Anti-stem cell factor antibodies and methods of use thereof

Abstract

To provide anti-stem cell factor antibodies and methods of use thereof.SOLUTION: The disclosure relates to antibodies and antigen-binding fragments thereof that bind to Stem Cell Factor (SCF). The antibodies and antigen-binding fragments thereof specifically bind to SCF248. The disclosure further relates to methods for making the antibodies, and to methods of use of the antibodies including methods of treatment for inflammatory and / or fibrotic diseases and disorders. In some embodiments, the antibody or fragment thereof comprise a heavy chain variable region amino acid sequence set forth in SEQ ID NO:7 and a light chain variable region amino acid sequence set forth in SEQ ID NO:16.SELECTED DRAWING: None

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Application No. 62 / 900,927, filed September 16, 2019, the entire contents of which are incorporated herein by reference.

[0002] Field The present invention relates to antibodies and antigen-binding fragments thereof that bind to stem cell factor (SCF) and specific sites thereof, and methods of using such antibodies and antigen-binding fragments.

[0003] Instructions for electronically submitted text files The contents of the text file submitted electronically herewith, i.e., a computer-readable format copy of the Sequence Listing (file name: OPSL_001_01WO_SeqList_ST25; date of recording: September 16, 2020; file size: 58 kb), are incorporated herein by reference in their entirety. [Background technology]

[0004] Inflammatory diseases are a leading cause of morbidity and mortality worldwide. Some types of chronic inflammation can lead to fibrosis, which is the formation or development of excess fibrous connective tissue in an organ or tissue as a reparative or reactive process, distinct from the formation of fibrous tissue as a normal component of the organ or tissue. Chronic inflammation and fibrosis affect nearly every tissue and organ system, and fibrous tissue remodeling can influence cancer metastasis and accelerate chronic graft rejection in transplant patients.

[0005] Stem cell factor (SCF) and its receptor c-Kit are important factors in the maintenance of chronic inflammation and fibrotic diseases (El-Koraie, et al., Kidney Int. 60:167 (2001); Powell, et al., Am. J. Physiol. 289:G2 (2005); El Kossi, et al., Am. J. Kidney Dis. 41:785 (2003); Powell, et al., Am. J. Physiol. 277:C183 (1999); Ding et al. J Pathol. 2013 Jun;230(2):205-14.; Berlin et al. Lab Invest. 2006 Jun;86(6):557-65; Rasky et al. Am J Physiol Lung Cell Mol Physiol. 2020 Jan 1;318(1):L200-L211). c-Kit is a type III receptor tyrosine kinase present in many cell types (Orr-Urtreger et al., Development 109:911 (1990)). Immune cells, such as mast cells, eosinophils, and innate lymphoid cell types 2 and 3 (ILC2 and ILC3), are all c-Kit+ cells and may drive chronic inflammatory processes depending on the disease and organ involved. When an inflammatory response is initiated, various mediators, including SCF, activate c-Kit+ immune cells, resulting in the production of cytokines that transform fibroblasts into activated myofibroblasts. Myofibroblasts secrete extracellular matrix proteins, collagen, and fibronectin, leading to tissue fibrosis. Activated myofibroblasts, activated epithelial, endothelial, macrophage, eosinophil, mast cell, monocyte, and other cells also express SCF on their cell surface, further activating c-Kit+ immune cells, resulting in increased cytokine release and perpetuating inflammation. There is a need in the art for more effective and more specific treatments for inflammatory diseases, and the present disclosure addresses these and other needs. [Prior art documents] [Non-patent literature]

[0006]

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Summary of the Invention

Means for Solving the Problems

[0007] In one aspect, the disclosure provides antibodies and fragments thereof that specifically bind to stem cell factor (SCF). In some embodiments, the antibodies and fragments thereof specifically bind to the SCF isoform SCF248. In some embodiments, the antibodies and fragments thereof comprise heavy chain complementarity determining regions (CDRs), where heavy chain CDR1, CDR2, and CDR3 comprise SEQ ID NOs: 1, 2, and 3, respectively. In some embodiments, the antibodies and fragments thereof comprise light chain CDRs, where light chain CDR1, CDR2, and CDR3 comprise SEQ ID NOs: 4, 5, and 6, respectively. In some embodiments, the antibodies and fragments thereof comprise heavy chain CDR1, CDR2, and CDR3 comprise SEQ ID NOs: 1, 37, and 3, respectively. In some embodiments, the antibodies and fragments thereof comprise a heavy chain variable region comprising at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% identity to a sequence selected from the group consisting of SEQ ID NOs: 7, 8, 9, 10, 11, and 12. In some embodiments, the antibodies and fragments thereof comprise a light chain variable region comprising at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% identity to a sequence selected from the group consisting of SEQ ID NOs: 13, 14, 15, 16, and 17. In some embodiments, the antibodies and fragments thereof comprise a heavy chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 7, 8, 9, 10, 11, and 12, and a light chain variable region comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 13, 14, 15, 16, and 17.

[0008] In some embodiments, the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO:7 and a light chain variable region amino acid sequence set forth in SEQ ID NO:16. In some embodiments, the antibody or fragment thereof of claim 1 comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO:8 and a light chain variable region amino acid sequence set forth in SEQ ID NO:16. In some embodiments, the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO:9 and a light chain variable region amino acid sequence set forth in SEQ ID NO:16. In some embodiments, the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO:10 and a light chain variable region amino acid sequence set forth in SEQ ID NO:16. In some embodiments, the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO:11 and a light chain variable region amino acid sequence set forth in SEQ ID NO:16. In some embodiments, the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO:12 and a light chain variable region amino acid sequence set forth in SEQ ID NO:16.

[0009] In some embodiments, the antibody or fragment thereof is humanized. In some embodiments, the antibody is a monoclonal antibody. In some embodiments, the antibody comprises a human IgG1 domain or a human IgG4 domain. In some embodiments, the antibody is an antigen-binding fragment, the fragment being selected from Fab, F(ab')2, Fab', scFv, and a single domain antibody (sdAb).

[0010] In some embodiments, the antibody or fragment thereof blocks the interaction between SCF (e.g., SCF248) and c-Kit. In some embodiments, the antibody specifically binds to SCF248. In some embodiments, the antibody does not bind to SCF220. In some embodiments, the antibody prevents the interaction of SCF248 with c-kit by causing internalization of SCF, making it unavailable on the cell surface.

[0011] In one aspect, the present disclosure provides a pharmaceutical composition comprising the antibody or fragment thereof provided herein. In some embodiments, the pharmaceutical composition comprises a pharmaceutically acceptable carrier, diluent, or excipient.

[0012] In some embodiments, the present disclosure provides an isolated nucleic acid molecule encoding an antibody or fragment thereof provided herein. In some embodiments, the present disclosure provides an expression vector comprising a nucleic acid encoding the antibody or fragment thereof. In further specific embodiments, the present disclosure provides a recombinant host cell comprising the expression vector.

[0013] In one aspect, the present disclosure provides a method for producing an antibody that specifically binds to stem cell factor isoform 248 (SCF248), comprising immunizing a host animal with a peptide comprising SEQ ID NO:30 (ASSLRNDSSSSNRKAKNPPGD) or a fragment thereof, and obtaining the antibody from the immunized host animal. In some embodiments, the host animal is not human. In some embodiments, the fragment of SEQ ID NO:30 comprises at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, or 20 consecutive amino acids of SEQ ID NO:30. In some embodiments, a fragment of SEQ ID NO:30 comprises 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 consecutive amino acids of SEQ ID NO:30. In some embodiments, the N-terminal amino acid of a fragment of SEQ ID NO:30 is alanine at position 1 of the N-terminus of SEQ ID NO:30. In some embodiments, the method includes immunizing a host animal with a peptide consisting of SEQ ID NO:30. In some embodiments, antibodies from the immunized host animal are obtained from immune cells isolated from the host animal. In some embodiments, the method further includes generating a hybridoma using the immune cells. Thus, in some embodiments, the present disclosure provides hybridomas that produce the monoclonal antibodies described herein.

[0014] In one aspect, the disclosure provides an antibody or fragment thereof that specifically binds to SCF248, wherein the antibody or fragment thereof binds to an epitope comprising at least 8, at least 9, at least 10, at least 11, at least 12, or at least 13 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction of SCF248 with c-Kit. In a further embodiment, the epitope comprises SEQ ID NO: 33 or SEQ ID NO: 36. In yet a further embodiment, the epitope consists of SEQ ID NO: 33 or SEQ ID NO: 36.

[0015] In one aspect, the present disclosure provides compositions and methods for inhibiting the interaction between SCF and c-Kit. c-Kit is expressed on immune cells, hematopoietic stem cells, and some structural cells. SCF248, a c-kit ligand, is upregulated on myofibroblasts, activated epithelium, endothelium, macrophages, eosinophils, mast cells, monocytes, and the like. In some embodiments, the compositions and methods specifically inhibit the interaction between SCF248 and c-Kit. For example, in some embodiments, the compositions and methods specifically inhibit the interaction between SCF248 on myofibroblasts and c-Kit on immune cells. As another example, in some embodiments, the compositions and methods provided herein specifically inhibit the interaction between SCF248 on myofibroblasts, activated epithelium, endothelium, macrophages, eosinophils, mast cells, and / or monocytes and c-Kit on immune cells and / or structural cells. In some embodiments, the method comprises contacting SCF248 on myofibroblasts with an antibody or fragment thereof provided herein. In some embodiments, the antibody or fragment thereof provided herein blocks SCF248 from binding to c-Kit. In some embodiments, the blockage is mediated by steric hindrance. In some embodiments, the antibody or fragment thereof provided herein internalizes SCF248.

[0016] In some embodiments, the present disclosure provides a method for inhibiting inflammation in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof provided herein. In some embodiments, the present disclosure provides a method for inhibiting an inflammatory disease in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof provided herein. In further embodiments, the inflammatory disease is a chronic inflammatory disease. In some embodiments, the present disclosure provides a method for treating inflammation and / or a chronic inflammatory disease in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof provided herein.

[0017] In some embodiments, the present disclosure provides a method for inhibiting fibrosis in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof provided herein. In some embodiments, the present disclosure provides a method for treating a fibrotic disease in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof provided herein. In various embodiments, the method further comprises administering one or more additional therapies and / or therapeutic agents.

[0018] In some embodiments, the inflammatory or fibrotic disease is selected from the group consisting of urticaria, atopic dermatitis, bullous pemphigoid, scleroderma, systemic sclerosis, non-alcoholic steatohepatitis (NASH), primary sclerosing cholangitis, cirrhosis, pulmonary fibrosis (e.g., idiopathic pulmonary fibrosis (IPF), scleroderma pulmonary fibrosis), chronic obstructive pulmonary disease (COPD), acute respiratory distress syndrome (ARDS), cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, asthma, bleomycin lung, endomyocardial fibrosis, fibromyalgia, eosinophilic esophagitis, radiation fibrosis, rheumatoid arthritis, and inflammatory bowel disease. [Brief explanation of the drawings]

