarity-determining regions of the antibody (or binding protein). A single antigen may have more than one epitope. Epitopes may be conformational or linear. Conformational epitopes consist of spatially juxtaposed amino acids derived from different segments of a linear polypeptide chain. Linear epitopes consist of adjacent amino acid residues in a polypeptide chain.

[0033] As used herein, the “Fc chain” of dimer Fc refers to one of the two polypeptides that form its dimer Fc region, i.e., the polypeptide that contains the C-terminal constant region of the immunoglobulin heavy chain and has the ability to stably associate with another similar polypeptide. For example, the Fc chain of dimer IgG Fc is IgG C H 2 and IgG C HIt contains three constant domain sequences. The Fc chain of the dimer Fc ("Fc region") can be one of various Ig classes, e.g., IgA, IgD, IgE, IgG, or IgM. These classes are also denoted as α, δ, ε, γ, and μ, respectively. Some of these may be further divided into subclasses (isotypes), e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2.

[0034] The terms “Fc receptor” and “FcR” are used to describe receptors that bind to the Fc region of an antibody. For example, an FcR may be a human FcR with a natural sequence. Generally, FcRs are those that bind to IgG antibodies (γ receptors) and include the FcγRI, FcγRII, and FcγRIII subclass receptors, including allele variants and alternatively spliced ​​forms of these receptors. FcγRII receptors include FcγRIIA ("activating receptor") and FcγRIIB ("inhibiting receptor"), which have similar amino acid sequences that differ primarily in their cytoplasmic domains. Other isotypes of immunoglobulins can also be bound by certain FcRs (see, e.g., Janeway et al., Immuno Biology: the immune system in health and disease, (Elsevier Science Ltd., NY) (4th edition, 1999)). The activating receptor FcγRIIA contains an immunoreceptor tyrosine-based activation motif (ITAM) in its cytoplasmic domain. The inhibitory receptor FcγRIIB contains an immunoreceptor tyrosine-based inhibition motif (ITIM) in its cytoplasmic domain (reviewed in Daeron, Annu. Rev. Immunol. 15:203-234 (1997)). FcRs are reviewed in Ravetch and Kinet, Annu. Rev. Immunol. 9:457-92 (1991); Capel et al., Immunomethods 4:25-34 (1994); and de Haas et al., J. Lab. Clin. Med. 126:330-41 (1995). Other FcRs (including those to be identified in the future) are encompassed by the term "FcR" as used herein.The term also includes FcRn, the neonatal receptor responsible for the transfer of maternal IgG to the fetus (Guyer et al., J. Immunol. 117:587 (1976); and Kim et al., J. Immunol. 24:249 (1994)).

[0035] As used herein, the term “humanized” refers, when used in reference to an antibody (or binding protein), to a form of non-human (e.g., mouse) antibody that is a chimeric antibody. A “humanized antibody” contains the smallest sequence derived from a non-human immunoglobulin. Typically, a humanized antibody is a human immunoglobulin (recipient or acceptor antibody) in which the hypervariable region residues of the recipient are replaced by hypervariable regions derived from a non-human species (donor antibody), such as mouse, rat, rabbit, or non-human primate, having the desired specificity, affinity, and capacity. In some cases, Fv framework region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues. Furthermore, humanized antibodies may contain residues not found in either the recipient or donor antibody. These modifications are made to further refine antibody performance, such as binding affinity. Generally, a humanized antibody comprises at least one, and typically substantially all, of two variable domains, where all or substantially all of the hypervariable loops correspond to those of a non-human immunoglobulin, and all or substantially all of the framework region is a human immunoglobulin sequence, the framework region may contain one or more amino acid substitutions to improve binding affinity. In some embodiments, six or fewer amino acid substitutions in the heavy chain and three or fewer amino acid substitutions in the light chain are used in the framework region. The humanized antibody also optionally comprises at least a portion of the immunoglobulin constant region (Fc), typically that of a human immunoglobulin.

[0036] "Percent (%) identity" refers to the degree to which two sequences (nucleotides or amino acids) have the same residues at the same positions in their alignment. For example, "the amino acid sequence is X% identical to sequence number Y" refers to the % identity of the amino acid sequence to sequence number Y, and is further detailed as X% of the residues in the amino acid sequence being identical to the residues in the sequence disclosed in sequence number Y. Generally, computer programs are employed for such calculations. Exemplary programs for comparing and aligning pairs of sequences include ALIGN (Myers and Miller, 1988), FASTA (Pearson and Lipman, 1988; Pearson, 1990), and gapped BLAST (Altschul et al., 1997), BLASTP, BLASTN, or GCG (Devereux et al., 1984).

[0037] As used herein, the term “pharmaceutical composition” refers to a combination of an active agent and a carrier, inert, or active agent that makes the composition particularly suitable for in vivo or ex vivo diagnostic or therapeutic use.

[0038] As used herein, the term “pharmaceutically acceptable carrier” refers to any of the standard pharmaceutically acceptable carriers (e.g., phosphate-buffered saline solutions, water, emulsions (e.g., oil / water or water / oil emulsions), and various types of wetting agents). The above compositions may also include stabilizers and preservatives. For examples of carriers, stabilizers, and adjuvants, see, for example, Martin, Remington's Pharmaceutical Sciences, 15th edition, Mack Publ. Co., Easton, PA (1975).

[0039] As used herein, “polypeptide” may be used interchangeably with “protein” and refers to a sequence of at least two amino acids linked to one another by a peptide bond. In some embodiments, a polypeptide may comprise at least 3 to 5 amino acids, each of which is linked to another by at least one peptide bond. Those skilled in the art will recognize that a polypeptide may comprise one or more “non-natural” amino acids or other entities (which may nevertheless be incorporated into the polypeptide chain). In some embodiments, a polypeptide may be glycosylated. For example, a polypeptide may comprise one or more covalently linked sugar moieties. In some embodiments, a single “polypeptide” (e.g., an antibody polypeptide) may comprise two or more individual polypeptide chains, which in some cases may be linked to one or more disulfide bonds or other means.

[0040] As used herein, the term “reference level” generally refers to a level considered “normal” for comparison purposes, e.g., a level of a suitable control. For example, in the context of the half-life of a protein (e.g., a binding protein) (e.g., serum half-life), the reference level may refer to the half-life of a “reference binding protein,” which may be, for example, an immunoglobulin of the same class (e.g., IgG1, IgG2, or IgG4) having a wild-type Fc region or an Fc region lacking a half-life-extending mutation. In some embodiments, the reference binding protein includes the same complementarity-determining region as that of the binding protein of interest (e.g., an IL-23 binding protein as described herein). In some embodiments, the reference binding protein includes the same variable regions (e.g., heavy chain and / or light chain variable regions) as that of the binding protein of interest (e.g., an IL-23 binding protein as described herein).

[0041] As used herein, the phrases "specifically binds to" or "selectively binds to" with respect to a target (e.g., IL-23) refer to a binding reaction in which, when referring to a binding protein as described herein, the binding protein binds to the target with a higher affinity, higher binding capacity, and / or longer duration than it binds to structurally different targets. In representative embodiments, the binding protein has at least 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 25-fold, 50-fold, 100-fold, 1,000-fold, 10,000-fold, or greater affinity for a specific target compared to an unrelated target when tested under the same affinity assay conditions. The terms "specific binding to", "specifically binds to", or "specific for" a particular target, as used herein, for example, the equilibrium dissociation constant K of the target to which a molecule binds d (e.g., 10 -5 M, 10 -6 M, 10 -7 M, 10 -8 M, 10 -9 M, 10 -10 M, 10 -11 M, or 10 -12 M, or less) can be indicated by that molecule. In some embodiments, the binding protein can specifically bind to an epitope on a target that is conserved across species (e.g., structurally conserved across species), such as between human and non-human primate species (e.g., structurally conserved between human and non-human primate species). In some embodiments, the binding protein can bind exclusively to a predetermined target, e.g., exclusively to IL-23, and may not bind to other interleukins.

