WO2016191289A2 - Biomarker for inflammatory disorders and uses thereof - Google Patents

Abstract

Disclosed herein are methods of screening, diagnosing, treating, evaluating efficacy, and/or monitoring inflammatory disorders such as rheumatoid arthritis and treatments for those inflammatory disorders by inhibiting or antagonizing IL-1 β and/or IL-17. The methods may involve measuring one or more biomarkers selected from collagen, aggrecan, ΤΊΜΡ-1, TRAP-5b, GM-CSF, CD4, CD40, MIP-1α, TNF, CD11b, IL-6, IL-10, IL-15, and IL-13Rα2.

Description

BIOMARKER FOR INFLAMMATORY DISORDERS AND USES THEREOF

[0001] This application claims priority to U.S. Provisional Application

No. 62/165,609, filed May 22, 2015, which is expressly incorporated herein by reference in its entirety for any purpose.

[0002] The present disclosure relates to methods of screening, diagnosing, treating, evaluating, and/or monitoring inflammatory disorders such as rheumatoid arthritis.

[0003] Inflammation plays a role in the pathology of many diseases.

Inflammatory disorders are complex pathological conditions driven by various mediators and networks. A component behind these pathological conditions is the cytokine network. Studies have shown that cytokine-neutralizing agents can be effective at treating inflammatory disorders. One of the most common human inflammatory disorders is rheumatoid arthritis (RA), characterized by a chronic inflammatory reaction in the synovium of joints. A complex interaction of proinflammatory cytokines and chemokines expressed in disease joints are believed to play a pivotal role in inducing inflammation in joints of RA patients. Several cytokine-neutralizing agents are approved for treatment of RA, including antibodies targeting tumor necrosis factor a (TNFa), interleukin-1 (IL- 1), or interleukin-6 (IL-6). [Genovese M., et al. Ann Rheum Dis 2013;72:863- 869].

[0004] Diseases with an inflammatory component are often treated with general or specific immunosuppressants, biologic agents, antibiotics and/or anti-inflammatory pharmaceuticals. However, a large degree of inter-patient variability has been observed in patient response to treatment, e.g., with inhibitors of TNF-a, IL-1 , IL-6, and/or IL-17. Thus, multiple pathways may be involved in inflammation in a given patient, and identifying suitable biomarkers may help determine a patient's suitability for a particular treatment. However, given the complexity of the pathways involved in inflammation, the art has not yet provided suitable biomarkers to identify patients susceptible to a particular treatment (such as inhibition or antagonization of 1L-1 β and IL-17).

[0005] One potential method of treatment for inflammatory disorders is to disrupt/neutralize two or more inflammatory targets (e.g., targets such as TNF, IL-1 , IL-6, and IL17) simultaneously. For instance, bispecific antibodies or other multi-specific constructs could be used for simultaneous targeting. US Patent No. 7,612,181 (incorporated herein by reference in its entirety) provides a novel family of dual variable domain (DVD-lg) binding proteins capable of binding two or more antigens with high affinity. DVD-lg molecules are unique binding proteins comprised of two variable domains fused to N-terminal constant regions. The variable domains may be directly fused to one another or connected via synthetic peptide linkers of assorted length and amino acid composition. DVD-lg binding proteins may be engineered with intact and functional Fc domains, allowing them to mediate appropriate effector functions. U.S. Patent Publication 2014/0271458 and International Application WO

2014/144280 (incorporated herein by reference in their entirety) provide exemplary DVD-lg binding proteins capable of targeting IL-1 β and IL-17 simultaneously.

[0006] Despite the multiple treatment options, some patients still fail to respond to anti-cytokine therapies or lose response effectiveness over time. Given the large degree of inter-patient variability in the response to the monotherapies, combination therapies or multifunctional constructs of different cytokine-neutralizing reagents hold potential promise to reach a broader patient population. Additional work is needed, however, to correctly target these mono- and multi-specific therapies to the patients most likely to respond to those treatments. In addition, inflammatory disorders often have a slow progression and it is difficult to assess the disease modifying properties of therapies that slow down or halt the progression of tissue structural changes. Accordingly, there is a need for solutions that provide for improved methods of treatment, screening, diagnosing, and/or monitoring of inflammatory disorders during the course of treatment.