[0019] [Figure 1]1 shows an outline of the tissue injury / inflammatory disease process. [Figure 2] 1 shows an exemplary mechanism for 5H10, an anti-SCF248 antibody of the present disclosure. The 5H10 antibody is referred to as "OpSCF" in the figures. [Figure 3] Shown are the SCF isoforms SCF220 and SCF248, as well as the monomeric cleaved extracellular domain, SCF165, which is released upon cleavage of SCF248 at a cleavage site within the Exon 6 region. [Figure 4A] 1 is a set of histograms showing binding of the mouse 5H10 antibody to control cells that do not express SCF (left panel), cells that express SCF220 but not SCF248 (middle panel), and cells that express SCF248 but not SCF220 (right panel). [Figure 4B] Binding of the murine 5H10 antibody to a 165 amino acid truncated SCF extracellular domain (ECD) and the complete 194 amino acid SCF ECD is shown. [Figure 5] Shown is the mean fluorescence intensity (MFI) measured by flow cytometry after exposure of cultured human IPF myofibroblasts to pHrodo red-labeled 2G8, 5H10, or control IgG antibodies. [Figure 6] Figure 1 shows activation of the P13K / ACT and MEK / ERK pathways of c-kit signaling after contacting eosinophils with SCF248-expressing cells in the presence of 5H10 antibody or IgG control. The 5H10 antibody significantly reduced activation of both pathways. [Figure 7] Binding of murine 2G8 and 5H10 antibodies to S1 / S14 hSCF248 (ATCC® CRL2454™) early passage cells (A) and late passage cells (B) is shown by flow cytometry. [Figure 8A] Binding of murine 2G8 and murine 5H10 to S1 / S14 hSCF220 (ATCC® CRL2453™) early passage and hygromycin B-treated cells is shown. [Figure 8B] Binding of murine 2G8 and murine 5H10 to S1 / S14 hSCF248 early passage cells and hygromycin B treated cells is shown. [Figure 9A] Binding of 2G8 humanized variants at different antibody concentrations to S1 / S14 hSCF220 cells is shown by flow cytometry. [Figure 9B] Binding of different concentrations of 2G8 humanized variants to S1 / S14 hSCF248 cells is shown by flow cytometry. [Figure 10A] Binding of 5H10 humanized variants at different antibody concentrations to S1 / S14 hSCF248 cells is shown by flow cytometry. [Figure 10B] Binding of 5H10 humanized variants at different antibody concentrations to S1 / S14 hSCF220 cells is shown by flow cytometry. [Figure 11A] Binding of 2G8 humanized variants at different antibody concentrations to S1 / S14 hSCF248 cells is shown by flow cytometry. [Figure 11B] Binding of 5H10 humanized variants at different antibody concentrations to S1 / S14 hSCF248 cells is shown by flow cytometry. [Figure 11C] Binding of 5H10 humanized variants at different antibody concentrations to S1 / S14 hSCF248 cells is shown by flow cytometry. In Figure 11C, the VH shown is paired with VK3. [Figure 11D] Binding of 5H10 humanized variants at different antibody concentrations to S1 / S14 hSCF248 cells is shown by flow cytometry. In Figure 11D, the 5H10 antibody shown is VH1 / VK3. [Figure 12A]The figure shows the change in CCL11 mRNA levels after preincubation of human IPF myofibroblasts (Mfb) with a positive control (irrelevant antibody) or the antibodies indicated below each bar. The mouse parent antibody is designated "5H10" in the figure. The humanized 5H10 antibodies VH1 / VK3, VH2 / VK3, VH3 / VK3, VH4 / VK3, and VH5 / VK3 were also tested, as indicated. The antibody concentrations tested were 1 μg / mL or 10 μg / mL. [Figure 12B] Figure 1 shows the change in collagen 1A1 mRNA levels after preincubation of human IPF myofibroblasts (Mfb) with a positive control (irrelevant antibody) or the antibodies indicated below each bar in the figure. The mouse parent antibody is designated "5H10" in the figure. Humanized 5H10 antibodies VH1 / VK3, VH2 / VK3, VH3 / VK3, VH4 / VK3, and VH5 / VK3 were also tested as indicated. Antibody concentrations tested were 1 μg / mL or 10 μg / mL. [Figure 12C] The figure shows the change in fibronectin mRNA levels after preincubation of human IPF myofibroblasts (Mfb) with a positive control (irrelevant antibody) or the antibodies indicated below each bar. The mouse parent antibody is designated "5H10" in the figure. The humanized 5H10 antibodies VH1 / VK3, VH2 / VK3, VH3 / VK3, VH4 / VK3, and VH5 / VK3 were also tested as indicated. The antibody concentrations tested were 1 μg / mL or 10 μg / mL. [Figure 12D] Figure 1 shows the change in collagen 3 mRNA levels after preincubation of human IPF myofibroblasts (Mfb) with a positive control (irrelevant antibody) or the antibodies indicated below each bar in the figure. The mouse parent antibody is designated "5H10" in the figure. Humanized 5H10 antibodies VH1 / VK3, VH2 / VK3, VH3 / VK3, VH4 / VK3, and VH5 / VK3 were also tested as indicated. Antibody concentrations tested were 1 μg / mL or 10 μg / mL. [Figure 13]Figure 1 shows the internalization of pHrodo red-labeled murine 5H10 antibody, VH0 / VK0 chimeric antibody, humanized variants VH1 / VK3, and VH2 / VK3. Arrows indicate exemplary cells showing internalized antibody. [Figure 14] 1 shows the effect of 5H10 humanized variants on PBMC viability. [Figure 15A] 1 shows CD4+ T cell responses induced by the indicated 5H10 humanized variants in an EpiScreen™ time course T cell proliferation assay. [Figure 15B] 1 shows CD4+ T cell responses induced by the indicated 5H10 humanized variants in an EpiScreen™ time course T cell proliferation assay. [Figure 15C] 1 shows CD4+ T cell responses induced by the indicated 5H10 humanized variants in an EpiScreen™ time course T cell proliferation assay. [Figure 15D] 1 shows CD4+ T cell responses induced by the indicated 5H10 humanized variants in an EpiScreen™ time course T cell proliferation assay. [Figure 15E] 1 shows CD4+ T cell responses induced by the indicated 5H10 humanized variants in an EpiScreen™ time course T cell proliferation assay. [Figure 15F] 1 shows CD4+ T cell responses induced by the indicated 5H10 humanized variants in an EpiScreen™ time course T cell proliferation assay. [Figure 15G] Figure 1 shows CD4+ T cell responses induced by the indicated 5H10 humanized variants in an EpiScreen™ time course T cell proliferation assay. Exenatide is a positive control. [Figure 15H] Figure 1 shows CD4+ T cell responses induced by the indicated 5H10 humanized variants in an EpiScreen™ time course T cell proliferation assay. KLH is a positive control. [Figure 16] 1 shows the analysis of variance (ANOVA) of the EpiScreen™ time course T cell proliferation assay. [Figure 17] Lung histology of a bleomycin control animal (left panel) and an animal treated with bleomycin and 5H10 (20 mg / kg) is shown. [Figure 18] 1 shows the reduction in lung hydroxyproline in animals treated with bleomycin and control IgG, and in animals treated with bleomycin and 5H10 (20 mg / kg; referred to in the figure as anti-SCF248). [Figure 19] The change in body weight over time in naive mice, mice treated with bleomycin and a control Ig antibody (Bleo+cIg) to induce pulmonary fibrosis, and mice treated with bleomycin and murine 5H10 antibody (referred to in the figures as anti-SCF248) is shown as a percentage of the body weight measured on day 0. Antibodies were administered at the indicated time points. [Figure 20] Figure 1 shows a decrease in mRNA for inflammatory cytokines and myofibroblast activation markers (TGFβ, CCL2, Col1a1, fibronectin (fn), smooth muscle actin (acta2), and stem cell factor (kitlg)) in animals treated with bleomycin and control IgG, and in animals treated with bleomycin and 5H10 (20 mg / kg; referred to as anti-SCF248 in the figure). [Figure 21] 1 shows the reduction of pulmonary mast cells, eosinophils, and ILC2 lymphocytes in animals treated with bleomycin and control IgG, and in animals treated with bleomycin and 5H10 (20 mg / kg; referred to in the figure as anti-SCF248). [Figure 22] Pulmonary function tests, as measured by forced expiratory volume (left panel), forced expiratory rate (middle panel), and change in lung pressure (right panel), show significant improvement in animals treated with bleomycin and 5H10 (20 mg / kg; referred to in the figure as anti-SCF248) compared with animals treated with bleomycin and control IgG. [Figure 23]FIG. 1 shows a schematic diagram of the study design in an in vivo chronic allergic asthma model used to test humanized antibodies. [Figure 24] Figure 1 shows the results of treatment with the humanized 5H10 antibody in an in vivo model of chronic allergic asthma. (A) Airway resistance was measured and significantly reduced in animals treated with VH1 / VK3 compared to PBS controls. IL-13 mRNA (B), collagen 1 mRNA (C), and collagen 3 mRNA (D) were also reduced in lung tissue in animals treated with VH1 / VK3 compared to chronic asthma (PBS) controls. (E) SCF248 mRNA expression was also reduced in animals treated with the VH1 / VK3 5H10 antibody compared to PBS controls. [Figure 25] Figure 1 shows that antibody VH1 / VK3 reduced mRNA levels of mucus proteins Gob5 (A), IL-13 (B), and IL-5 (C) in vivo at concentrations of 1 mg / kg and 5 mg / kg. DETAILED DESCRIPTION OF THE INVENTION

[0020] Stem cell factor (SCF) is a key mediator of acute and chronic inflammation, fibrotic diseases, and tissue remodeling diseases. The interaction of SCF with c-Kit on immune cells initiates and perpetuates inflammation and fibrosis. The present disclosure provides compositions and methods for inhibiting the interaction between SCF and c-Kit. In one embodiment, the present disclosure provides compositions and methods for preventing SCF248, the inflammatory form of SCF, from interacting with c-Kit to reduce and / or prevent immune cell activation. Accordingly, the present disclosure provides methods for treating chronic inflammation and fibrotic and tissue remodeling diseases. In one embodiment, the present disclosure provides compositions and methods for reducing the accumulation (e.g., proliferation and / or retention) of immune cells in an organ or tissue. For example, the present disclosure provides compositions and methods for preventing SCF248 from interacting with c-Kit to reduce and / or prevent the accumulation of immune cells in an organ or tissue. In some embodiments, the present disclosure provides compositions and methods for reducing and / or preventing the activation and / or accumulation of mast cells, eosinophils, type 2 innate lymphoid (ILC2) cells, and type 3 innate lymphoid (ILC3) cells in an organ or tissue.

[0021] In particular, the present disclosure provides antibodies and fragments thereof that specifically bind to SCF and block or inhibit its interaction with c-Kit. In some embodiments, the antibodies and fragments provided herein bind to SCF and inhibit the activity of c-Kit and c-Kit+ cells. The present disclosure also provides methods for producing antibodies and fragments thereof that specifically bind to SCF, as well as diagnostic and therapeutic methods of their use. In one aspect, the antibodies and fragments provided herein specifically bind to SCF248, an SCF isoform that promotes inflammation. Thus, the present disclosure provides certain effective compositions and methods for inhibiting inflammation and fibrosis and treating chronic inflammatory and fibrotic diseases.

[0022] definition As used herein, the term "antibody" refers to a binding protein having at least one antigen-binding domain. The antibodies and fragments thereof of the present invention may be whole antibodies or any fragments thereof. Thus, the antibodies and fragments of the present invention include monoclonal antibodies or fragments thereof, antibody variants or fragments thereof, and immunoconjugates. Antigen-binding fragments include Fab fragments, Fab' fragments, F(ab')2 fragments, bispecific Fab dimers (Fab2), trispecific Fab trimers (Fab3), Fv, single-chain Fv proteins ("scFv"), bis-scFv, (scFv)2, minibodies, diabodies, triabodies, tetrabodies, disulfide-stabilized Fv proteins ("dsFv"), single-domain antibodies (sdAbs, nanobodies), heavy-chain-only antibodies (e.g., camelid VHHs, camelid nanobodies, shark Ig NARs), and the portion of a full-length antibody responsible for antigen binding. An isolated antibody or antigen-binding fragment thereof is an antibody that has been identified and separated and / or recovered from a component of its natural environment.

[0023] In some embodiments, the antibodies and antigen-binding fragments thereof are isolated antibodies and fragments thereof. Accordingly, the present invention provides isolated antibodies and antigen-binding fragments thereof, as well as nucleic acids encoding such antibodies and fragments, and compositions comprising such isolated antibodies, fragments, and nucleic acids. The term "isolated" refers to a compound of interest (e.g., an antibody or nucleic acid) that has been separated from its natural environment. The present invention also provides pharmaceutical compositions comprising an isolated antibody or fragment thereof, or a nucleic acid encoding such an antibody or fragment, and further comprising one or more pharmaceutically acceptable carriers. Pharmaceutically acceptable carriers include, for example, excipients, diluents, encapsulating materials, fillers, buffers, or other agents.

[0024] As used herein, the term "derived from," when used in reference to a molecule or polypeptide to which a reference antibody or other binding protein is directed, means a molecule or polypeptide that is specific for and capable of binding to the same epitope as the reference antibody or other binding protein.

[0025] As used herein, the phrase "specific for" may mean that the antibody does not bind to the target solely through non-specific interactions, and this property can be determined by comparison with an isotype control or the like. Specific binding may include, but is not necessarily required to, exclusive binding to a single target. In various embodiments, the antibodies provided herein specifically bind to SCF248 and do not bind to SCF220.

[0026] The term "host cell" refers to a cell that has been transformed or is transformable with a nucleic acid sequence and thereby expresses a gene of interest. The term includes the progeny of the parent cell, whether or not the progeny is identical in morphology or genetic make-up to the original parent cell, so long as the gene of interest is present.

[0027] A "variant" of a polypeptide (e.g., an antigen-binding protein or antibody) comprises an amino acid sequence in which one or more amino acid residues have been inserted, deleted, and / or substituted into the amino acid sequence compared to another polypeptide sequence. Variants include antibodies and fragments thereof having a recited percent identity to the antibodies or fragments provided herein or to antibodies or fragments having a recited DNA or amino acid sequence.