[0042] The terms “subject,” “recipient,” “individual,” “host,” and “patient” are used interchangeably herein and, in some embodiments, refer to any mammalian subject, in particular humans, to whom diagnosis, treatment, or therapy is desired. “Mammal” with respect to the purpose of treatment refers to any animal classified as a mammal (including humans, domesticated animals and livestock, as well as laboratory animals, zoo animals, sports animals, or pet animals (e.g., dogs, horses, cats, cows, sheep, goats, pigs, mice, rats, rabbits, guinea pigs, monkeys, etc.)). In some embodiments, the mammal is human. None of these terms require the supervision of a medical professional.

[0043] As used herein, “treatment” of a condition or “treatment” of a condition (e.g., a condition as described herein) is an approach to obtain a beneficial or desired outcome (e.g., a clinical outcome). Beneficial or desired outcomes may include, but are not limited to, the alleviation or improvement of one or more symptoms or conditions, whether detectable or undetectable; a reduction in the severity of a disease, disorder, or condition; a stable (i.e., non-exacerbating) state of a disease, disorder, or condition; prevention of the spread of a disease, disorder, or condition; delay or slowing of the progression of the disease, disorder, or condition; palliation of the disease, disorder, or condition; and remission, whether partial or total. “Palliating” a disease, disorder, or condition means that the severity of the disease, disorder, or condition and / or its undesirable clinical manifestations are reduced and / or the time course of its progression is slowed or prolonged compared to the degree or course of the condition in the absence of treatment.

[0044] The terms “variable domain” and “variable region” are used interchangeably herein and refer to portions of the antibody (or binding protein) or immunoglobulin domains that exhibit variability in their sequence and are involved in determining the specificity and binding affinity of a particular antibody. The variability is not uniformly distributed throughout the variable domain of the antibody; it is concentrated in the respective subdomains of the heavy and light chain variable regions. These subdomains are referred to as “hypervariable regions” or “complementarity-determining regions” (CDRs). The more conserved (i.e., non-hypervariable) portions of the variable domain are referred to as “framework” regions (FRMs or FRs), which provide a scaffold for six CDRs in three-dimensional space to form an antigen-binding surface.

[0045] Throughout the detailed description, where a composition is described as having, including, or comprising certain components, or where a process and method is described as having, including, or comprising certain steps, it is intended that there exist compositions of the disclosure that are essentially composed of or consist of such described components, and that there exist processes and methods of the disclosure that are essentially composed of or consist of such described process steps.

[0046] As a general rule, unless otherwise specified, percentages of a composition are determined by weight. Furthermore, if a variable is not defined, its previous definition prevails.

[0047] IL-23 binding protein In one aspect, a protein is provided that can bind to IL-23 ("IL-23-binding protein") and contains a modified Fc region. In some embodiments, the IL-23-binding protein may bind to an epitope of human IL-23, for example, specifically. In some embodiments, the IL-23-binding protein may bind to an epitope that is specific to IL-23 (e.g., an epitope on the p19 subunit of IL-23 (IL-23A)). In some embodiments, the IL-23-binding protein may bind to an epitope shared with other molecules such as IL-12 (e.g., an epitope on the p40 subunit of IL-23 (IL-23B)).

[0048] In some embodiments, the IL-23 binding protein is an antibody or a fragment thereof. In some embodiments, the antibody or fragment thereof is a monoclonal antibody or a fragment thereof. In some embodiments, the antibody or fragment thereof is a chimeric antibody or a fragment thereof. In some embodiments, the antibody or fragment thereof is a humanized antibody or a fragment thereof. In some embodiments, the antibody or antigen-binding fragment thereof is a human antibody.

[0049] In certain embodiments, the IL-23 binding protein comprises a) a heavy chain variable region (VH) containing CDRH1, CDRH2, and CDRH3 of any of antibodies 1-7 (see Table 1); b) a light chain variable region (VL) containing CDRL1, CDRL2, and CDRL3 of the same antibodies from antibodies 1-7; and c) an Fc region containing amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS).

[0050] In a particular embodiment, the IL-23 binding protein includes a) a heavy chain variable region (VH) containing CDRH1, CDRH2, and CDRH3 of any of the antibodies 1-7 (see Table 1); b) a light chain variable region (VL) containing CDRL1, CDRL2, and CDRL3 of the same antibodies 1-7; and c) a modified Fc that extends the half-life of the IL-23 binding protein compared to an IL-23 binding protein that does not contain the modified Fc.

[0051] In certain embodiments, the IL-23 binding protein specifically binds to an epitope of IL-23 (e.g., an epitope on the p19 subunit of IL-23) and includes an Fc region comprising amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS).

[0052] The amino acid sequences of exemplary CDRs of the IL-23 binding proteins of this disclosure are provided in Table 1. Antibodies 1, 2, 3, 4, and 5 can bind to the p19 subunit of IL-23 ("IL-23p19"). Antibodies 6 and 7 can bind to the p40 subunit of IL-23 ("IL-23p40"). Table 1. CDR arrangement [Table 1-1] [Table 1-2] [Table 1-3]

[0053] The amino acid sequences of the exemplary heavy chain variable region (VH) and light chain variable region (VL) of exemplary IL-23 binding proteins are provided in Table 2. Table 2. Sequences of the heavy chain variable region (VH) and light chain variable region (VL) of exemplary IL-23 binding proteins. [Table 2-1] [Table 2-2]

[0054] In some embodiments, the IL-23 binding protein includes a heavy chain variable region (VH) containing an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 92.5%, at least 95%, at least 97.5%, or at least 99% sequence identity with the VH of any of the antibodies 1 to 7 (see Table 2); and a) a light chain variable region (VL) containing an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 92.5%, at least 95%, at least 97.5%, or at least 99% sequence identity with the VL of the same antibody 1 to 7.

[0055] FC modification The IL-23 binding proteins of this disclosure typically include a modified Fc region (as further described herein). The Fc region typically includes one or more Fc chains. The IgG Fc chain typically includes two constant weight domains (C H 2 and C H 3) and C above H It includes a hinge region connected to two domains. Typically, an Fc region may contain two Fc domains that dimerize with each other; however, an Fc region may have more than two Fc chains, as can be present in a single chain or, for example, in some antibody forms.

[0056] In some embodiments, the IL-23 binding protein includes an IgG1 Fc region (e.g., a human IgG1 Fc region). That is, the Fc region has an amino acid sequence substantially similar to that of the Fc region in wild-type IgG1 Fc, except that it has certain residues at certain positions as noted herein. In some embodiments, the wild-type IgG1 Fc is human IgG1 Fc, where each Fc chain has the amino acid sequence of SEQ ID NO: 141. In some embodiments, the IL-23 binding protein includes an Fc region, where each Fc chain has an amino acid sequence that is at least 85%, at least 87.5%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to that of the Fc chain in wild-type IgG1 Fc.

[0057] In some embodiments, the IL-23 binding protein includes an IgG2 Fc region (e.g., a human IgG2 Fc region). That is, the Fc region has an amino acid sequence substantially similar to that of the Fc region in wild-type IgG2 Fc, except that it has certain residues at certain positions as noted herein. In some embodiments, the wild-type IgG2 Fc is human IgG2 Fc, where each Fc chain has the amino acid sequence of SEQ ID NO: 143. In some embodiments, the IL-23 binding protein includes an Fc region, where each Fc chain has an amino acid sequence that is at least 85%, at least 87.5%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to that of the Fc chain in wild-type IgG2 Fc.

[0058] In some embodiments, the IL-23 binding protein includes an IgG4 Fc region (e.g., a human IgG4 Fc region). That is, the Fc region has an amino acid sequence substantially similar to that of the Fc region in wild-type IgG4 Fc, except that it has certain residues at certain positions as noted herein. In some embodiments, the wild-type IgG4 Fc is human IgG4 Fc, where each Fc chain has the amino acid sequence of SEQ ID NO: 142. In some embodiments, the IL-23 binding protein includes an Fc region, where each Fc chain has an amino acid sequence that is at least 85%, at least 87.5%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to that of the Fc chain in wild-type IgG4 Fc.

[0059] In certain embodiments, the Fc region is modified (e.g., substituted) by one or more amino acid residues. In certain embodiments, such modification alters the half-life of a molecule containing the Fc region (e.g., a binding protein) by altering (e.g., enhancing) its binding to an Fc receptor (e.g., neonatal Fc receptor (FcRn)).

[0060] Unless otherwise specified herein, the numbering of amino acid residues in the Fc region or constant region follows the EU numbering system (also known as the EU index), as described in Kabat et al., Sequences of Proteins of Immunological Interest, 5th edition. Public Health Service, National Institutes of Health, Bethesda, MD, 1991.

[0061] An example amino acid sequence of the Fc sequence is provided in Table 3. Table 3. Exemplary Fc sequences [Table 3-1] Table 3-2 Table 3-3 Table 3-4 Table 3-5 Table 3-6 Table 3-7 Table 3-8 Table 3-9 Table 3-10 Table 3-11 Table 3-12 Table 3-13 Table 3-14 Table 3-15 Table 3-16 Table 3-17 Table 3-18 Table 3-19

[0062] In some embodiments, the IL-23 binding protein includes an Fc region with one or more modifications in SEQ ID NO: 141. In some embodiments, the IL-23 binding protein includes an Fc region with one or more modifications in SEQ ID NO: 142. In some embodiments, the IL-23 binding protein includes an Fc region with one or more modifications in SEQ ID NO: 143. In some embodiments, the Fc region includes an amino acid sequence having at least 80% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 141 to 143. In some embodiments, the Fc region includes an amino acid sequence having at least 85% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 141 to 143. In some embodiments, the Fc region includes an amino acid sequence having at least 90% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 141 to 143. In some embodiments, the Fc region includes an amino acid sequence having at least 95% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 141 to 143. In some embodiments, the Fc region includes an amino acid sequence having at least 96% sequence identity with an amino acid sequence following any one of sequence numbers 141 to 143. In some embodiments, the Fc region includes an amino acid sequence having at least 97% sequence identity with an amino acid sequence following any one of sequence numbers 141 to 143. In some embodiments, the Fc region includes an amino acid sequence having at least 98% sequence identity with an amino acid sequence following any one of sequence numbers 141 to 143. In some embodiments, the Fc region includes an amino acid sequence having at least 99% sequence identity with an amino acid sequence following any one of sequence numbers 141 to 143. In some embodiments, the Fc region includes an amino acid sequence following any one of sequence numbers 141 to 143.

[0063] In some embodiments, one or more modifications in the modified Fc region are selected from the group consisting of: S298A, E333A, K334A, K326A, F243L, R292P, Y300L, V305I, P396L, F243L, R292P, Y300L, L235V, P396L, F243L, S239D, I332E, A330L, S267E, L328F, D265S, S239E, K326A, A327H, G237F, K326E, G236A, D270L, H268D, S324T, L234F, N325L, V266L, and S267D. In some embodiments, one or more modifications in the modified Fc region are selected from the group consisting of: S228P, M252Y, S254T, T256E, T256D, T250Q, H285D, T307A, T307Q, T307R, T307W, L309D, Q411H, Q311V, A378V, E380A, M428L, N434A, N434S, N297A, D265A, L234A, L235A, and N434W.

[0064] In some embodiments, the modified Fc region includes a specific combination of amino acid substitutions selected from the group consisting of: L234A / L235A; V234A / G237A; L235A / G237A / E318A; S228P / L236E; H268Q / V309L / A330S / A331S; C220S / C226S / C229S / P238S;C226S / C229S / E3233P / L235V / L235A;L234F / L235E / P331S;C226S / P230S;L234A / G237A;L234A / L235A / G237A;Q311R / M428L;and L234A / L235A / P329G.

[0065] In some embodiments, the modified Fc region comprises a specific combination of amino acid substitutions selected from the group consisting of: M428L / N434S (LS); M252Y / S254T / T256E (YTE); T250Q / M428L; T307A / E380A / N434A; T256D / T307Q (DQ); T256D / T307W (DW); M252Y / T256D (YD); T307Q / Q311V / A378V (QVV); T256D / H285D / T307R / Q311V / A378V (DDRVV); L309D / Q311H / N434S (DHS); S228P / L235E (SPLE); L234A / L235A (LALA); M428L / N434A (LA); L234A / G2LAGA / QVV, LALAGA / QVV;LALAPG / QVV;N297A / DDRVV;D265A / DDRVV;LALA / DDRVV;LAGA / DDRVV;LALAGA / DDRVV;LALAPG / DDRVV;SP / Q311R / M428L;SPLE / Q311R / M428L; N297A / Q311R / M428L; D265A / Q311R / M428L; LALA / Q311R / M428L; LAGA / Q311R / M428L; LALAGA / Q311R / M428L; and LALAPG / Q311R / M428L. In some embodiments, the modified Fc region includes a specific combination of amino acid substitutions selected from the group consisting of M428L / N434S(LS) and M252Y / S254T / T256E(YTE). In some embodiments, the modified Fc region includes the M428L / N434S(LS) modification (e.g., SEQ ID NO: 159, SEQ ID NO: 176, SEQ ID NO: 183). In some embodiments, the modified Fc region includes the M252Y / S254T / T256E(YTE) modification (e.g., SEQ ID NO: 152, SEQ ID NO: 173, SEQ ID NO: 182).

[0066] In some embodiments, the IL-23 binding protein includes modifications to improve its ability to mediate effector function. Such modifications are known in the art and include afucosylation or manipulation of the affinity of the Fc region to the activating receptor (primarily FCGR3a for antibody-dependent cell-mediated injury (ADCC)) and to C1q for complement-dependent cell-mediated injury (CDC).

[0067] In some aspects, the IL-23 binding protein described above contains an Fc region (e.g., IgG1 Fc region) with reduced fucose content at position Asn 297 (EU numbering) compared to naturally occurring Fc regions. Such Fc regions are known to improve ADCC. In some aspects, such antibodies contain no fucose at position Asn 297.

[0068] In some embodiments, the IL-23 binding protein includes an Fc region having one or more amino acid substitutions that improve ADCC (e.g., substitutions at one or more positions among positions 298, 333, and 334 of the Fc region). In some embodiments, the IL-23 binding protein includes an Fc region having one or more amino acid substitutions at positions 239, 332, and 330.

[0069] In some embodiments, the Fc region includes an amino acid sequence having at least 80% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 144-252. In some embodiments, the Fc region includes an amino acid sequence having at least 85% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 144-252. In some embodiments, the Fc region includes an amino acid sequence having at least 90% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 144-252. In some embodiments, the Fc region includes an amino acid sequence having at least 95% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 144-252. In some embodiments, the Fc region includes an amino acid sequence having at least 96% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 144-252. In some embodiments, the Fc region includes an amino acid sequence having at least 97% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 144-252. In some embodiments, the Fc region includes an amino acid sequence having at least 98% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 144 to 252. In some embodiments, the Fc region includes an amino acid sequence having at least 99% sequence identity with an amino acid sequence following any one of SEQ ID NOs: 144 to 252. In some embodiments, the Fc region includes an amino acid sequence following any one of SEQ ID NOs: 144 to 252.

[0070] In some embodiments, the IL-23-binding protein includes an Fc region in the oligosaccharide bound to the Fc region, having at least one galactose residue. Such antibody variants may have improved CDC function.

[0071] In some embodiments, the IL-23 binding protein includes one or more modifications that improve or reduce C1q binding and / or CDC.

[0072] In a particular embodiment, the Fc region comprises one or more amino acid substitutions, where the one or more substitutions result in an increase in one or more of the following compared to an Fc without the one or more substitutions: antibody half-life, ADCC activity, ADCP activity, or CDC activity. In a particular embodiment, the one or more amino acid substitutions result in an increased antibody half-life at pH 6.0 compared to a binding protein containing a wild-type Fc region. In a particular embodiment, the binding protein has an increased half-life that is approximately 10,000 times, 1,000 times, 500 times, 100 times, 50 times, 20 times, 10 times, 9 times, 8 times, 7 times, 6 times, 5 times, 4.5 times, 4 times, 3.5 times, 3 times, 2.5 times, 2 times, 1.95 times, 1.9 times, 1.85 times, 1.8 times, 1.75 times, 1.7 times, 1.65 times, 1.6 times, 1.55 times, 1.50 times, 1.45 times, 1.4 times, 1.35 times, 1.3 times, 1.25 times, 1.2 times, 1.15 times, 1.1 times, or 1.05 times longer than the binding protein containing the wild-type Fc region.

[0073] In a particular embodiment, the Fc region comprises one or more amino acid substitutions, where the one or more substitutions result in a decrease in one or more of the ADCC activity, ADCP activity, or CDC activity compared to an Fc without the one or more substitutions.