[0028] The term "identity" refers to the relationship between the sequences of two or more polypeptide molecules or two or more nucleic acid molecules, as determined by aligning and comparing the sequences. "Percent identity," "percent homology," "sequence identity," "sequence homology," and the like refer to the percent of identical residues between amino acids or nucleotides in the compared molecules, and are calculated based on the size of the smallest molecule being compared. For such calculations, gaps in the alignment, if any, are preferably addressed by a specific mathematical model or computer program (i.e., "algorithm"). Methods that can be used to calculate the identity of aligned nucleic acids or polypeptides include those described in Computational Molecular Biology, (Lesk, AM, ed.), 1988, New York: Oxford University Press; Biocomputing Informatics and Genome Projects, (Smith, DW, ed.), 1993, New York: Academic Press; Computer Analysis of Sequence Data, Part I, (Griffin, AM, and Griffin, HG, eds.), 1994, New Jersey: Humana Press; von Heinje, G., 1987, Sequence Analysis in Molecular Biology, New York: Academic Press; Sequence Analysis Primer, (Gribskov, M. and Devereux, J., eds.), 1991, New York: M. Stockton Press; and Carillo et al.,1988,SIAM J. Applied Math. 48: 1073. In calculating percent identity, the sequences being compared are typically aligned to give the highest match between the sequences.

[0029] The term "light chain" includes full-length light chains and fragments thereof containing sufficient variable region sequence to confer binding specificity. A full-length light chain contains a variable region domain and a constant region domain. The variable region domain of a light chain is at the amino-terminus of the polypeptide. Light chains include kappa chains and lambda chains.

[0030] The term "heavy chain" includes full-length heavy chains and fragments thereof containing sufficient variable region sequence to confer binding specificity. A full-length heavy chain contains a variable region domain and three constant region domains (C H 1. C H 2, and C H 3) The variable heavy domain is at the amino terminus of the polypeptide and comprises C H The CH3 domain is at the carboxyl terminus, with the CH3 domain being closest to the carboxy terminus of the polypeptide. The heavy chain can be of any isotype, including IgG (including IgG1, IgG2, IgG3, and IgG4 subtypes), IgA (including IgA1 and IgA2 subtypes), IgM, and IgE. The term "isotype" refers to the antibody class encoded by the heavy chain constant region gene. In some embodiments, the antibodies provided herein have an IgG4 heavy chain or an IgG4 heavy chain containing certain amino acid mutations. For example, in some embodiments, the IgG4 contains a mutation at position 228 to inhibit Fab arm exchange (EU numbering scheme (Kabat et al., Sequence of proteins of immunologic interest, 5th ed. Bethesda, MD, NIH 1991)). For example, in some embodiments, the IgG4 heavy chain is an IgG4 S228P heavy chain. In some embodiments, the heavy chain contains one or more amino acid mutations that reduce binding to Fc receptors, thereby reducing or eliminating antibody effector function. For example, the heavy chain may contain a mutation at one or more of positions 233, 234, 235, 236, 237, 265, 309, 331, and 409 (EU numbering).

[0031] The term "variable region" or "variable domain" refers to a portion of an antibody's light and / or heavy chain, typically comprising approximately the amino-terminal 120-130 amino acids in the heavy chain and approximately the amino-terminal 100-110 amino acids in the light chain. In certain embodiments, the variable regions of different antibodies vary extensively in amino acid sequence, even among antibodies of the same species. The variable region of an antibody typically determines the specificity of that particular antibody for its target. As used herein, the term "target" refers to a molecule or portion of a molecule capable of binding by an antigen-binding protein. In certain embodiments, a target may have one or more epitopes. In certain embodiments, a target is an antigen. The use of "antigen" in the phrase "antigen-binding protein" simply means that the protein sequence comprising the antigen can be bound by an antibody. In this context, it is not required that the protein be foreign or capable of inducing an immune response.

[0032] The term "epitope" includes any determinant capable of binding by an antigen-binding protein (e.g., an antibody or T-cell receptor). An epitope is a region of an antigen bound by an antigen-binding protein that targets the antigen; if the antigen is a protein, the epitope includes specific amino acids that directly contact the antigen-binding protein. Epitopes are most often found on proteins, but in some cases may be found on other types of molecules (e.g., nucleic acids). Epitopes may include chemically active surface groupings of molecules such as amino acids, sugar side chains, phosphoryl or sulfonyl groups, and may have specific three-dimensional structural characteristics and / or specific charge characteristics. Generally, antibodies specific for a particular target antigen will preferentially recognize epitopes on the target antigen in a complex mixture of proteins and / or macromolecules. Antibody epitopes can be linear or conformational. In various embodiments, the epitopes provided herein are linear epitopes.

[0033] The use of the singular includes the plural unless otherwise specified. The word "a" or "an" means "at least one" unless otherwise specified. The use of "or" means "and / or" unless otherwise specified. The meaning of the phrase "at least one" is equivalent to the meaning of the phrase "one or more." Furthermore, the use of the term "including" and other forms (e.g., "includes" and "included") is not limiting. Furthermore, terms such as "element" or "component" encompass both elements or components containing one unit and elements or components containing two or more units unless otherwise specified. As used herein, the term "about" refers to an amount greater or less than the stated parameter value, for example, plus or minus 5% or 10% of the object that "about" modifies, or as one of ordinary skill in the art would recognize from the context (e.g., approximately 50% of the interval between values). The term "about" also includes the value being referred to.

[0034] stem cell factor At least two forms of SCF exist in humans, each with distinct structures and activities. SCF220 functions in several homeostatic functions, including hematopoiesis and spermatogenesis, and is found in the bone marrow, testis, and other tissues and organs. SCF220 is slowly cleavable and is sometimes referred to as "membrane SCF." In contrast, SCF248 is rapidly cleavable and contains a cleavage site in exon 6, located between the N-terminal c-kit binding domain and the transmembrane domain. SCF248 is also sometimes referred to as "soluble SCF." Exon 6 is omitted from SCF220 by alternative splicing, and therefore SCF220 lacks this cleavage site. The monomeric extracellular domain (SCF165) is a cleavage product and serves as a plasma biomarker for chronic inflammatory diseases. Although plasma may also contain detectable levels of SCF extracellular domains derived from SCF220, it is expected that the majority of detectable extracellular domains are SCF165. SCF248 is an isoform found in myofibroblasts, activated epithelial cells, and other cells, and activates immune cells during inflammation, contributing to the perpetuation of fibrosis. More specifically, SCF248 binds to c-Kit on immune cells, initiating cytokine production and activating fibroblasts to transform into myofibroblasts, which secrete extracellular matrix proteins, collagen, and fibronectin. In addition to activated myofibroblasts, activated epithelial, endothelial, macrophage, eosinophil, mast cell, monocyte, and other cells also express SCF on their cell surface, activating more c-Kit+ immune cells, resulting in the release of additional cytokines and immune activation and fibrotic responses.

[0035] The antibodies and antigen-binding fragments thereof disclosed herein are specific for SCF. In some embodiments, the antibodies and fragments thereof are specific for human SCF. In some embodiments, the antibodies and fragments thereof are specific for SCF248. In some embodiments, the antibodies bind to SCF248 and not to other isoforms of SCF. In some embodiments, the antibodies bind to SCF248 and not to SCF220. In some embodiments, the disclosure provides methods for producing antibodies or fragments thereof specific for SCF248. The disclosure provides exemplary antibodies and fragments specific for SCF248, as well as methods for producing and using such antibodies and fragments. In some embodiments, the antibodies and fragments thereof provided herein bind to SCF248 and block the interaction between SCF248 and c-Kit, thereby interrupting the positive feedback loop between SCF248 expressed on various cell types and cKit+ immune cells.

[0036] Antibodies and fragments The present disclosure provides antibodies (including monoclonal antibodies) and fragments thereof. The antibody fragments specific for SCF (e.g., SCF248) provided herein are sometimes referred to herein as antigen-binding fragments, meaning that they contain a portion of the parent antibody that is capable of binding to the target antigen (SCF (e.g., SCF248)). The terms "antibody fragment," "antigen-binding fragment," and the like are used interchangeably herein. Examples of antigen fragments include Fab fragments, Fab' fragments, F(ab)' fragments, Fv fragments, isolated CDR regions, bispecific Fab dimers (Fab2), trispecific Fab trimers (Fab3), single chain Fv proteins ("scFv"), bis-scFv, (scFv)2, minibodies, diabodies, triabodies, tetrabodies, disulfide stabilized Fv proteins ("dsFv"), single domain antibodies (sdAb, nanobodies), heavy chain only antibodies (e.g. camelid VHH, camelid nanobody, shark Ig NAR), and the portion of a full length antibody responsible for antigen binding.

[0037] A "Fab fragment" is a fragment of one light chain and one heavy chain C H The heavy chain of a Fab molecule cannot form disulfide bonds with another heavy chain molecule. An "Fab' fragment" contains one light chain and a portion of one heavy chain, which contains the VH domain and C H 1 domain, plus C H 1 Domain and C H The "F(ab')2 fragment" contains two light chains and two heavy chains, and these heavy chains are connected to the C(ab')2 domains. H 1 Domain and C HThe F(ab')2 fragment contains a portion of the constant region between the two heavy chains, resulting in the formation of an interchain disulfide bond between the two heavy chains. Thus, the F(ab')2 fragment is composed of two Fab' fragments, which are held together by disulfide bonds between the two heavy chains. The Fv fragment contains the variable regions of both the heavy and light chains and lacks the constant region. An "scFv" is an Fv molecule in which the variable regions of the heavy and light chains are connected by a flexible linker to form a single polypeptide chain that forms an antigen-binding region.

[0038] In some embodiments, the antibodies and fragments thereof provided herein are defined by their complementarity-determining regions (CDRs). CDRs are portions of the variable chains within an antibody, and each of the light chain variable region and heavy chain variable region contains three CDRs: CDR1, CDR2, and CDR3. The CDRs of an antibody determine its antigen specificity. In certain embodiments, definitive definition of the CDRs and identification of the residues comprising the antibody binding site are accomplished by elucidating the structure of the antibody and / or the structure of an antibody-ligand complex. In certain embodiments, this can be accomplished by any of a variety of techniques known to those skilled in the art (e.g., X-ray crystallography). In certain embodiments, various analytical methods can be used to identify or approximate CDR regions. Examples of such methods include, but are not limited to, the Kabat definition, the Chothia definition, the AbM definition, and the contact definition.

[0039] The Kabat definition is a standard for numbering residues in antibodies and is typically used to identify CDR regions. See, e.g., Johnson & Wu, Nucleic Acids Res., 28:214-8 (2000). The Chothia definition is similar to the Kabat definition, but takes into account the location of certain structural loop regions. See, e.g., Chothia et al., J. Mol. Biol., 196:901-17 (1986); Chothia et al., Nature, 342:877-83 (1989). The AbM definition uses an integrated suite of computer programs for modeling antibody structure produced by the Oxford Molecular Group. See, e.g., Martin et al., Proc Natl Acad Sci (USA), 86:9268-9272 (1989); "AbM™, A Computer Program for Modeling Variable Regions of Antibodies," Oxford, UK; Oxford Molecular, Ltd. The AbM definition models the tertiary structure of antibodies from primary sequence using a combination of knowledge databases and first-principles methods, such as those described in Samudrala et al., "Ab Initio Protein Structure Prediction Using a Combined Hierarchical Approach" (PROTEINS, Structure, Function and Genetics Suppl., 3:194-198 (1999)). Contact definitions are based on analysis of available complex crystal structures. See, e.g., MacCallum et al., J. Mol. Biol., 5:732-45 (1996).

[0040] Antibodies and fragments thereof can also include recombinant polypeptides, fusion proteins, and bispecific antibodies. The anti-SCF antibodies and fragments thereof disclosed herein can be of the IgG1, IgG2, IgG3, or IgG4 isotype. In one embodiment, the anti-SCF antibodies and fragments thereof disclosed herein are of the IgG1 or IgG4 isotype. The anti-SCF antibodies and fragments thereof of the present invention can be derived from any species, including, but not limited to, mouse, rat, rabbit, primate, llama, camel, goat, shark, chicken, and human. The SCF antibodies and fragments thereof can be chimeric, humanized, or fully human. In one embodiment, the anti-SCF antibody is a murine antibody. In another embodiment, the anti-SCF antibody is a chimeric antibody. In a further embodiment, the chimeric antibody is a mouse-human chimeric antibody. In another embodiment, the antibody is derived from a mouse and humanized.

[0041] A "chimeric antibody" is an antibody having at least a portion of a heavy chain variable region and at least a portion of a light chain variable region from one species and at least a portion of a constant region from another species. For example, in one embodiment, a chimeric antibody can contain a murine variable region and a human constant region.

[0042] A "humanized antibody" is an antibody that contains complementarity-determining regions (CDRs) derived from a non-human antibody and framework and constant regions derived from a human antibody. For example, an anti-SCF antibody provided herein may contain CDRs derived from one or more mouse antibodies and human framework and constant regions. Thus, in one embodiment, a humanized antibody provided herein binds to the same epitope on SCF as the mouse antibody from which the antibody's CDRs are derived.