[0074] In a particular embodiment, the Fc region binds an Fcγ receptor selected from the group consisting of FcγRI, FcγRIIa, FcγRIIb, FcγRIIc, FcγRIIIa, and FcγRIIIb. In a particular embodiment, the Fc region binds the Fcγ receptor with higher affinity at pH 6.0 compared to binding proteins containing a wild-type Fc region.

[0075] In some embodiments, the IL-23 binding protein includes, for example, human serum or an extended half-life in humans (i.e., serum half-life). In some embodiments, the IL-23 binding protein includes a half-life of at least about 14 weeks, 28 weeks, 42 weeks, 56 weeks, 70 weeks, 84 weeks, 96 weeks, or longer than 96 weeks. In some embodiments, the IL-23-binding protein has a half-life in the range of approximately 14 to 96 days, approximately 14 to 84 days, approximately 14 to 70 days, approximately 14 to 56 days, approximately 14 to 42 days, approximately 14 to 28 days, approximately 28 to 96 days, approximately 28 to 84 days, approximately 28 to 70 days, approximately 28 to 56 days, approximately 28 to 42 days, approximately 42 to 96 days, approximately 42 to 84 days, approximately 42 to 70 days, or approximately 42 to 56 days. In some embodiments, the IL-23-binding protein has a half-life in the range of approximately 42 to 56 days. In some embodiments, the IL-23-binding protein has a half-life in the range of approximately 74 to 93 days. In some embodiments, the IL-23 binding protein has a half-life of at least about 50 days, at least about 55 days, at least about 60 days, at least about 65 days, at least about 70 days, at least about 75 days, at least about 80 days, at least about 85 days, at least about 90 days, or at least about 93 days. In some embodiments, the IL-23 binding protein has a half-life of about 50 days, about 55 days, about 60 days, about 65 days, about 70 days, about 75 days, about 80 days, about 85 days, about 90 days, or about 93 days. Methods for measuring the half-life are known in the art. In some embodiments, the half-life is measured in non-human primates. In some embodiments, the half-life is measured in humans. In some embodiments, the half-life is measured after intravenous administration. In some embodiments, the half-life is measured after subcutaneous administration.

[0076] In some embodiments, the IL-23 binding protein has a half-life at least 20% longer than that of the comparer antibody. In some embodiments, the comparer antibody contains the same complementarity-determining region and variable region, but contains a different Fc region. In some embodiments, the half-life of the IL-23 binding protein is at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% longer than the half-life of the comparer antibody. In some embodiments, the half-life of the IL-23 binding protein is at least 2 times, at least 3 times, at least 4 times, at least 5 times, at least 6 times, at least 7 times, at least 8 times, at least 9 times, or at least 10 times longer than the half-life of the comparer antibody.

[0077] Pharmaceutical composition This disclosure also encompasses methods for using a pharmaceutical composition containing a therapeutically effective amount of the above-mentioned IL-23 binding protein. The composition may be formulated for use in various drug delivery systems. One or more physiologically acceptable excipients or carriers may also be included in the composition for appropriate formulation. Suitable formulations for use in this disclosure can be found in Remington's Pharmaceutical Sciences, Mack Publishing Company, Philadelphia, Pa., 17th edition, 1985. For a brief review of methods for drug delivery, see, for example, Langer (Science 249:1527-1533, 1990).

[0078] In some embodiments, the pharmaceutical composition may include, for example, formulation materials for modifying, maintaining, or preserving the pH, volumetric osmolality, viscosity, clarity, color, isotonicity, odor, sterility, stability, dissolution rate or release rate, or adsorption or osmosis of the composition.In such embodiments, suitable formulation materials include, but are not limited to, the following: amino acids (e.g., glycine, glutamine, asparagine, arginine, or lysine); antimicrobial agents; antioxidants (e.g., ascorbic acid, sodium sulfite, or sodium bisulfite); buffers (e.g., borates, bicarbonates, Tris-HCl, citrates, phosphates, or other organic acids); bulking agents (e.g., mannitol or glycine); chelating agents (e.g., ethylenediaminetetraacetic acid (EDTA)); complexing agents (e.g., caffeine, polyvinylpyrrolidone, β-cyclodextrin, or hydroxypropyl-β-cyclodextrin); fillers; monosaccharides; dissaccharides; and other carbohydrates (e.g., glucose, mannose, or dextrin); proteins (e.g., serum albumin, gelatin, or immunoglobulin); colorants, flavoring agents, and diluents; emulsifiers; hydrophilic polymers (e.g., polyvinylpyrrolidone); Low molecular weight polypeptides; salt-forming counterions (e.g., sodium); preservatives (e.g., benzalkonium chloride, benzoic acid, salicylic acid, thimerosal, phenethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid, or hydrogen peroxide); solvents (e.g., glycerin, propylene glycol, or polyethylene glycol); sugar alcohols (e.g., mannitol or sorbitol); suspending agents; surfactants or wetting agents (e.g., Pluronic®, PEG, sorbitan esters, polysorbates (e.g., polysorbate 20, polysorbate), Triton®, tromethamine, lecithin, cholesterol, tyroxapole); stability enhancers (e.g., sucrose or sorbitol); tonicity enhancers (e.g., alkali metal halides, preferably sodium chloride or potassium chloride, mannitol, sorbitol); delivery vehicles; diluents; additives and / or pharmaceutical agents (Remington's Pharmaceutical Sciences, See the 18th edition (Mack Publishing Company, 1990).

[0079] In some embodiments, the pharmaceutical composition does not contain citrate.

[0080] In some embodiments, the pharmaceutical composition may include nanoparticles, such as polymer nanoparticles, liposomes, or micelles.

[0081] In some embodiments, the pharmaceutical composition may include sustained-release or controlled-release formulations. Techniques for formulating sustained-release or controlled-release means (e.g., liposome carriers, bio-erodible microparticles or porous beads and depot injections) are also known to those skilled in the art. Sustained-release preparations may include, for example, porous polymer microparticles or semipermeable polymer matrices in the form of molded articles (e.g., films) or microcapsules. Examples of sustained-release matrices include polyesters, hydrogels, polylactides, copolymers of L-glutamic acid and γ-ethyl-L-glutamate, poly(2-hydroxyethyl methacrylate), ethylene vinyl acetate, or poly-D(-)-3-hydroxybutyric acid. The sustained-release composition may also include liposomes, which may be prepared by any of several methods known in the art.

[0082] The pharmaceutical compositions comprising the IL-23 binding protein disclosed herein may be presented in dosage unit form and may be prepared by any suitable method. The pharmaceutical compositions should be formulated to be compatible with their intended route of administration. Examples of routes of administration include intravenous (IV), intradermal, inhalation, transdermal, topical, transmucosal, intrathecal, and rectal administration. In some embodiments, the IL-23 binding protein disclosed herein is administered intravenously or subcutaneously. In some embodiments, the IL-23 binding protein disclosed herein is administered intravenously. In some embodiments, the IL-23 binding protein disclosed herein is administered subcutaneously.

[0083] Useful formulations can be prepared by methods known in the field of pharmacy. See, for example, Remington's Pharmaceutical Sciences, 18th edition (Mack Publishing Company, 1990). Suitable formulation components for parenteral administration include sterile diluents (e.g., water for injection, saline solution, non-volatile oil, polyethylene glycol, glycerin, propylene glycol, or other synthetic solvents); antibacterial agents (e.g., benzyl alcohol or methylparaben); antioxidants (e.g., ascorbic acid or sodium bisulfite); chelating agents (e.g., EDTA); buffering agents (e.g., acetate, citrate, or phosphate); and agents for adjusting tonicity (e.g., sodium chloride or dextrose). In some embodiments, formulations for parenteral administration do not contain citrate.

[0084] For intravenous or subcutaneous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL™ (BASF, Parsippany, NJ), or phosphate-buffered saline (PBS). These carriers should be stable under manufacturing and storage conditions and protected from microorganisms. These carriers may be solvents or dispersion media containing, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, and liquid polyethylene glycol), and suitable mixtures thereof.

[0085] Intravenous or subcutaneous drug delivery formulations may be contained in syringes, pens, or bags. In some embodiments, the bag is connected to a channel including a tube and / or needle. In some embodiments, the formulation is a lyophilized formulation or a liquid formulation.

[0086] These compositions can be sterilized by conventional sterilization techniques or by sterile filtration. The resulting aqueous solutions can be packaged for immediate use or lyophilized, and the lyophilized preparations can be combined with a sterile aqueous carrier before administration.

[0087] A polyol (which may act as a tonicifier and stabilize the IL-23 binding protein) may also be included in the formulation. The polyol is added to the formulation in an amount that may vary with respect to the desired isotonicity of the formulation. In some embodiments, the aqueous formulation is isotonic. The amount of polyol added may also be varied with respect to the molecular weight of the polyol; for example, a small amount of monosaccharide (e.g., mannitol) may be added compared to a dissaccharide (e.g., trehalose). In some embodiments, the polyol used in the formulation as a tonicity agent is mannitol.

[0088] Detergents or surfactants may also be added to the formulation. Exemplary detergents include nonionic detergents (e.g., polysorbates (e.g., polysorbate 20, 80, etc.) or poloxamers (e.g., poloxamer 188)). The amount of detergent added is such that it reduces aggregation of the formulated antibody and / or minimizes the formation of particulate matter in the formulation and / or reduces adsorption. In some embodiments, the formulation may contain a surfactant that is a polysorbate. In some embodiments, the formulation may contain the detergent polysorbate 80 or Tween® 80. Tween® 80 is a term used to describe polyoxyethylene (20) sorbitan monooleate (see Fiedler, Lexikon der Hifsstoffe, Editio Cantor Verlag Aulendorf, 4th edition, 1996).

[0089] In some embodiments, the protein products of the present disclosure are formulated as liquid formulations. In some embodiments, the liquid formulations are prepared in combination with sugars to a level that provides stabilization. In some embodiments, the liquid formulations are prepared in an aqueous carrier. In some embodiments, the stabilizer is added in an amount less than or equal to the amount that would produce a viscosity undesirable or inappropriate for intravenous administration. In some embodiments, the sugar is a dissaccharide, for example, sucrose. In some embodiments, the liquid formulations may also contain one or more of buffers, surfactants, and preservatives.

[0090] In some embodiments, the pH of the liquid formulation is set by the addition of a pharmaceutically acceptable acid and / or base. In some embodiments, the pharmaceutically acceptable acid is hydrochloric acid. In some embodiments, the base is sodium hydroxide.

[0091] The aqueous carriers of interest described herein are pharmaceutically acceptable (safe and non-toxic for administration to humans) and are useful for the preparation of liquid formulations. Illustrative carriers include sterile water for injection (SWFI), bacteriostatic water for injection (BWFI), pH-buffered solutions (e.g., phosphate-buffered saline), sterile saline solutions, Ringer's solution, or dextrose solutions.

[0092] Preservatives may be added to the formulations herein as needed to reduce bacterial activity. The addition of preservatives may facilitate the production of multi-use (multiple-dose) formulations, for example.

[0093] The IL-23 binding protein described above can be freeze-dried to produce a freeze-dried formulation containing the protein and a cryoprotectant. The cryoprotectant may be a sugar, for example, a disaccharide. In some embodiments, the cryoprotectant is sucrose or maltose. The freeze-dried formulation may also contain one or more of a buffering agent, a surfactant, a filler, and / or a preservative.

[0094] The amount of sucrose or maltose useful for stabilizing the above lyophilized drug product can be at least 1:2 protein to sucrose or maltose by weight ratio. In some embodiments, the above protein to sucrose or maltose weight ratio is 1:2 to 1:5. In some embodiments, the pH of the above formulation is set by adding a pharmaceutically acceptable acid and / or base before lyophilization. In some embodiments, the pharmaceutically acceptable acid is hydrochloric acid. In some embodiments, the pharmaceutically acceptable base is sodium hydroxide.

[0095] Treatment method In a particular embodiment, a method for treating inflammatory bowel disease in a patient requiring treatment for inflammatory bowel disease is described herein, the method comprising the step of subcutaneously or intravenously administering an effective amount of an IL-23 binding protein containing a modified Fc region to the patient. In a particular embodiment, a method for treating inflammatory bowel disease in a patient requiring treatment for inflammatory bowel disease, the method comprising the step of subcutaneously or intravenously administering to the patient an effective amount of IL-23 binding protein as disclosed herein, for example, a) heavy chain variable region (VH) comprising complementarity-determining regions (CDRs) (i) CDR-H1, (ii) CDR-H2, and (iii) CDR-H3; b) light chain variable region (VL) comprising CDRs (i) CDR-L1, (ii) CDR-L2, and (iii) CDR-L3; and c) Fc region comprising amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS), wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 are (i) Sequence numbers 1, 8, 15, 22, 29, and 36, respectively (ii) Sequence numbers 2, 9, 16, 23, 30, and 37, respectively (iii) Sequence IDs 3, 10, 17, 24, 31, and 38, respectively (iv) Sequence numbers 4, 11, 18, 25, 32, and 39, respectively (v) Sequence numbers 5, 12, 19, 26, 33, and 40, respectively (vi) Sequence numbers 6, 13, 20, 27, 34, and 41, respectively (vii) Sequence numbers 7, 14, 21, 28, 35, and 42, respectively. A method comprising the amino acid sequence is described herein.

[0096] In a particular embodiment, a method for treating inflammatory bowel disease in a patient requiring treatment for inflammatory bowel disease, the method comprising an effective amount of IL-23 binding protein as disclosed herein, for example, a) heavy chain variable region (VH) comprising complementarity-determining regions (CDRs) (i) CDR-H1, (ii) CDR-H2, and (iii) CDR-H3; b) light chain variable region (VL) comprising CDR (i) CDR-L1, (ii) CDR-L2, and (iii) CDR-L3; and c) modified Fc that extends the half-life of the IL-23 binding protein compared to an IL-23 binding protein without modified Fc, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 are (i) Sequence numbers 1, 8, 15, 22, 29, and 36, respectively (ii) Sequence numbers 2, 9, 16, 23, 30, and 37, respectively (iii) Sequence IDs 3, 10, 17, 24, 31, and 38, respectively (iv) Sequence numbers 4, 11, 18, 25, 32, and 39, respectively (v) Sequence numbers 5, 12, 19, 26, 33, and 40, respectively (vi) Sequence numbers 6, 13, 20, 27, 34, and 41, respectively (vii) Sequence numbers 7, 14, 21, 28, 35, and 42, respectively. A method comprising the step of subcutaneously or intravenously administering an IL-23 binding protein containing the amino acid sequence to the patient is further described herein.

[0097] In a particular embodiment, a method for treating inflammatory bowel disease in a patient requiring treatment for inflammatory bowel disease, the method comprising the step of subcutaneously or intravenously administering an effective amount of IL-23-binding protein to the patient, wherein the IL-23-binding protein specifically binds to an epitope of IL-23 and comprises an Fc region including amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS), is further described herein.

[0098] In some embodiments, the inflammatory bowel disease is Crohn's disease or ulcerative colitis. In some embodiments, the inflammatory bowel disease is ulcerative colitis. In some embodiments, the inflammatory bowel disease is Crohn's disease.

[0099] In a particular embodiment, a method for treating an inflammatory disease in a patient requiring treatment for an inflammatory disease is described herein, the method comprising the step of subcutaneously or intravenously administering an effective amount of an IL-23 binding protein containing a modified Fc region to the patient. In a particular embodiment, a method for treating an inflammatory disease in a patient requiring treatment of the inflammatory disease, the method comprising an effective amount of IL-23 binding protein as disclosed herein, for example, a) a heavy chain variable region (VH) comprising complementarity-determining regions (CDRs) (i) CDR-H1, (ii) CDR-H2, and (iii) CDR-H3; b) a light chain variable region (VL) comprising CDRs (i) CDR-L1, (ii) CDR-L2, and (iii) CDR-L3; and c) an Fc region comprising amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS), wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 are (i) Sequence numbers 1, 8, 15, 22, 29, and 36, respectively (ii) Sequence numbers 2, 9, 16, 23, 30, and 37, respectively (iii) Sequence IDs 3, 10, 17, 24, 31, and 38, respectively (iv) Sequence numbers 4, 11, 18, 25, 32, and 39, respectively (v) Sequence numbers 5, 12, 19, 26, 33, and 40, respectively (vi) Sequence numbers 6, 13, 20, 27, 34, and 41, respectively (vii) Sequence numbers 7, 14, 21, 28, 35, and 42, respectively. A method comprising the step of subcutaneously or intravenously administering an IL-23 binding protein containing the amino acid sequence to the above-mentioned patient is described herein.