[0043] In some embodiments, the antibodies and fragments thereof provided herein comprise a heavy chain and a light chain, each of which comprises three CDRs. Exemplary amino acid sequences of heavy chain CDR1, CDR2, and CDR3 (HCDR1, HCDR2, and HCDR3, respectively) and light chain CDR1, CDR2, and CDR3 (LCDR1, LCDR2, and LCDR3, respectively) are set forth in Table 1 below. Exemplary amino acid sequences of heavy and light chain variable regions are also set forth in Table 1. In some embodiments, the present disclosure provides antibodies designated herein as "5H10" and "2G8." The heavy chain variable regions of humanized 5H10 or 2G8 are designated herein as VH1, VH2, VH3, VH4, and VH5. 5H10 VH0 is the variable heavy chain of the murine parent antibody generated by the methods described herein. VH1, VH2, VH3, VH4, and VH5 are each humanized heavy chain variable regions derived from 5H10 VH0 or 2G8 VH0. The 5H10 antibody contains a kappa light chain. The murine parent antibody variable light chain is referred to herein as 5H10 VK0. VK1, VK2, VK3, and VK4 are each humanized light chain variable regions derived from VK0. The 2G8 antibody contains a lambda light chain. The murine parent antibody variable light chain is referred to herein as 2G8 VL0. VL1, VL2, VL3, and VL4 are each humanized light chain variable regions derived from VL0. [Table 1-1] [Table 1-2]

[0044] One of skill in the art will understand that the variable heavy and variable light chains may be independently selected from the antibodies provided herein or may be mixed and matched. Thus, in some embodiments, the antibodies and fragments thereof provided herein include any of the following: VH0 / VK0, VH0 / VK1, VH0 / VK2, VH0 / VK3, VH0 / VK4, VH1 / VK0, VH1 / VK1, VH1 / VK2, VH1 / VK3, VH1 / VK4, VH2 / VK0, VH2 / VK1, VH2 / VK2, VH2 / VK3, The combinations include heavy and light chains selected from the group consisting of VH2 / VK4, VH3 / VK0, VH3 / VK1, VH3 / VK2, VH3 / VK3, VH3 / VK4, VH4 / VK0, VH4 / VK1, VH4 / VK2, VH4 / VK3, VH4 / VK4, VH5 / VK0, VH5 / VK1, VH5 / VK2, VH5 / VK3, and VH5 / VK4.

[0045] In some embodiments, the present disclosure provides antibodies or fragments thereof comprising an amino acid sequence at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 7 to 12. In some embodiments, the present disclosure provides antibodies or fragments thereof comprising a heavy chain variable region set forth in a sequence selected from the group consisting of SEQ ID NOs: 7 to 12. In some embodiments, the present disclosure provides antibodies or fragments thereof comprising an amino acid sequence at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 7 to 11, wherein the antibodies or fragments comprise heavy chain CDR1, CDR2, and CDR3 identical to SEQ ID NOs: 1, 2, and 3, respectively. In some embodiments, the present disclosure provides an antibody or fragment comprising an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% homology to the amino acid sequence of SEQ ID NO: 12, and comprising heavy chain CDR1, CDR2, and CDR3 identical to SEQ ID NOs: 1, 37, and 3, respectively.

[0046] In some embodiments, the present disclosure provides antibodies or fragments thereof comprising an amino acid sequence at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 13 to 17. In some embodiments, the present disclosure provides antibodies or fragments thereof comprising a light chain variable region set forth in a sequence selected from the group consisting of SEQ ID NOs: 13 to 17. In some embodiments, the present disclosure provides antibodies or fragments thereof comprising an amino acid sequence at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 13 to 17, wherein the antibodies or fragments comprise light chain CDR1, CDR2, and CDR3 identical to SEQ ID NOs: 4, 5, and 6, respectively.

[0047] In some embodiments, the present disclosure provides antibodies or fragments comprising amino acid sequences having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% homology to the following: SEQ ID NO:7 and SEQ ID NO:13; SEQ ID NO:7 and SEQ ID NO:14; SEQ ID NO:7 and SEQ ID NO:15; SEQ ID NO:7 and SEQ ID NO:16; SEQ ID NO:7 and SEQ ID NO:17; ...8 and SEQ ID NO:13; SEQ ID NO:8 and SEQ ID NO:14; SEQ ID NO:8 and SEQ ID NO:15; SEQ ID NO:8 and SEQ ID NO:16; SEQ ID NO:8 and SEQ ID NO:17; SEQ ID NO:9 and SEQ ID NO:13; SEQ ID NO:9 and SEQ ID NO:14; SEQ ID NO:9 and SEQ ID NO:15; SEQ ID NO:9 and SEQ ID NO:16; SEQ ID NO:9 and SEQ ID NO:17; SEQ ID NO:10 and SEQ ID NO:13; SEQ ID NO:10 and SEQ ID NO:14; SEQ ID NO:10 and SEQ ID NO:15; SEQ ID NO:10 and SEQ ID NO:16; SEQ ID NO:10 and SEQ ID NO:17; SEQ ID NO:11 and SEQ ID NO:13; SEQ ID NO:11 and SEQ ID NO:14; SEQ ID NO:11 and SEQ ID NO:15; SEQ ID NO:11 and SEQ ID NO:16; SEQ ID NO:11 and SEQ ID NO:17; SEQ ID NO:12 and SEQ ID NO:13; SEQ ID NO:12 and SEQ ID NO:14; SEQ ID NO:12 and SEQ ID NO:15; SEQ ID NO:12 and SEQ ID NO:16; or SEQ ID NO:12 and SEQ ID NO:17.

[0048] In certain embodiments, antibodies and fragments thereof comprise a heavy and light chain combination selected from the group consisting of VH1 / VK1, VH1 / VK2, VH1 / VK3, VH2 / VK1, VH2 / VK2, VH2 / VK3, VH3 / VK1, VH3 / VK2, VH3 / VK3, VH4 / VK1, VH4 / VK2, VH4 / VK3, VH5 / VK1, VH5 / VK2, and VH5 / VK3. In some embodiments, the antibody comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:8 or SEQ ID NO:9, and an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO:16. In some embodiments, the antibody or fragment thereof comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO: 8 or SEQ ID NO: 9, and an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to SEQ ID NO: 16, and the antibody or fragment comprises heavy chain CDR1, CDR2, and CDR3 identical to SEQ ID NOs: 1, 2, and 3, respectively, and light chain CDR1, CDR2, and CDR3 identical to SEQ ID NOs: 4, 5, and 6, respectively. In some embodiments, the antibody or fragment thereof comprises an amino acid sequence having at least 95% or at least 99% sequence identity to SEQ ID NO:8 or SEQ ID NO:9 and an amino acid sequence having at least 95% or at least 99% sequence identity to SEQ ID NO:16, and the antibody or fragment comprises heavy chain CDR1, CDR2, and CDR3 identical to SEQ ID NOs:1, 2, and 3, respectively, and light chain CDR1, CDR2, and CDR3 identical to SEQ ID NOs:4, 5, and 6, respectively. The antibody or fragment thereof may specifically bind to SCF248 but not to SCF220. In some embodiments, the antibody comprises a heavy chain variable region set forth in SEQ ID NO:8 and a light chain variable region set forth in SEQ ID NO:16.In some embodiments, the antibody comprises a heavy chain variable region set forth in SEQ ID NO:9 and a light chain variable region set forth in SEQ ID NO:16.

[0049] In some embodiments, the antibodies and fragments provided herein comprise a heavy chain variable region amino acid sequence set forth in SEQ ID NO: 7, 8, 9, 10, 11, or 12, or a variant thereof, and / or a light chain variable region amino acid sequence set forth in SEQ ID NO: 13, 14, 15, 16, or 17, or a variant thereof. Variants may contain 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions or deletions, or a combination thereof. In some embodiments, the amino acid substitutions are conservative substitutions. Anti-SCF antibodies disclosed herein having one or more amino acid substitutions, insertions, deletions, or a combination thereof in the CDRs or variable light or heavy chain regions retain the biological activity of a corresponding anti-SCF antibody without the amino acid substitutions, insertions, or deletions relative to the sequences provided herein. Thus, the variant anti-SCF antibodies provided herein retain specific binding to SCF248. As used herein, terms such as percent homology, sequence identity, sequence homology, etc. are used interchangeably and refer to the number of identical amino acid positions shared by two reference sequences divided by the total number of amino acid positions multiplied by 100.

[0050] In some embodiments, the present disclosure provides antibodies that bind to the same epitope as any one of the exemplary antibodies disclosed herein. Accordingly, in some embodiments, the present disclosure provides antibodies whose binding to SCF competes with the exemplary antibodies provided herein. For example, in some embodiments, the present disclosure provides antibodies that specifically bind to a region of the amino acid sequence set forth herein as SEQ ID NO:29. In some embodiments, the antibodies provided herein specifically bind to an epitope comprising the amino acid sequence of SEQ ID NO:33 (ASSLRNDSSSSNRK) or SEQ ID NO:36 (ASSLRNDSSSSNR). In some embodiments, the present disclosure provides antibodies that specifically bind to an epitope consisting of the amino acid sequence set forth in SEQ ID NO:33 or SEQ ID NO:36. In some embodiments, the present disclosure provides antibodies that specifically bind to an epitope comprising at least 5, at least 6, at least 7, at least 8, at least 9, or at least 10 consecutive amino acids of SEQ ID NO:33.

[0051] In some embodiments, the antibodies and fragments thereof provided herein comprise HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and / or LCDR3 of the heavy and light chain variable regions provided herein, or variants thereof. Thus, in some embodiments, the antibodies and fragments thereof provided herein include antibodies whose HCDR is the HCDR of SEQ ID NO: 7, 8, 9, 10, 11, or 12, and / or whose LCDR is the LCDR of SEQ ID NO: 13, 14, 15, 16, or 17. For example, in some embodiments, the antibodies and fragments thereof comprise amino acids 31-35, 50-65, and 95-102 of any one of the heavy chain variable regions provided herein, as defined by the Kabat numbering scheme. In some embodiments, the antibodies and fragments thereof comprise amino acids 24-34, 50-56, and 89-97 of any one of the light chain variable regions provided herein, as defined by the Kabat numbering scheme.

[0052] Exemplary humanized antibodies are provided herein. Additional anti-SCF antibodies comprising the heavy and light chain CDRs provided herein, or variants thereof, can be generated using any human framework sequences and are encompassed by the present invention. In one embodiment, framework sequences suitable for use in the present invention include framework sequences structurally similar to those provided herein. Further modifications in the framework regions can be made to improve the properties of the antibodies provided herein. Such further framework modifications can include chemical modifications, point mutations to reduce immunogenicity or remove T-cell epitopes, or backmutations to residues within the original germline sequence.

[0053] In some embodiments, such framework modifications include modifications corresponding to the mutations exemplified herein, including backmutations to germline sequences. For example, in one embodiment, one or more amino acids in the human framework regions of the VH and / or VL of a humanized antibody provided herein are backmutated to the corresponding amino acid in the parent murine antibody. The invention also encompasses humanized antibodies that bind to SCF (e.g., SCF248) and contain framework modifications corresponding to the exemplary modifications described herein for any suitable framework sequence, as well as other framework modifications that otherwise improve the properties of the antibody. In other embodiments, the antibodies provided herein contain one or more mutations that improve stability, improve solubility, alter glycosylation, and / or reduce immunogenicity, e.g., by targeted amino acid changes that reduce deamidation or oxidation, reduce isomerization, optimize hydrophobic core and / or charge cluster residues, remove hydrophobic surface residues, optimize residues involved in the interface between the variable heavy and variable light chains, and / or modify the isoelectric point.

[0054] The anti-SCF antibodies and fragments thereof provided herein can further comprise Fc region modifications to alter effector function. Fc modifications can be amino acid insertions, deletions, or substitutions, or chemical modifications. For example, Fc region modifications can be made to increase or decrease complement fixation, increase or decrease antibody-dependent cellular cytotoxicity, or increase or decrease the half-life of the antibody. Some Fc modifications increase or decrease the affinity of the antibody for an Fcγ receptor (e.g., FcγRI, FcγRII, FcγRIII, or FcRn). Various Fc modifications have been described in the art (e.g., Shields et al., J Biol. Chem 276;6591 (2001); Tai et al. Blood 119;2074 (2012); Spiekermann et al. J Exp. Med 196;303 (2002); Moore et al. mAbs 2:2;181 (2010); Medzihradsky Methods in Molecular Biology 446;293 (2008); Mannan et al. Drug Metabolism and Disposition 35;86 (2007); and Idusogie et al. J Immunol 164;4178 (2000). In some embodiments, the glycosylation pattern of the Fc region is altered. In other embodiments, the Fc region is modified by pegylation (e.g., by reacting the antibody or fragment thereof with polyethylene glycol (PEG)). Exemplary Fc modifications include modifications at one or more amino acid positions selected from the group consisting of 228, 233, 234, 235, 236, 241, 248, 265, 297, 309, 331, and 409 (Kabat numbering; Kabat et al., Sequences of Immunological Interest, Fifth Edition, National Institute of Health, Bethesda, Md. (1991). In some embodiments, the antibody has a modification to reduce or eliminate effector function. In some embodiments, the antibody is an IgG1 antibody with one or more Fc modifications selected from the group consisting of E233P, L234V, L234A, L235V, L235A, G236 (deletion), D265A, D270A, N297A, and N297Q. In some embodiments, the antibody is an IgG4 antibody with one or more Fc modifications selected from the group consisting of S228P, E233P, F234A, F234V, L235A, L235V, S241P, L248E, D265A, D265T, L309L, and R409K. In some embodiments, the anti-SCF antibodies provided herein comprise an S241P mutation and an L248E mutation.