[0100] In a particular embodiment, a method for treating an inflammatory disease in a patient requiring treatment for an inflammatory disease, the method comprising an effective amount of IL-23 binding protein as disclosed herein, for example, a) heavy chain variable region (VH) comprising complementarity-determining regions (CDRs) (i) CDR-H1, (ii) CDR-H2, and (iii) CDR-H3; b) light chain variable region (VL) comprising CDR (i) CDR-L1, (ii) CDR-L2, and (iii) CDR-L3; and c) modified Fc that extends the half-life of the IL-23 binding protein compared to an IL-23 binding protein without modified Fc, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 are (i) Sequence numbers 1, 8, 15, 22, 29, and 36, respectively (ii) Sequence numbers 2, 9, 16, 23, 30, and 37, respectively (iii) Sequence IDs 3, 10, 17, 24, 31, and 38, respectively (iv) Sequence numbers 4, 11, 18, 25, 32, and 39, respectively (v) Sequence numbers 5, 12, 19, 26, 33, and 40, respectively (vi) Sequence numbers 6, 13, 20, 27, 34, and 41, respectively (vii) Sequence numbers 7, 14, 21, 28, 35, and 42, respectively. A method comprising the step of subcutaneously or intravenously administering an IL-23 binding protein containing the amino acid sequence to the patient is further described herein.

[0101] In a particular embodiment, a method for treating an inflammatory disease in a patient requiring treatment for the inflammatory disease, the method comprising the step of subcutaneously or intravenously administering an effective amount of IL-23-binding protein to the patient, wherein the IL-23-binding protein specifically binds to an epitope of IL-23 and comprises an Fc region including amino acid modifications M252Y, S254T, and T256E(YTE) and / or M428L and N434S(LS), is further described herein.

[0102] In some embodiments, the inflammatory disease is psoriasis. In some embodiments, the inflammatory disease is psoriatic arthritis. In some embodiments, the inflammatory disease is hidradenitis suppurativa.

[0103] In some embodiments, the IL-23 binding protein is administered in doses of approximately 75 mg to approximately 150 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 250 mg to approximately 750 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 300 mg to approximately 700 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 300 mg to approximately 600 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 300 mg to approximately 500 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 300 mg to approximately 400 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 400 mg to approximately 700 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 400 mg to approximately 600 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 300 mg to approximately 500 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 500 mg to approximately 700 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 500 mg to approximately 600 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 600 mg to approximately 700 mg. In some embodiments, the IL-23 binding protein is administered in doses of approximately 75 mg, approximately 100 mg, approximately 125 mg, approximately 150 mg, approximately 175 mg, approximately 200 mg, approximately 225 mg, approximately 250 mg, approximately 275 mg, approximately 300 mg, approximately 350 mg, approximately 400 mg, approximately 450 mg, approximately 500 mg, approximately 550 mg, approximately 600 mg, approximately 650 mg, or approximately 700 mg.

[0104] In some embodiments, the IL-23 binding protein is administered intravenously, intratumorally, intramuscularly, subcutaneously, intralesionally, intratestinally, intracolonally, intrarectally, intratrapouchely, or intraperitoneally. In some embodiments, the IL-23 binding protein is administered via parenteral routes (e.g., intravenously, intramuscularly, subcutaneously, intra-arterially, or intraperitoneally). In some embodiments, the IL-23 binding protein is administered intravenously or subcutaneously. In some embodiments, the IL-23 binding protein is administered intravenously. In some embodiments, the IL-23 binding protein is administered subcutaneously.

[0105] The administration of the above IL-23 binding protein may occur at various intervals. In some embodiments, the IL-23 binding protein is administered to the patient at least once at intervals longer than 8 weeks. In some embodiments, the interval is about 12 to about 26 weeks. In some embodiments, the interval is about 12 to about 22 weeks. In some embodiments, the interval is about 12 to about 18 weeks. In some embodiments, the interval is about 12 to about 14 weeks. In some embodiments, the interval is about 16 to about 26 weeks. In some embodiments, the interval is about 16 to about 22 weeks. In some embodiments, the interval is about 16 to about 18 weeks. In some embodiments, the interval is about 20 to about 26 weeks. In some embodiments, the interval is about 20 to about 22 weeks. In some embodiments, the interval is about 12 weeks. In some embodiments, the interval is about 16 weeks. In some embodiments, the interval is about 26 weeks.

[0106] Subject In certain embodiments, the subject to whom the IL-23 binding protein is administered in accordance with this disclosure is a mammal (e.g., a primate). In some embodiments, the subject is a human.

[0107] The subjects described above are those who have inflammatory bowel disease (e.g., Crohn's disease or ulcerative colitis) or inflammatory disease (e.g., psoriasis, psoriatic arthritis, or hidradenitis suppurativa), exhibit at least one symptom of the inflammatory bowel disease or inflammatory disease described above, are diagnosed with the inflammatory bowel disease or inflammatory disease described above, and / or may be identified as being at risk of the inflammatory bowel disease or inflammatory disease described above.

[0108] Psoriasis is a skin condition that causes itchy, scaly rashes, most commonly appearing on the knees, elbows, trunk, and scalp. The most common type of psoriasis, plaque psoriasis, causes dry, itchy, scaly, raised skin patches (plaques). These patches typically appear on the elbows, knees, hips, and scalp.

[0109] Psoriatic arthritis (PsA) refers to chronic inflammatory arthritis associated with psoriasis. Approximately 1 in 20 individuals with psoriasis develop arthritis along with psoriasis, and in about 75% of cases, psoriasis precedes arthritis. PsA itself manifests in various forms ranging from mild to severe arthritis, typically affecting the fingers and spine.

[0110] In other embodiments, the subject being considered may have inflammatory bowel disease, Crohn's disease, and / or ulcerative colitis, exhibit at least one symptom of them, have been diagnosed with them, and / or be identified as being at risk of them. For example, Crohn's disease can be a chronic, long-term disease that causes inflammation in the subject's GI duct and may affect any part of that GI duct from mouth to anus. Crohn's disease may be considered to belong to a larger group of conditions associated with inflammatory bowel disease or IBD. IBD affects the digestive system but can also cause problems in other parts of the body. Ulcerative colitis (another form of IBD) affects the large intestine (colon).

[0111] Route of administration In certain embodiments, the administration step includes systemic administration. In certain embodiments, systemic administration includes parenteral administration, such as intravenous, intra-arterial, intraperitoneal, subcutaneous, intramuscular, or intradermal administration. In some embodiments, systemic administration includes enteral administration, such as trans-gastroenteric administration or oral administration.

[0112] In some embodiments, the administration step includes intravenous administration. In some embodiments, the administration step includes subcutaneous administration.

[0113] In some embodiments, the administration step includes intramuscular, intralesional, intrasmall intestinal, intracolonic, intrarectal, or intravesical administration.

[0114] Outcome In many embodiments, the methods disclosed herein produce measurable improvements in the subjects, such as an improvement or cessation of symptoms. For example, such improvements may include improvements in clinical scores, or scores from surveys or questionnaires, that are relevant to or appropriate for the assessment of a disease or condition as referred herein (e.g., psoriasis or psoriatic arthritis), or inflammatory bowel disease (e.g., ulcerative colitis and / or Crohn's disease). For example, in some embodiments, reductions in the psoriasis area and severity index (PASI), such as a 50% or 75% reduction in the PASI score, are achieved.

[0115] Alternatively, or furthermore, improvements in the above scores or quality of life, as determined by any or any combination of the following instructions, may be achieved by the methods of this disclosure: the Dermatology Life Quality Index (DLQI), Investigator Global Assessment (IGA), Nail Psoriasis Severity Index (NAPSI), Physician Global Assessment (PGA), or Medical Outcomes Study Short Form (36) Health Survey (SF-36), or Treatment Satisfaction Questionnaire for Medication (TSQM).

[0116] In some embodiments, improvements in clinical scores or quality of life are achieved as measured by any or a combination of the following: the Composite Psoriatic Disease Activity Index (CPDAI), the PsA Disease Activity Score (PASDAS), the Group for Research and Assessment of Psoriasis and Psoriatic Arthritis (GRAPPA) Composite Exercise (GRACE) index, the Psoriatic Arthritis Impact of Disease (PsAID) questionnaire, or the Work Productivity and Activity Impairment (WPAI) questionnaire.

[0117] Preparation method The IL-23 binding proteins described above can be prepared using recombinant DNA techniques well known to those skilled in the art. For example, one or more isolated polynucleotides encoding the IL-23 binding proteins can be ligated to other suitable nucleotide sequences (including, for example, constant region coding sequences and expression regulatory sequences) to generate a conventional gene expression construct (i.e., an expression vector) encoding the desired IL-23 binding protein. The preparation of the defined gene construct is within the conventional art of the art.

[0118] Nucleic acids encoding a desired IL-23 binding protein can be incorporated (ligated) into an expression vector, which can then be introduced into host cells through conventional transfection or transformation techniques. Exemplary host cells include E. coli cells, Chinese hamster ovary (CHO) cells, human embryonic kidney 293 (HEK 293) cells, HeLa cells, baby hamster kidney (BHK) cells, monkey kidney cells (COS), human hepatocellular carcinoma cells (e.g., Hep G2), and melanoma cells that do not otherwise produce IgG proteins. Transformed host cells can be grown under conditions that allow the host cells to express the gene encoding the IL-23 binding protein.

[0119] Specific expression and purification conditions vary depending on the expression system used. For example, if a gene is to be expressed in E. coli, it is first cloned into an expression vector by positioning the engineered gene downstream from a suitable bacterial promoter (e.g., Trp or Tac) and a prokaryotic signaling sequence. The expressed protein may be secreted. The expressed protein may accumulate in refractile bodies or inclusion bodies, which can be collected after the disruption of the cells by French press or sonication. The refractile bodies are then solubilized, and the protein can be refolded and / or cleaved by methods known in the art.

[0120] If the manipulated gene is to be expressed in a eukaryotic host cell (e.g., a CHO cell), it is first inserted into an expression vector containing a suitable eukaryotic promoter, secretory signal, poly(A) sequence, and stop codon. Optionally, the vector or gene construct may also contain enhancers and introns. In embodiments involving a fusion protein containing an IL-23 binding protein or a portion thereof, the expression vector optionally includes a sequence encoding all or part of the constant region, enabling the expression of all or part of the heavy or light chain. The gene construct can be introduced into a eukaryotic host cell using conventional techniques.

[0121] In some embodiments, an N-terminal signal sequence is included in the protein construct to express an IL-23 binding protein. Exemplary N-terminal signal sequences include those derived from interleukin-2, CD-5, IgG κ light chain, trypsinogen, serum albumin, and prolactin.

[0122] Following transfection, a single clone can be isolated for cell banking using methods known in the art (e.g., limiting dilution, ELISA, FACS, microscopy, or Clonepix). The clone can be cultured under conditions suitable for bioreactor scale-up, and the expression of the IL-23 binding protein can be maintained.

[0123] The above IL-23 binding proteins can be isolated and purified using methods known in the art (including centrifugation, depth filtration, cell lysis, homogenization, freeze-thaw cycles, affinity purification, gel filtration, ion exchange chromatography, hydrophobic interaction exchange chromatography, and mixed-mode chromatography). [Examples]

[0124] Examples Example 1: Determination of antibody affinity for IL-23 The binding affinity (KD) of antibodies against human IL-23 (hIL-23) (antibodies 1 with or without the M252Y, S254T, and T256E(YTE) mutations) was determined using BIACORE. TM The determination was made via surface plasmon resonance (SPR) using an 8K SPR system (Cytiva). Series S Sensor Chip CM5 (Cytiva) sensor chips were prepared by immobilizing goat anti-human IgG (Jackson ImmunoResearch) and were ultimately used to determine the binding kinetic rate and affinity constant in HBS-P+ electrophoresis buffer (10 mM HEPES pH 7.4, 150 mM NaCl, 0.05% surfactant P20) at 25°C. The anti-IL23 mAb construct (diluted to 2 μg / mL) was captured on flow cell 2 (active) for 60 seconds at a flow rate of 10 μL / min. Recombinant human IL-23 was prepared at concentrations of 0 nM, 1.23 nM, 3.7 nM, 11.11 nM, 33.33 nM, and 100 nM, and injected into flow cell 1 (reference) and flow cell 2 (active) at a flow rate of 50 μL / min over 180 seconds. The samples were then injected into freshly captured mAbs in a multi-cycle manner by regenerating the capture surface with 10 mM glycine (pH 1.5) injected at a flow rate of 30 μL / min over 30 seconds. The apparent binding rate constant (ka) and dissociation rate constant (kd) were determined using a kinetic binding model. Their ratio provides the apparent equilibrium dissociation constant or affinity constant (KD = kd / ka). The results are shown in Table 4.

[0125] Under the tested conditions, the kd (and therefore KD) of antibody 1 and antibody 1 with the YTE mutation was found in BIACORE TM The detection limit of 8K SPR is 1 × 10⁻⁶ -6 The dissociation rate was 1 / s (both antibodies tested had a dissociation rate below this limit), so a conclusion could not be reached. Table 4. Antibody affinity for IL-23 [Table 4]

[0126] Example 2: Inhibition of IL-23-induced activation of STAT3 in HEK 293F IL-23R+ / IL-12Rβ1+ STAT3 luciferase reporter cells. The functional activity of antibodies blocking IL-23-induced biological activity (antibody 1 with or without the YTE mutation) was evaluated by inhibiting STAT3 activation in HEK 293F cells expressing IL-23R, IL-12Rβ1, and STAT3-linked luciferase reporters. Briefly, reporter cells were seeded into 96-well plates. A mixture of hIL-23 and purified IL-23 antibody was associated at room temperature for 30 minutes before being added to the cells (resulting in final concentrations of 20 ng / mL of hIL-23 and 0–50 nM of antibody). Cells were incubated at 37°C for 5 hours and subsequently lysed in One-Glo luciferase assay buffer (Promega) at room temperature for 10 minutes under light-shielding conditions. Luminescence was quantified using a SpectraMax® M5e plate reader (Molecular Devices), and subsequent data were analyzed using GraphPad Prism. IC 50 The value was determined as the antibody concentration required to inhibit the maximum luminescence signal detected in a 20 ng / mL hIL-23-only incubation by 50%.

[0127] As shown in Table 5, antibody 1 and antibody 1 YTE exhibit comparable inhibition of IL-23-induced activation of STAT3. Table 5. Inhibition of IL-23-induced STAT3 activation [Table 5] Value relative to antibody 1, normalized to 1.000.

[0128] Example 3: Inhibition of IL-23-induced phosphorylation of STAT3 in DB cells The functional activity of antibodies that block IL-23-induced biological activity (antibody 1 with or without the YTE mutation) was evaluated using inhibition of STAT3 phosphorylation in DB cells. In short, 6 × 10 5 DB cells were seeded in TC-treated round-bottom 96-well plates. A mixture of hIL-23 and purified IL-23 antibody was allowed to associate at room temperature for 1 hour before adding to the cells (resulting in final concentrations of 25 ng / mL of hIL-23 and 0–10 nM of antibody). Cells were incubated at 37°C for 1 hour, and phosphorylated STAT3 (pSTAT3) was quantified using the pSTAT3 Sandwich ELISA kit (Cell Signaling Technology). Subsequent data were analyzed using GraphPad Prism. IC 50 The value was determined as the antibody concentration required for 50% inhibition of the maximum pSTAT3 level detected in an incubation of hIL-23 alone at 25 ng / mL.

[0129] As shown in Table 6, antibody 1 and antibody 1 YTE exhibit comparable inhibition of IL-23-induced phosphorylation of STAT3. Table 6. Inhibition of IL-23-induced STAT3 phosphorylation [Table 6] Value relative to antibody 1, normalized to 1.000.