[0055] In various embodiments, the present disclosure provides an antibody provided herein comprising a human IgG4 constant region set forth in SEQ ID NOs: 40 and 41. In various embodiments, the present disclosure provides an antibody comprising at least about 80%, at least about 85%, at least about 90%, at least about 95%, or about 99% sequence identity to SEQ ID NOs: 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50. In various embodiments, the present disclosure provides an antibody comprising a heavy chain set forth in SEQ ID NO: 40 and a light chain set forth in SEQ ID NO: 41. In various embodiments, the present disclosure provides an antibody comprising a heavy chain set forth in SEQ ID NO: 42, 43, 44, 45, or 46 and a light chain set forth in SEQ ID NO: 47, 48, 49, or 50. In various embodiments, the present disclosure provides an antibody comprising a heavy chain set forth in SEQ ID NO: 42 and a light chain set forth in SEQ ID NO: 49. In various embodiments, the present disclosure provides an antibody comprising a heavy chain set forth in SEQ ID NO: 43 and a light chain set forth in SEQ ID NO: 49. In embodiments, the present disclosure provides an antibody comprising a heavy chain set forth in SEQ ID NO: 44 and a light chain set forth in SEQ ID NO: 49. In embodiments, the present disclosure provides an antibody comprising a heavy chain set forth in SEQ ID NO: 45 and a light chain set forth in SEQ ID NO: 49. In embodiments, the present disclosure provides an antibody comprising a heavy chain set forth in SEQ ID NO: 46 and a light chain set forth in SEQ ID NO: 49.

[0056] In some embodiments, the present disclosure provides methods for generating antibodies that specifically bind to SCF248. The SCF248 isoform of SCF contains exon 6, which contains a cleavage site between two alanine residues (amino acids 16 and 17 of SEQ ID NO: 34, which shows the amino acid sequence of exon 6). Previous anti-SCF antibodies were generated by immunizing mice with a peptide spanning exon 6 and part of exon 7 (see, e.g., U.S. Patent No. 8,911,729, incorporated herein by reference in its entirety for all purposes). Because SCF220 is associated with homeostatic activity, any cross-reactivity with SCF220 is considered harmful, resulting in various off-target effects in subjects. Advantageously, the antibodies provided herein bind to SCF248 with high specificity. In some embodiments, the antibodies provided herein are specific for SCF248 and do not bind to SCF220. Thus, the antibodies provided herein can specifically inhibit the interaction between SCF248 and c-Kit, which induces and perpetuates chronic inflammatory responses and fibrosis. Furthermore, the antibodies provided herein can reduce the interaction between SCF248 and c-Kit by specifically inducing the internalization of SCF. Thus, in some embodiments, the present disclosure provides antibodies specific for SCF248 that are safe and effective for various inflammatory and fibrotic diseases discussed herein and known in the art.

[0057] For preparation of monoclonal antibodies, any technique which provides for the production of antibody molecules by continuous cell lines in culture can be used (see, e.g., Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring, NY). (See, e.g., Kozbor et al., J. Immunol. 1999, 14:131-132, 2001). Such techniques include, but are not limited to, the hybridoma technique originally developed by Kohler and Milstein, as well as the trioma technique, the human B cell hybridoma technique (see, e.g., Kozbor et al., J. Immunol. 1999, 14:131-132, 2001). al., Immunol. Today, 4:72 (1983)), and EBV-hybridoma techniques for producing human monoclonal antibodies (Cole et al., Monoclonal Antibodies and Cancer Therapy, Alan R. Liss, Inc., pp. 77-96 (1985)). Alternatively, antibodies may be made by recombinant DNA methods. In some embodiments, antibodies in accordance with the present disclosure can be made by isolating monoclonal antibodies from phage display libraries using, for example, the techniques described in Clackson et al., Nature 352:624-28 (1991) and Marks et al., J. Mol. Biol. 222(3):581-97 (1991). In some embodiments, the antibodies are fully human, constructed by combining Fv clone variable domain sequence(s) selected from human-derived phage display or yeast display library(ies) with known human constant domain sequence(s).

[0058] In some embodiments provided herein, antibodies are prepared from hybridomas. Using the hybridoma method, mice, hamsters, or other suitable host animals are immunized by injecting an immunizing peptide to induce lymphocytes to produce antibodies that specifically bind to the immunizing antigen. Alternatively, lymphocytes may be immunized in vitro. After immunization, lymphocytes are isolated and fused with a suitable myeloma cell line, for example, using polyethylene glycol, to form hybridoma cells, which can then be selected away from unfused lymphocytes and myeloma cells. The hybridomas produce monoclonal antibodies specifically directed against the selected antigen as determined by immunoprecipitation, immunoblotting, or an in vitro binding assay (e.g., radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA)), and the hybridomas can then be grown in vitro (e.g., in culture) using standard methods (Goding, Monoclonal Antibodies: Principles and Practice, Academic Press, 1986), or grown in vivo as ascites tumors in an animal. Monoclonal antibodies can then be purified from the culture medium or ascites fluid as described above for polyclonal antibodies.

[0059] In some embodiments, the antibodies provided herein are produced using a mouse hybridoma system. Hybridoma production in mice is a well-established procedure. Immunization protocols and techniques for isolating immunized spleen cells for fusion are known in the art. Fusion partners (e.g., mouse myeloma cells) and fusion procedures are also known. Embodiments of the techniques herein provide antibodies (e.g., monoclonal antibodies) produced from hybridomas prepared by immunizing mice with peptides that are portions or fragments of the SCF protein.

[0060] In some embodiments, antibodies specific for SCF248 provided herein are generated by immunizing mice with a peptide having an amino acid sequence located primarily or exclusively within exon 6. For example, the immunizing peptide includes any stretch of five or more amino acids within SEQ ID NO: 34. As another example, the immunizing peptide includes any stretch of five or more amino acids beginning at amino acid position 20 of SEQ ID NO: 29. As another example, the immunizing peptide includes any stretch of five or more amino acids beginning at amino acid position 20 of SEQ ID NO: 29 and ending at any one of positions 25-38 of SEQ ID NO: 29. Thus, in some embodiments, the immunizing peptide includes the amino acid sequence of exon 6 from the cleavage site onwards, and is either entirely contained within exon 6 or includes only one, two, three, four, or five amino acids of exon 7. In some embodiments, the immunizing peptide comprises or consists of SEQ ID NO: 30. In some embodiments, the immunizing peptide comprises any of the peptides provided herein or conservative variants thereof. Conservative variants may include one, two, three, four, or five amino acid substitutions or deletions, or a combination thereof. As noted above, in some embodiments, antibodies generated using the immunizing peptides provided herein have an epitope that resides entirely or predominantly within exon 6. By "predominantly within" is meant that at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% of the peptide resides within exon 6. In some embodiments, the epitope begins at the exon 6 cleavage site (i.e., between the alanines at amino acid positions 19 and 20 of SEQ ID NO:29) and extends to the end of exon 6. In some embodiments, the epitope begins at the exon 6 cleavage site and extends to the first, second, third, fourth, or fifth N-terminal amino acid of the transmembrane domain. In some embodiments, the epitope comprises or consists of SEQ ID NO:33. In some embodiments, the antibody designated herein as 5H10 (including murine, chimeric, and humanized 5H10 antibodies) binds to an epitope of SCF comprising or consisting of SEQ ID NO:33.

[0061] In some embodiments, the methods provided herein were used to generate the antibody designated herein as 5H10. In some embodiments, the antibody "5H10" is also designated herein as "OpSCF." Antibody 5H10 advantageously binds with high specificity to SCF248 and does not bind to SCF220. The amino acid sequences of the murine parent antibody 5H10 and its humanized variants are provided herein (see Table 1).

[0062] In one embodiment, the invention provides bispecific or multispecific antibodies specific for SCF and at least one other antigen or epitope. The anti-SCF antibodies and fragments thereof provided herein can be tested for binding to SCF using the binding assays provided herein or any other binding assay known in the art.

[0063] Unless otherwise indicated, the practice of the present invention will employ techniques well known in the art and described in, for example, Methods in Molecular Biology, Humana Press; Molecular Cloning: A Laboratory Manual, second edition (Sambrook et al., 1989); Current Protocols in Immunology (JEColiganet al., eds., 1991); Immunobiology (CA Janeway and P. Travers, 1997); Antibodies (P. Finch, 1997); Antibodies: a practical approach (D. Catty., ed., IRL Press, 1988-1989); Monoclonal antibodies: a practical approach (P. Shepherd and C. Dean, eds., Oxford University Press, 2000); Phage display: a laboratory manual (C. Barbas III et al., Cold Spring Harbor Laboratory Press, 2001); and Using antibodies: a laboratory manual (E. Harlow and D. Lane (Cold Spring Harbor Laboratory Press, 1999)) are used.

[0064] Treatment method In one aspect, the present disclosure provides methods for treating and / or preventing any disease or condition associated with immune cell migration, activation, and / or proliferation via the interaction of SCF248 with c-Kit on immune cells. Accordingly, in some embodiments, the present disclosure provides methods for inhibiting or preventing immune cell activation, as well as methods for treating or preventing various diseases and disorders involving inflammation by reducing or preventing the accumulation of immune cells in an organ or tissue. In some embodiments, the immune cells are selected from the group consisting of mast cells, innate lymphoid cells (ILCs (e.g., ILC2 or ILC3 cells)), and eosinophils.

[0065] As used herein, the term "treatment" or "treating" refers to both therapeutic and prophylactic treatment. Subjects in need of treatment include those who already have a disease or condition, as well as those who are at risk of developing a disease or condition and for whom the prevention, delay, or alleviation of the disease or condition is the goal. As used herein, the term "subject" refers to mammals (e.g., rodents, cats, dogs, and primates). Preferably, the subject according to the present invention is a human. As used herein, the term "therapeutically effective amount" refers to the amount of a compound or composition required to provide a therapeutic and / or prophylactic benefit to a subject.

[0066] In one aspect, the present invention provides a method for treating an inflammatory, fibrotic, and / or tissue remodeling disease in a subject. In some embodiments, the inflammatory disease is a chronic inflammatory disease.

[0067] Chronic inflammatory, fibrotic, and tissue remodeling diseases include diseases of the lung, kidney, liver, heart, skin, connective tissue, and other tissues. Exemplary inflammatory, fibrotic, or tissue remodeling diseases include, but are not limited to, pulmonary fibrosis (e.g., idiopathic pulmonary fibrosis (IPF), scleroderma pulmonary fibrosis, scleroderma-related interstitial lung disease (SSc-ILD), pulmonary fibrosis associated with pulmonary infection or pneumonia, pulmonary fibrosis associated with systemic lupus erythematosus and / or rheumatoid arthritis, sarcoidosis), chronic obstructive pulmonary disease (COPD), acute respiratory distress syndrome (ARDS), cystic fibrosis, peribronchial fibrosis, bleomycin lung, hypersensitivity pneumonitis, asthma, fibrothorax, mediastinal fibrosis, chronic sinusitis, urticaria (e.g., chronic idiopathic urticaria), atopic dermatitis, and the like. Dermatitis, dermatomyositis, nodular subepidermal fibrosis, scleroderma, keloids, renal fibrosis, chronic kidney disease, glomerulonephritis, chronic renal allograft rejection, nephropathy (e.g., IgA nephropathy, focal segmental glomerulosclerosis, rapidly progressive glomerulonephritis, crescentic glomerulonephritis, lupus nephritis, hypertensive nephropathy, or diabetic nephropathy), nonalcoholic steatohepatitis (NASH), liver cirrhosis, liver fibrosis, primary sclerosing cholangitis, primary biliary cirrhosis, fibromyalgia, gingival fibrosis, radiation-induced fibrosis, eosinophilic esophagitis, arthrofibrosis, and atrial fibrosis, endomyocardial fibrosis, parenchymal fibrosis, fibrous histiocytoma, or glial scarring.

[0068] In some embodiments, the antibodies and fragments thereof disclosed herein can be administered to a subject by at least one route selected from parenteral, subcutaneous, intramuscular, intravenous, intra-articular, intrabronchial, intraperitoneal, intracapsular, intrachondral, intrasinus, intracavity, intracerebral, intraventricular, intracolonic, intracervical, intragastric, intrahepatic, intramyocardial, intraosseous, intrapelvic, intrapericardial, intraperitoneal, intrapleural, intraprostatic, intrapulmonary, intrarectal, intrarenal, intraretinal, intraspinal, intrasynovial, intrathoracic, intratympanic, intrauterine, intravesical, intravitreal, bolus, subconjunctival, oral, vaginal, rectal, buccal, sublingual, intranasal, intratumor, and transdermal.

[0069] In various embodiments, the antibodies and fragments thereof disclosed herein can be administered to a subject in need thereof in combination with one or more additional therapies, which can be procedures, such as surgical procedures, or therapeutic agents (e.g., drugs designed to alleviate or reduce the symptoms of a disease or disorder associated with fibrosis and / or inflammation).

[0070] The present invention will be further described by reference to the following examples, although it should be noted that these examples, like the above-described embodiments, are illustrative and should not be construed as limiting the scope of the invention in any way. [Example]

[0071] The following examples are presented for the purpose of illustrating various embodiments of the present disclosure and are not intended to limit the disclosure in any way. Those skilled in the art will recognize that variations of the examples and other uses are encompassed within the spirit of the disclosure as defined by the claims.

[0072] An overview of the tissue injury / disease process is summarized in Figure 1. The disease process initiates inflammation. Cytokine production by c-Kit+ immune cells induces fibroblast transformation into activated myofibroblasts, which express SCF248 on their surface. Expression of SCF248 on the surface of myofibroblasts and other cells activates more immune cells, resulting in the release of IL-4, IL-9, IL-13, IL-25, TGFβ, and other cytokines, perpetuating inflammation. Myofibroblasts secrete extracellular matrix proteins, collagen, and fibronectin, leading to fibrosis and remodeling disorders (e.g., pulmonary fibrosis, dermal fibrosis, severe asthma, and other diseases).

[0073] An exemplary mechanism for an antibody of the disclosure that targets SCF248 (which antibody is referred to herein as OpSCF and / or 5H10) is summarized in FIG.

[0074] As shown above, SCF has two isoforms resulting from alternative splicing: SCF248 and SCF220. SCF248 and SCF220 differ by exon 6. SCF220 is associated with homeostatic functions, while SCF248 is associated with inflammation and fibrosis. SCF248 activates immune cells during inflammation and is sometimes referred to as "lytic SCF." SCF248 is expressed in various cell types, including myofibroblasts, activated epithelium, endothelium, macrophages, eosinophils, mast cells, and monocytes (FIG. 3). The SCF248 isoform results in the cleavage of a monomeric truncated extracellular domain called SCF165. The amino acid sequence of exon 6 is provided herein as SEQ ID NO: 34.

[0075] Example 1: Production of anti-SCF mAbs using hybridoma technology A peptide containing ASSLRNDSSSSNRKAKNPPGD (SEQ ID NO: 30) was used to generate antibodies that bind to SCF248. The immunizing peptide included a portion of exon 6, i.e., the SCF248 isoform of stem cell factor. Specifically, the immunizing peptide included a portion of exon 6 beginning after the cleavage site defined herein. Mice were immunized with the peptide set forth in SEQ ID NO: 30 using standard protocols. Quantitation of high-titer serum antibodies indicated successful immunization and the generation of fusion hybridomas. Culture supernatants were analyzed for SCF-specific antibodies from individual clones and selected based on specificity. Hybridomas producing specific monoclonal antibodies against the peptide were expanded, and the most highly titered monoclonals were then tested in biologically relevant cultures. Antibody 5H10 was highly specific for SCF248 without cross-reactivity with SCF220. No other monoclonal antibodies produced by hybridomas were highly specific for SCF248 without cross-reactivity with SCF220. Therefore, 5H10 was selected for further characterization, development, and chimerization, and subsequent humanization.

[0076] Example 2. Binding of 5H10 to the complete extracellular domain of SCF248 The mouse 5H10 antibody, obtained as described in Example 1, was directly conjugated to a fluorescent marker, and the labeled antibody was incubated with S1 / S14 hSCF248 cells expressing SCF248, S1 / S14 hSCF220 cells expressing SCF220, or control cells not expressing SCF. Binding of the labeled antibody to the cells was assessed by flow cytometry. The specificity of 5H10 for SCF248 and the lack of cross-reactivity with SCF220 are shown in Figure 4A.

[0077] The binding of the mouse 5H10 antibody to a truncated extracellular domain (ECD) of SCF containing only amino acids 1 to 165 and to the complete ECD of SCF containing amino acids 1 to 194 was assessed by ELISA. This antibody bound to the complete SCF ECD but not to the truncated SCF ECD (Fig. 4B), indicating that this antibody is specific for the complete extracellular domain and does not bind to the monomeric truncated ECD circulating in the blood.

[0078] To evaluate the ability of the 2G8 and 5H10 antibodies to internalize SCF248 on myofibroblasts, these antibodies were labeled with pHrodo red, which is colorless at neutral pH and fluoresces red at the low pH of the endosome. The labeled antibodies were incubated with cultured human IPF myofibroblasts for 45 minutes, and the red fluorescence was visualized by microscopy. As shown in Figure 5, the dye-labeled antibodies were rapidly internalized, whereas the control IgG was not. 5H10 was internalized more rapidly than 2G8, resulting in higher fluorescence.

[0079] SCF induces c-kit to signal via two distinct pathways: the MEK / ERK pathway and the P13K / AKT pathway. We investigated whether the murine 5H10 antibody inhibits intracellular signaling in c-kit-positive cells via one or both of these pathways. Eosinophils were incubated with SCF248-expressing cell lines in the presence of either 5H10 or an IgG control, and phosphorylated protein expression was measured using a BioRad Bio-Plex assay system. 5H10 significantly reduced phospho-MEK and phospho-AKT levels, indicating that the antibody significantly reduced c-kit-mediated intracellular signaling (Figure 6).

[0080] Taken together, these studies demonstrate that antibody 5H10 specifically binds and internalizes SCF248 without cross-reacting with SCF220 isoforms or the truncated ECD. Furthermore, 5H10 significantly inhibits intracellular signaling pathways that perpetuate inflammation in c-kit-positive cells.

[0081] Example 3: Humanization of murine antibody 5H10 Chimeric antibodies derived from 5H10 were produced by subcloning the heavy and light chain variable domains into a vector with a human IgG4 backbone. The chimeric antibodies were expressed and purified using standard protocols. 2G8 is a previously developed antibody that binds to SCF248 and SCF220 and contains a lambda light chain. The chimeric heavy and light chains of 2G8 were designated VH0 and VL0, respectively. The SCF248-specific antibody 5H10 provided herein contains a kappa light chain. The chimeric heavy and light chains of 5H10 were designated VH0 and VK0, respectively.

[0082] This chimeric antibody was humanized. The humanized heavy chains retained the same complementarity-determining regions (CDRs) but more "human" framework regions. Several humanized variants of each of the 2G8 and 5H10 variable heavy chains (referred to herein as VH1, VH2, VH3, VH4, and VH5) were generated. Humanized kappa light chain variants of 5H10 (referred to herein as VK1, VK2, VK3, and VK4) were also generated. The humanized lambda light chains of 2G8 were designated VL1, VL2, VL3, and VL4. The chimeric and humanized light and heavy chain combinations of 2G8 and 5H10 tested are shown in Tables 2 and 3, respectively. As shown in Table 2, certain heavy and light chain combinations of the 5H10 antibody variants conferred high binding to hSCF248. The binding data used to determine the binding scores is presented in Example 5 below. [Table 2] [Table 3]

[0083] Binding affinity was also assessed using BiaCore analysis. BiaCore data showed that the affinities of all humanized 5H10 antibodies with VK1, VK2, or VK3 light chains for the immobilized SCF248 peptide antigen were very similar to the binding affinity of the parental murine 5H10 using this assay. Humanized 5H10 antibodies with VK4 light chains did not bind to this peptide. [Table 4]

[0084] Example 4. Evaluation of anti-SCF chimeric antibody binding by flow cytometry The binding of the chimeric antibodies 2G8 and 5H10 was tested using S1 / S14 hSCF248 cells, a transfected cell line expressing SCF248. An anti-SCF antibody was used as a positive control for SCF binding. A human IgG4 antibody was used as a negative isotype control. Early passage (P3) S1 / S14 hSCF248 cells were compared with late passage (P5) cells. As expected, the negative control (human IgG4 antibody) did not bind to either cell population. 5H10 bound to cells at early passages (Figure 7A) but was not detected at later passages (Figure 7B). This is due to the loss of SCF248 expression with multiple passages. Similarly, the maximum mean fluorescence intensity (MFI) detected with 10 μg / mL 2G8 was approximately 4-fold reduced at P5 compared to P3.

[0085] Similar observations were made with cells treated with hygromycin B. Hygromycin selection was used to enrich for the fraction of S1 / S14 cells expressing the relevant human SCF sequence. The binding of two chimeric anti-SCF mAbs, 2G8 and 5H10, and a humanized anti-SCF antibody was assessed by flow cytometry. The binding and specificity of the chimeric antibodies 2G8 and 5H10 were tested using S1 / S14 hSCF248 and S1 / S14 hSCF220 cells (SCF248+ and SCF220+, respectively). The anti-SCF antibody was used as a positive control for SCF binding. A human IgG4 antibody was used as a negative isotype control. Hygromycin B-treated cells were compared with early passage (P3) cells. The maximum mean fluorescence intensity (MFI) and antibody dose-response curves were similar to those of early passage (P3) cells (Figures 8A and 8B).

[0086] Example 5. Binding evaluation of 2G8 and 5H10 humanized mAbs by flow cytometry The binding and specificity of chimeric antibodies 2G8 and 5H10, as well as their humanized variants, were tested using S1 / S14 hSCF248 cells and S1 / S14 hSCF220 cells (SCF248+ and SCF220+, respectively). In both sets of experiments, a human IgG4 antibody was used as a negative isotype control antibody. The isotype control did not bind to either S1 / S14 hSCF248 or S1 / S14 hSCF220 cells (Figures 9A, 9B, 10A, and 10B). A commercially available anti-SCF antibody (Abcam, catalog no. EP665Y / ab52603) was found to bind to S1 / S14 hSCF220 cells and weakly bind to S1 / S14 hSCF248 cells (only at a 1:25 dilution) (Figures 9A, 9B, 10A, and 10B). 2G8 VH0 / VL0 (chimeric 2G8) showed stronger binding than its humanized clones. However, 2G8 VH0 / VL0 also showed binding to S1 / S14 hSCF220 cells at antibody concentrations of 3.3 and 10 μg / mL (Figures 9A and 9B). 5H10 VH0 / VK0 showed higher binding than the humanized clones 5H10 VH3 / VK2, 5H10 VH3 / VK3, 5H10 VH4 / VK2, 5H10 VH4 / VK3, 5H10 VH5 / VK2, and 5H10 VH5 / VK3 (Figure 10A). Neither 5H10 nor any of its humanized variants were observed to bind to S1 / S14 hSCF220 cells (Figure 10B). The 50% maximal binding (BC) in this study was 0.01. 50 ) differences between the indicated 5H10 humanized variants are shown in Table 5. Binding of the 5H10 clone reached saturation at 3.3 μg / mL on S1 / S14 hSCF248 cells (Figure 10A). [Table 5]

[0087] The binding of additional humanized variants of 5H10 and 2G8 was tested using S1 / S14 hSCF248 cells. 2G8 VH0 / VL0 (chimeric form) showed stronger cell binding than the humanized variants (Figure 11A). The binding profiles of the humanized 5H10 clones 5H10 VH1 / VK1, 5H10 VH1 / VK2, 5H10 VH1 / VK3, 5H10 VH2 / VK2, and 5H10 VH2 / VK3 were comparable to that of 5H10 VH0 / VK0 (Figure 11B). Consistent with the Biacore data presented above, the humanized variant 5H10 VH1 / VK4 lost target binding (Figure 11B). The isotype control did not bind to S1 / S14 hSCF248 cells. Based on the data presented in these studies, the humanized 2G8 and 5H10 mAbs were assigned binding scores for S1 / S14 hSCF248 cells and are presented in Tables 2 and 3 above.

[0088] In a separate experiment, the binding of 5H10 clones VH1 / VK3, VH2 / VK3, VH3 / VK3, VH4 / VK3, and VH5 / VK3 to SCF248-expressing cell lines was assessed by flow cytometry. As shown in Figures 11C and 11D, VH1 / VK3 and VH2 / VK3 exhibited high binding, maximal at 1 μg / mL. The negative control was secondary antibody alone. No binding was observed to the control SCF220-expressing cell line (not shown).