[0130] Example 4. Pharmacokinetic (PK) analysis of IL-23p19-binding protein in cynomolgus monkeys To evaluate the effect of half-life-extending mutations on the pharmacokinetics (PK) of IL-23p19-binding protein, male cynomolgus monkeys (Macaca fascicularis) weighing between 2.4 and 3.4 kg were administered a single bolus dose (5 mg / kg) of IL-23p19-binding protein (Antibody 1; see sequences in Tables 1 and 2) containing the Fc LALA mutation and with and without the Fc YTE mutation (see SEQ ID NOs. 148 and 152 in Table 3, respectively) on day 0, either intravenously (IV) and / or subcutaneously (SC). Serum samples were collected periodically throughout the study.

[0131] The half-lives were determined from cynomolgus monkey serum samples for each dose cohort up to day 56, and the mean PK curves are shown in Figure 1 for IV administration and in Figure 2 for SC administration. The β-elimination half-lives of the IL-23p19-binding protein with the Fc YTE mutation were observed to be 30 days and 24 days for IV administration and SC administration, respectively. In contrast, the observed half-lives of the reference antibody (without the Fc YTE mutation) were 11 days and 10 days for IV administration and SC administration, respectively.

[0132] From scaling the half-life observed in cynomolgus monkeys of IL-23p19-binding protein with YTE modification, a human half-life of 74–93 days was estimated.

[0133] Therefore, this example demonstrates the enhanced half-life of an IL-23p19-binding protein having a half-life extension mutation in the Fc region.

[0134] Equal parts This disclosure can be embodied in other specific forms without departing from its intent or essential features. The embodiments described herein should therefore be considered illustrative rather than limiting in all respects the disclosure described herein. Accordingly, the scope of the disclosure is indicated not by the foregoing description but by the appended claims, and all modifications that fall within the meaning and equivalent scope of the claims are intended to be encompassed within the claims.

Claims

1. a) Complementarity-determining region (CDR) (i) CDR-H1, (ii) CDR-H2, and (iii) CDR-H3, including heavy chain variable region (VH); and b) CDR (i) CDR-L1, (ii) CDR-L2, and (iii) CDR-L3 including a light chain variable region (VL); and c) Fc region including amino acid modifications M252Y, S254T, and T256E (YTE) and / or M428L and N434S (LS) An IL-23 binding protein containing, Here, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 are defined as follows: (i) Sequence numbers 1, 8, 15, 22, 29, and 36, respectively (ii) Sequence numbers 2, 9, 16, 23, 30, and 37, respectively (iii) Sequence numbers 3, 10, 17, 24, 31, and 38, respectively (iv) Sequence numbers 4, 11, 18, 25, 32, and 39, respectively (v) Sequence IDs 5, 12, 19, 26, 33, and 40, respectively (vi) Sequence IDs 6, 13, 20, 27, 34, and 41, respectively, or (vii) Sequence numbers 7, 14, 21, 28, 35, and 42, respectively An IL-23 binding protein containing the following amino acid sequence.

2. The IL-23 binding protein according to claim 1, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs: 1, 8, 15, 22, 29, and 36, respectively.

3. The IL-23 binding protein according to claim 1, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs: 2, 9, 16, 23, 30, and 37, respectively.

4. The IL-23 binding protein according to claim 1, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs: 3, 10, 17, 24, 31, and 38, respectively.

5. The IL-23 binding protein according to claim 1, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs: 4, 11, 18, 25, 32, and 39, respectively.

6. The IL-23 binding protein according to claim 1, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs. 5, 12, 19, 26, 33, and 40, respectively.

7. The IL-23 binding protein according to claim 1, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs: 6, 13, 20, 27, 34, and 41, respectively.

8. The IL-23 binding protein according to claim 1, wherein CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 each contain the amino acid sequences of SEQ ID NOs: 7, 14, 21, 28, 35, and 42, respectively.

9. The aforementioned VH and VL are, (i) Sequence IDs 127 and 134, respectively (ii) Sequence IDs 128 and 135, respectively (iii) Sequence numbers 129 and 136, respectively (iv) Sequence IDs 130 and 137, respectively (v) Sequence IDs 131 and 138, respectively (vi) Sequence IDs 132 and 139, respectively, or (vii) Sequence IDs 133 and 140, respectively A sequence having at least 80% sequence identity with the amino acid sequence of, The IL-23 binding protein according to claim 1.

10. The IL-23 binding protein according to claim 9, wherein VH comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 127, and VL comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO:

134.

11. The IL-23 binding protein according to claim 9, wherein VH comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 128, and VL comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO:

135.

12. The IL-23 binding protein according to claim 9, wherein VH comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 129, and VL comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO:

136.

13. The IL-23 binding protein according to claim 9, wherein VH comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 130, and VL comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO:

137.

14. The IL-23 binding protein according to claim 9, wherein VH comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 131, and VL comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO:

138.

15. The IL-23 binding protein according to claim 9, wherein VH comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 132, and VL comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO:

139.

16. The IL-23 binding protein according to claim 9, wherein VH comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO: 133, and VL comprises a sequence having at least 80% sequence identity with the amino acid sequence of SEQ ID NO:

140.

17. The IL-23 binding protein according to any one of claims 1 to 16, wherein the Fc region is an IgG1, IgG2, or IgG4 immunoglobulin Fc region.

18. The IL-23 binding protein according to claim 17, wherein the Fc region is an IgG1 immunoglobulin region.

19. The IL-23 binding protein according to claim 17, wherein the Fc region is an IgG2 immunoglobulin region.

20. The IL-23 binding protein according to claim 17, wherein the Fc region is an IgG4 immunoglobulin region.

21. a) Complementarity-determining regions (CDRs): (i) CDR-H1, (ii) CDR-H2, and (iii) CDR-H3, including heavy-chain variable regions (VHs); b) CDR (i) CDR-L1, (ii) CDR-L2, and (iii) CDR-L3 including a light chain variable region (VL); and c) A modified Fc region that extends the half-life of the IL-23 binding protein compared to an IL-23 binding protein that does not contain the modified Fc region. An IL-23 binding protein containing, Here, CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 are defined as follows: (i) Sequence numbers 1, 8, 15, 22, 29, and 36, respectively (ii) Sequence numbers 2, 9, 16, 23, 30, and 37, respectively (iii) Sequence numbers 3, 10, 17, 24, 31, and 38, respectively (iv) Sequence numbers 4, 11, 18, 25, 32, and 39, respectively (v) Sequence IDs 5, 12, 19, 26, 33, and 40, respectively (vi) Sequence IDs 6, 13, 20, 27, 34, and 41, respectively, or (vii) Sequence numbers 7, 14, 21, 28, 35, and 42, respectively An IL-23 binding protein containing the following amino acid sequence.

22. An IL-23 binding protein, wherein the IL-23 binding protein specifically binds to an epitope of IL-23 and comprises an Fc region including amino acid modifications M252Y, S254T, and T256E (YTE) and / or M428L and N434S (LS).

23. A method for treating inflammatory bowel disease in a patient requiring treatment for inflammatory bowel disease, the method comprising the step of subcutaneously or intravenously administering an effective amount of the IL-23 binding protein described in any one of claims 1 to 22 to the patient.

24. The method according to claim 23, wherein the inflammatory bowel disease is Crohn's disease or ulcerative colitis.

25. The method according to claim 24, wherein the inflammatory bowel disease is ulcerative colitis.

26. The method according to claim 24, wherein the inflammatory bowel disease is Crohn's disease.

27. The method according to any one of claims 23 to 26, wherein the administration step includes subcutaneous administration.

28. The method according to any one of claims 23 to 26, wherein the administration step includes intravenous administration.

29. A pharmaceutical composition comprising a therapeutically effective amount of the IL-23 binding protein according to any one of claims 1 to 28 and a pharmaceutically acceptable additive.

30. A method for treating an inflammatory disease in a patient requiring treatment for the inflammatory disease, the method comprising the step of subcutaneously or intravenously administering to the patient an effective amount of the IL-23 binding protein according to any one of claims 1 to 22 or the pharmaceutical composition according to claim 29.

31. The method according to claim 30, wherein the inflammatory disease is psoriasis.

32. The method according to claim 30, wherein the inflammatory disease is psoriatic arthritis.

33. The method according to claim 30, wherein the inflammatory disease is hidradenitis suppurativa.

34. The method according to any one of claims 30 to 33, wherein the administration step includes subcutaneous administration.

35. The method according to any one of claims 30 to 33, wherein the administration step includes intravenous administration.

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