[0089] Example 6. In vitro blocking of the interaction between SCF and c-kit The humanized 5H10 antibody was tested in vitro for its ability to inhibit the SCF-c-kit interaction and the inflammatory feed-forward loop. Cultured human IPF myofibroblasts (Mfb), which express surface SCF248, were overlaid with LAD2 mast cells, an SCF-responsive cell line. Without any other intervention, Mfb stimulates LAD2 cells, which in turn stimulate Mfb to produce additional cytokines and extracellular matrix proteins. In this assay, the readouts of the inflammation and feed-forward loop are mRNA for CCL11, collagens 1 and 3, and fibronectin.

[0090] The murine 5H10 antibody and humanized (VH1 / VK3, VH2 / VK3, VH3 / VK3, VH4 / VK3, and VH5 / VK3) 5H10 antibodies were preincubated with Mfb at concentrations of 1 μg / mL and 10 μg / mL to evaluate their ability to inhibit the feed-forward loop. The results are shown in Figures 12A-12D. The humanized VH1 / VK3 antibody consistently inhibited the SCF-c-kit interaction, even at low concentrations.

[0091] To evaluate the ability of the humanized antibodies (5H10, VH1 / VK3, and VH2 / VK3) to internalize SCF248 on myofibroblasts, these antibodies were labeled with pHrodo red, which is colorless at neutral pH and fluoresces red at the low pH of the endosome. The labeled antibodies were incubated with cultured human IPF myofibroblasts for 45 minutes, and the red fluorescence was visualized by microscopy. As shown in Figure 13, the humanized antibodies were rapidly internalized, as were the parental mouse antibody 5H10 and the chimeric antibody (VH0 / VK0).

[0092] Example 7. Immunogenicity of chimeric antibody 5H10 and humanized lead candidates The immunogenicity of five humanized antibodies, 5H10 VH1 / VK3, 5H10 VH2 / VK3, 5H10 VH3 / VK3, 5H10 VH4 / VK3, and 5H10 VH5 / VK3, was compared with that of the chimeric antibody 5H10 VH0 / VK0.

[0093] An initial assessment of any cytotoxic effect of the samples on PBMC viability was performed on the five donors used in the EpiScreen™ time course assay. CD8+ T cell-depleted PBMCs were incubated with the samples, and cell viability was quantified on day 7 using a Luna-FL™ Automated Cell Counter. The mean viability of PBMCs from five mAb-treated donors was similar to that of cells treated with medium alone, ranging from 83% to 90% (Figure 14). KLH (Pierce, Life Technologies, UK) was used as the neoantigen. Exenatide (Bydueon, AstraZeneca, UK) was used as the clinical benchmark control. Figures 15A-H and Table 6 demonstrate the results of an EpiScreen™ time-course T cell proliferation assay of CD4+ T cell responses induced by samples and controls. The clinical benchmark exenatide and the neoantigen KLH both induced positive proliferative responses. 5H10 VH1 / VK3, 5H10 VH2 / VK3, and 5H10 VH3 / VK3 induced low rates of positive responses (SI ≥ 1.90, p < 0.05), ranging from 4% to 8%. Sample 5H10 VH5 / VK3 induced higher positive responses in 17% of the donor cohort. 5H10 VH0 / VK0 and 5H10 VH4 / VK3 did not induce any positive responses. The mean magnitude of the positive T cell proliferative responses ranged from 2.14 to 3.61 across all samples (Table 7).

[0094] An analysis of variance (ANOVA) of the entire proliferation data set (using the maximum magnitude of proliferation on days 5-8) was used to determine whether there were statistically significant differences in the maximum magnitude of the CD4+ T cell response to the test conditions relative to each other and relative to the clinical benchmark, exenatide (Figure 16). The maximum magnitude of the T cell proliferation response to exenatide was statistically higher than the responses to all samples (Table 8). [Table 6] [Table 7] [Table 8]

[0095] In summary, the risk of clinical immunogenicity was determined by measuring ex vivo T cell responses in the EpiScreen™ time-course T cell assay using peripheral blood mononuclear cells (PBMCs) isolated from 24 healthy donors representative of European and North American populations (based on HLA allotypes). T cell responses were measured using a proliferation assay ([ 3 H-thymidine incorporation) were measured. The results showed that four of the lead humanized antibodies had low clinical immunogenicity potential.

[0096] Example 8. Evaluation of murine 5H10 in a bleomycin model of pulmonary inflammation and fibrosis The in vivo efficacy of 5H10 was evaluated using an animal model of bleomycin pulmonary fibrosis. Bleomycin is a chemotherapeutic agent that causes pulmonary fibrosis in humans and animals. C57BL6 mice were administered bleomycin intratracheally on day 1 and 20 mg / kg 5H10 or an isotype-matched control antibody intraperitoneally on days 8 and 12. Samples were collected on day 17. 5H10-treated animals showed significant improvement in lung histology (Figure 17), reduction in lung hydroxyproline (a quantitative measure of fibrosis; Figure 18), maintenance of body weight over time (Figure 19), reduction in mRNA for inflammatory cytokines and myofibroblast activation markers (Figure 20), and reduction in lung mast cells, eosinophils, and ILC2 lymphocytes (Figure 21). Pulmonary function tests also significantly improved (Figure 22). Thus, this study demonstrated that 5H10 was effective in reducing fibrosis and inflammation and improving lung function in an in vivo model of pulmonary fibrosis.

[0097] Example 9. Evaluation of humanized 5H10 antibody in a chronic allergic asthma model The humanized 5H10 antibody was tested in an in vivo model of chronic allergic asthma. Mice were sensitized intraperitoneally and subcutaneously with cockroach antigen (CRA), followed by intranasal boosting on days 14, 18, 22, and 26. On days 26, 29, 32, and 34, mice received intraperitoneal administration of the indicated humanized 5H10 antibody or a PBS control at 20 mg / kg, or an irrelevant control antibody. On days 30 and 34, CRA was also administered intratracheally. Samples were collected on day 35. A schematic diagram of the study design is shown in Figure 23.

[0098] Animals treated with antibody 5H10 VH1 / VK3 had significantly less airway resistance than PBS controls and other humanized variant antibodies (Figure 24A). Furthermore, animals treated with VH1 / VK3 or VH2 / VK3 showed a significant decrease in lung IL-13 mRNA compared to chronic asthma controls (i.e., compared to animals administered CRA without any antibody treatment (PBS control)) (Figure 24B). Furthermore, VH1 / VK3 antibody-treated animals showed a significant decrease in matrix gene expression (collagen 1 mRNA shown in Figure 24C, collagen 3 mRNA shown in Figure 24D). SCF248 also showed a significant decrease in lung IL-13 mRNA compared to chronic asthma controls (i.e., compared to animals administered CRA without any antibody treatment (PBS control)). mRNA expression was also significantly reduced in VH1 / VK3-treated animals (Figure 24E). Figures 25A, 25B, and 25C show that VH1 / VK3 administration reduced the mRNA levels of the mucus protein Gob5 and the cytokines IL-13 and IL-5 at concentrations of 1 mg / kg and 5 mg / kg. Thus, these data demonstrate that the VH1 / VK3 antibody blocks cytokine and matrix gene expression in vivo in a chronic asthma model.

[0099] Example 12. Phase 1a clinical trial to evaluate the safety, pharmacokinetics, and pharmacodynamics of anti-SCF248 antibodies The Phase 1a trial will enroll 110 healthy volunteers, with the primary objectives being to obtain safety evaluation and accurate pharmacokinetic and pharmacodynamic data for the humanized anti-SCF248 antibody 5H10.

[0100] Single or multiple ascending doses of humanized 5H10 or placebo will be administered either intravenously or subcutaneously. Six to eight subjects will be included in each cohort. Starting doses will be based on toxicology studies conducted in accordance with GLP (Good Laboratory Practice). Baseline pharmacokinetics will be obtained throughout development and product lifespan. Pharmacodynamics will be assessed by assessing serum inflammatory markers such as SCF165, as well as circulating c-kit+ cell counts, including mast cell precursors and type 2 innate lymphoid (ILC2) cells.

[0101] Example 13. Clinical trial to evaluate the safety and efficacy of anti-SCF248 antibodies in patients The clinical trial will enroll patients with inflammatory disorders (e.g., atopic dermatitis, chronic urticaria, pulmonary fibrosis, and / or other conditions). One objective of the study is to establish a dose-response relationship between the humanized 5H10 antibody and pharmacodynamic markers in affected patients, such as the number of circulating c-kit+ cells (e.g., mast cell precursors and type 2 innate lymphoid cells (ILC2) cells). Inflammatory biomarkers (e.g., ADAM8, CCL17, EPX, RNASE3, CCL2, CCL5, tryptase, histamine, and SCF165) will also be measured.

[0102] The 5H10 antibody will be administered in a single ascending dose to each subject group. The starting dose will be based on pharmacodynamic biomarkers from the Phase 1a study. Patients will receive treatment for one, two, three, or more months. The results of this study will demonstrate that the humanized 5H10 antibody is effective in stabilizing and / or treating and / or preventing the progression of inflammatory and fibrotic disorders.

[0103] All publications, patents, and patent applications cited herein are specifically incorporated herein by reference in their entirety. While the described invention has been described with reference to specific embodiments thereof, those skilled in the art will recognize that various modifications may be made and equivalents may be substituted without departing from the spirit and scope of the invention. In addition, many modifications may be made to adapt a particular situation, material, composition of matter, process, process step(s) to the objective, spirit, and scope of the described invention. All such modifications are intended to be within the scope of the claims appended hereto. The present invention provides, for example, the following items. (Item 1) An antibody or fragment thereof that specifically binds to stem cell factor (SCF), wherein the antibody comprises heavy chain CDR1, CDR2, and CDR3 comprising SEQ ID NOs: 1, 2, and 3, respectively. (Item 2) 2. The antibody or fragment thereof of item 1, wherein the antibody further comprises light chain CDR1, CDR2, and CDR3 comprising SEQ ID NOs: 4, 5, and 6, respectively. (Item 3) 3. The antibody or fragment thereof of item 1 or 2, wherein the antibody comprises a heavy chain variable region having at least 80% identity to a sequence selected from SEQ ID NOs: 7, 8, 9, 10, and 11. (Item 4) 4. The antibody or fragment thereof according to item 3, wherein the antibody comprises a heavy chain variable region having at least 90% identity to a sequence selected from SEQ ID NOs: 7, 8, 9, 10, and 11. (Item 5) 3. The antibody or fragment thereof of item 1 or 2, wherein the antibody comprises a light chain variable region having at least 80% identity to a sequence selected from SEQ ID NOs: 13, 14, 15, and 16. (Item 6) 6. The antibody or fragment thereof according to item 5, wherein the antibody comprises a light chain variable region having at least 90% identity to a sequence selected from SEQ ID NOs: 13, 14, 15, and 16. (Item 7) 2. The antibody or fragment thereof according to item 1, wherein the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence selected from SEQ ID NOs: 7, 8, 9, 10, and 11, and a light chain variable region amino acid sequence selected from SEQ ID NOs: 13, 14, 15, and 16. (Item 8) 2. The antibody or fragment thereof according to item 1, wherein the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO: 7 and a light chain variable region amino acid sequence set forth in SEQ ID NO: 16. (Item 9) 2. The antibody or fragment thereof according to item 1, wherein the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO: 8 and a light chain variable region amino acid sequence set forth in SEQ ID NO: 16. (Item 10) 2. The antibody or fragment thereof according to item 1, wherein the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO: 9 and a light chain variable region amino acid sequence set forth in SEQ ID NO: 16. (Item 11) 2. The antibody or fragment thereof according to item 1, wherein the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO: 10 and a light chain variable region amino acid sequence set forth in SEQ ID NO: 16. (Item 12) 2. The antibody or fragment thereof according to item 1, wherein the antibody or fragment thereof comprises a heavy chain variable region amino acid sequence set forth in SEQ ID NO: 11 and a light chain variable region amino acid sequence set forth in SEQ ID NO: 16. (Item 13) Item 14. The antibody or fragment thereof according to Item 1 or 2, wherein the antibody is humanized. 14. The antibody or fragment thereof according to any one of items 1 to 13, wherein the antibody is a monoclonal antibody. (Item 15) 15. The antibody of item 14, wherein the antibody comprises a human IgG4 domain. (Item 16) 16. The antibody of item 15, wherein the IgG4 domain comprises an S241P mutation at amino acid residue 241 and an L248E mutation at amino acid residue 248, wherein the numbering of the residues is according to the Kabat numbering system. (Item 17) 17. The antibody according to any one of items 14 to 16, wherein the antibody comprises a heavy chain set forth in SEQ ID NO: 42 and a light chain set forth in SEQ ID NO: 49. (Item 18) 17. The antibody according to any one of items 14 to 16, wherein the antibody comprises a heavy chain set forth in SEQ ID NO: 43 and a light chain set forth in SEQ ID NO: 49. (Item 19) 19. The antibody or fragment thereof according to any one of items 1 to 18, wherein the antibody or fragment thereof blocks the interaction between SCF and c-Kit. (Item 20) 19. The antibody or fragment thereof according to any one of items 1 to 18, wherein the antibody or fragment thereof causes internalization of SCF. (Item 21) 21. The antibody or fragment thereof according to any one of items 1 to 20, wherein the antibody specifically binds to SCF248. (Item 22) 22. The antibody or fragment thereof according to any one of items 1 to 21, wherein the antibody does not bind to SCF220. (Item 23) The fragments may be Fab, F(ab' )2 23. The antibody or fragment thereof according to any one of items 1 to 14 or 17 to 22, wherein the antibody or fragment thereof is selected from a Fab', scFv, and a single domain antibody (sdAb). (Item 24) 24. An isolated nucleic acid molecule encoding the antibody or fragment thereof according to any one of items 1 to 23. (Item 25) 24. An expression vector comprising a nucleic acid segment encoding the antibody or fragment thereof according to any one of items 1 to 23. (Item 26) A recombinant host cell comprising the expression vector of item 25. (Item 27) A method for producing an antibody that specifically binds to stem cell factor isoform 248 (SCF248), comprising immunizing a host animal with a peptide comprising SEQ ID NO: 30 or a fragment thereof, and obtaining the antibody from the immunized host animal. (Item 28) 28. The method of item 27, wherein the fragment comprises at least 5 amino acids. (Item 29) 28. The method of claim 27, wherein the fragment comprises at least 10 amino acids. (Item 30) 28. The method of item 27, wherein the peptide consists of SEQ ID NO: 30. (Item 31) 28. The method of claim 27, wherein the peptide comprises at least 5 amino acids and the N-terminal amino acid of the peptide is alanine at position 1 of SEQ ID NO: 30. (Item 32) 32. The method of claim 31, wherein the peptide comprises at least 10 amino acids. (Item 33) 33. The method according to any one of items 27 to 32, wherein the antibodies from the immunized host animal are obtained from immune cells isolated from the host animal. (Item 34) 34. The method of claim 33, further comprising producing hybridomas using the immune cells. (Item 35) An antibody prepared by the method according to any one of Items 27 to 34, which specifically binds to SCF248. (Item 36) 24. A method for inhibiting chronic inflammation in a subject in need thereof, comprising administering to the subject the antibody or fragment thereof according to any one of items 1 to 23. (Item 37) 24. A method for inhibiting fibrosis in a subject in need thereof, comprising administering to the subject the antibody or fragment thereof according to any one of items 1 to 23. (Item 38) 24. A method for treating a chronic inflammatory or fibrotic disease in a subject in need thereof, the method comprising administering to the subject the antibody of any one of items 1 to 23. (Item 39) 39. The method of item 38, wherein the chronic inflammatory disease or the fibrotic disease is selected from the group consisting of urticaria, atopic dermatitis, non-alcoholic steatohepatitis (NASH), primary sclerosing cholangitis, pulmonary fibrosis, chronic obstructive pulmonary disease (COPD), acute respiratory distress syndrome (ARDS), cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, asthma, bleomycin lung, scleroderma, liver cirrhosis, endomyocardial fibrosis, fibromyalgia, and eosinophilic esophagitis. (Item 40) Item 39. The method of item 39, wherein the pulmonary fibrosis is idiopathic pulmonary fibrosis or scleroderma pulmonary fibrosis. (Item 41) 38. The method of claim 37, wherein the method further comprises administering an additional therapeutic agent to the subject. (Item 42) An antibody or fragment thereof that specifically binds to SCF248, wherein the antibody or fragment thereof binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction between SCF248 and c-Kit. (Item 43) 43. The antibody or fragment thereof according to item 42, wherein the epitope comprises SEQ ID NO: 33. (Item 44) 43. The antibody or fragment thereof according to item 42, wherein the epitope consists of SEQ ID NO: 33. (Item 45) 43. The antibody or fragment thereof of item 42, wherein the antibody or fragment thereof blocks SCF248 from binding to c-Kit on immune cells. (Item 46) 1. A method for inhibiting activation of an immune cell, comprising contacting the immune cell with an antibody or fragment thereof that specifically binds to SCF248, wherein the antibody or fragment thereof binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction of SCF248 with c-Kit. (Item 47) 47. The method of item 46, wherein the method is performed in vitro, ex vivo, or in vivo. (Item 48) 1. A method for inhibiting inflammation in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof that specifically binds to SCF248, wherein the antibody or fragment thereof binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction of SCF248 with c-Kit. (Item 49) 1. A method for inhibiting fibrosis in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof that specifically binds to SCF248, wherein the antibody or fragment thereof binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction of SCF248 with c-Kit. (Item 50) 1. A method for treating an inflammatory disease or disorder in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof that specifically binds to SCF248, wherein the antibody or fragment thereof binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction of SCF248 with c-Kit. (Item 51) 51. The method of item 50, wherein the inflammatory disease or disorder is a chronic inflammatory disease or disorder. (Item 52) 1. A method for treating a fibrotic disease in a subject in need thereof, comprising administering to the subject an antibody or fragment thereof that specifically binds to SCF248, wherein the antibody or fragment thereof binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction of SCF248 with c-Kit. (Item 53) 53. The method according to any one of Items 50 to 52, wherein the inflammatory disease or inflammatory disorder or fibrotic disease is selected from the group consisting of urticaria, atopic dermatitis, non-alcoholic steatohepatitis (NASH), primary sclerosing cholangitis, pulmonary fibrosis, chronic obstructive pulmonary disease (COPD), acute respiratory distress syndrome (ARDS), cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, asthma, bleomycin lung, scleroderma, liver cirrhosis, endomyocardial fibrosis, fibromyalgia, and eosinophilic esophagitis. (Item 54) 54. The method of claim 53, wherein the pulmonary fibrosis is idiopathic pulmonary fibrosis or scleroderma pulmonary fibrosis. (Item 55) 1. An antibody or fragment thereof that specifically binds to SCF248, for use as a pharmaceutical, wherein the antibody or fragment thereof binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction between SCF248 and c-Kit. (Item 56) 1. An antibody or fragment thereof that specifically binds to SCF248, wherein the antibody binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and wherein the antibody inhibits the interaction of SCF248 with c-Kit, for use in a method of treatment, the method comprising inhibiting activation of immune cells in a subject, inhibiting inflammation in a subject, inhibiting fibrosis in a subject, treating an inflammatory disease or disorder, treating a chronic inflammatory disease or disorder, and / or treating a fibrotic disease. (Item 57) 46. ​​The antibody or fragment thereof according to any one of items 1 to 21, 34, or 42 to 45, or the pharmaceutical composition according to item 22, for use as a medicament. (Item 58) 46. ​​The antibody or fragment thereof according to any one of items 1 to 21, 34, or 42 to 45, or the pharmaceutical composition according to item 22, for use in a method for inhibiting chronic inflammation, inhibiting fibrosis, treating a chronic inflammatory disease, and / or treating a fibrotic disease in a subject. (Item 59) 59. The antibody or fragment thereof for use according to item 56 or 58, wherein the chronic inflammatory disease or the fibrotic disease is selected from the group consisting of urticaria, atopic dermatitis, non-alcoholic steatohepatitis (NASH), primary sclerosing cholangitis, pulmonary fibrosis, chronic obstructive pulmonary disease (COPD), acute respiratory distress syndrome (ARDS), cystic fibrosis, peribronchial fibrosis, hypersensitivity pneumonitis, asthma, bleomycin lung, scleroderma, liver cirrhosis, endomyocardial fibrosis, fibromyalgia, and eosinophilic esophagitis. (Item 60) 60. The antibody or fragment thereof for use according to item 59, wherein the pulmonary fibrosis is idiopathic pulmonary fibrosis or scleroderma pulmonary fibrosis. (Item 61) 61. The antibody or fragment thereof for use according to any one of items 55 to 60, wherein the use further comprises administering an additional therapeutic agent to the subject.

Claims

1. Use of an antibody for the manufacture of a medicament for treating a chronic inflammatory disease or a fibrotic disease in a subject in need thereof, comprising: the antibody specifically binds to stem cell factor isoform 248 (SCF248); the antibody comprises a heavy chain CDR1, CDR2, and CDR3; the amino acid sequences of the heavy chain CDR1, CDR2, and CDR3 consist of SEQ ID NOs: 1, 2, and 3, respectively; the antibody comprises a light chain CDR1, CDR2, and CDR3; Use of an antibody wherein the amino acid sequences of the light chain CDR1, CDR2, and CDR3 consist of SEQ ID NOs: 4, 5, and 6, respectively.

2. The antibody (a) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 8; and (b) a light chain variable region having the amino acid sequence of SEQ ID NO: 16 2. Use of the antibody of claim 1, comprising:

3. Use of the antibody described in claim 1, wherein the antibody is humanized.

4. Use of the antibody described in claim 1, wherein the antibody is a monoclonal antibody.

5. Use of the antibody described in claim 4, wherein the antibody comprises a human IgG4 domain.

6. Use of the antibody described in claim 5, wherein the IgG4 domain comprises a S241P mutation at amino acid residue 241 and a L248E mutation at amino acid residue 248, and the numbering of the residues is that of the Kabat numbering system.

7. Use of the antibody described in claim 4, wherein the antibody comprises a heavy chain of sequence number 42 and a light chain of sequence number 49. (a) the antibody blocks the interaction between SCF and c-Kit. (b) the antibody causes internalization of SCF, and / or (c) the antibody does not bind to SCF220; Use of the antibody of claim 1.

9. The use of the antibody described in claim 1, wherein the antibody binds to an epitope comprising at least 8 consecutive amino acids of SEQ ID NO: 33, and the antibody inhibits the interaction between SCF248 and c-Kit.

10. Use of the antibody described in claim 9, wherein the epitope consists of sequence number 33.

11. Use of the antibody described in claim 5, wherein the human IgG4 domain comprises the amino acid sequence of SEQ ID NO:

40.

12. Use of the antibody described in claim 1, wherein the antibody comprises a light chain human Ig kappa constant domain.

13. Use of the antibody described in claim 12, wherein the light chain human Ig kappa constant domain comprises the amino acid sequence of SEQ ID NO:

41.

14. Use of the antibody described in claim 1, wherein the antibody comprises a light chain human Ig lambda constant domain.

15. Use of an anti-stem cell factor 248 (SCF248) antibody for the manufacture of a medicament for treating a chronic inflammatory or fibrotic disease in a subject in need thereof, comprising: The antibody specifically binds to SCF248, (i) a heavy chain having the amino acid sequence of SEQ ID NO: 42; and (ii) a light chain having the amino acid sequence of SEQ ID NO: 49 Use of an anti-SCF248 antibody, which is an antibody comprising:

16. An isolated nucleic acid molecule encoding an antibody, comprising: The antibody heavy chain CDR1, CDR2, and CDR3 comprising SEQ ID NOs: 1, 2, and 3, respectively; and Light chain CDR1, CDR2, and CDR3 comprising SEQ ID NOs: 4, 5, and 6, respectively An isolated nucleic acid molecule comprising:

17. The antibody comprising: (a) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 8, and (b) a light chain variable region having the amino acid sequence of SEQ ID NO: 16 17. The isolated nucleic acid molecule of claim 16, comprising:

18. An expression vector comprising a nucleic acid segment encoding an antibody, The antibody heavy chain CDR1, CDR2, and CDR3 comprising SEQ ID NOs: 1, 2, and 3, respectively; and Light chain CDR1, CDR2, and CDR3 comprising SEQ ID NOs: 4, 5, and 6, respectively An expression vector comprising:

19. The antibody comprising: (a) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 8, and (b) a light chain variable region having the amino acid sequence of SEQ ID NO: 16 19. The expression vector of claim 18, comprising:

20. A recombinant host cell comprising the expression vector described in claim 18.

21. The antibody, (a) a heavy chain variable region having the amino acid sequence of SEQ ID NO: 8, and (b) a light chain variable region having the amino acid sequence of SEQ ID NO: 16 21. The recombinant host cell of claim 20, comprising:

22. A composition for treating a chronic inflammatory or fibrotic disease in a subject in need thereof, comprising an antibody, the antibody specifically binds to stem cell factor isoform 248 (SCF248); the antibody comprises a heavy chain CDR1, CDR2, and CDR3; the amino acid sequences of the heavy chain CDR1, CDR2, and CDR3 consist of SEQ ID NOs: 1, 2, and 3, respectively; the antibody comprises a light chain CDR1, CDR2, and CDR3; The amino acid sequences of the light chain CDR1, CDR2, and CDR3 consist of SEQ ID NOs: 4, 5, and 6, respectively.

23. A composition for treating a chronic inflammatory or fibrotic disease in a subject in need thereof, comprising an anti-stem cell factor 248 (SCF248) antibody, The antibody specifically binds to SCF248, (i) a heavy chain having the amino acid sequence of SEQ ID NO: 42; and (ii) a light chain having the amino acid sequence of SEQ ID NO: 49 The composition is an antibody comprising:

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