[0007] As disclosed herein, biomarkers such as collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, CD4, CD40, IL-15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL-10, and IL-13Ra2 can be used in the detection, diagnosis, treatment, and monitoring of patients with inflammatory disorders. Such markers may be detected at altered concentrations in biological samples from patients suffering from and/or being treated for inflammatory disorders. The altered expression levels can also be used diagnostically (e.g., altered expression levels of one or more biomarkers can serve as a diagnostic or screening marker to identify a symptomatic or pre-symptomatic subject suffering from an inflammatory disorder). The altered expression levels can also be used to monitor the efficacy of a course of treatment for an inflammatory disorder (e.g., if expression levels of a biomarker do not decrease over the course of treatment, this can indicate an ineffective treatment). Accordingly, disclosed herein are new methods of diagnosing, monitoring the progression of treatment, and/or adjusting the dose of a therapeutic agent for treating an inflammatory disorder by determining the expression levels of one or more biomarkers, such as one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP- 5b, CD4, CD40, IL-15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL-10, and IL- 13Ra2.

[0008] In certain aspects of the present disclosure, a method of treatment of a subject having an inflammatory disorder is provided, comprising determining the level of one or more biomarkers in a biological sample from the subject, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11b, IL-6, IL-10, IL-15, and IL-13Ra2; and administering one or more therapeutic agents that antagonize IL-1 β and IL-17 if the subject has a level of the biomarker that is greater than a reference level. In some embodiments, a DVD-lg, such as one of those described above targeting IL-1 β and IL17, may be used as a therapeutic agent in a method of treatment after measuring and determining the level of one or more biomarkers in a biological sample from a subject, followed by administering the DVD-lg if the level of the biomarker is greater than a reference level. In various embodiments, the measured biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11b, IL-6, IL-10, IL-15, and IL-13Ra2.

[0009] The level of expression of a biomarker in a sample obtained from a subject may be assayed by any of a wide variety of techniques and methods. In some embodiments, measuring and determining the biomarker level in a sample can comprise a technique from one or more of the following: fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high-throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof. In an embodiment, the method is Northern blot, ELISA, FACS, or flow cytometry.

[0010] The level of one or more biomarkers in a biological sample may be compared to a reference level from a reference sample in any of the above-mentioned techniques. The reference level may be the level in a reference sample obtained from a healthy subject, or a sample from a patient with the inflammatory disorder prior to or during treatment. In certain aspects, the biological sample may be a cell, tissue, fluid, organ, blood vessel, bone, cartilage, synovial membrane, synovial fluid, or another fluid from space within a joint cavity. In other aspects, the biological sample is whole blood or serum. In some embodiments, the reference level is an average of levels in more than one reference sample. In certain embodiments, the reference level is an average of the level in 2, 3, 4, 5, 6, 7, 8, 9, 10, or more reference samples.

[0011] In an embodiment, a combination therapy comprising therapeutic agents that antagonizes IL-1 β and IL-17 decreases the level of expression of at least one of collagen II, aggrecan, TIMP-1 , TRAP-5b, CD4, CD40, IL-15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL-10, and IL-13Ra2 biomarkers to a greater extent than a monotherapy comprising a therapeutic agent that antagonizes IL-1 β or a therapeutic agent that antagonizes IL-17.

[0012] In an embodiment, the subject may have been previously administered one or more therapeutic agents that antagonize IL-1 β and IL-17, and/or an alternative therapy. In other embodiments, the subject may be under consideration for treatment with one or more therapeutic agents that antagonize IL-1 β and IL-17 for the first time.

[0013] In various embodiments of the present disclosure, a method of treatment of a human subject having an inflammatory disorder is provided, comprising administering one or more therapeutic agents that antagonize IL-1 β and IL-17, determining the level of one or more biomarkers in a biological sample from a subject, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11b, IL-6, IL-10, IL-15, and IL-13Fta2, and administering a further dose of the one or more therapeutic agents if the levels of the one or more biomarkers in the biological sample are greater than the reference level. In certain aspects, a DVD-lg, such as one of those described above targeting IL-1 β and IL17, may be used.

[0014] In certain aspects, a method of monitoring treatment effectiveness is provided, comprising administering one or more therapeutic agents that antagonize IL-1 β and IL-17, determining the level of one or more biomarkers in a biological sample from a subject, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11b, IL-6, IL-10, IL- 15, and IL-13Ra2, and determining effectiveness if the markers are not elevated relative to a reference sample from a healthy subject. Treatment may be considered effective when the measured biomarker level is equal to or less than the reference level. In some embodiments, the treatment comprises

administering a DVD-lg, such as one of those described above targeting IL-1 β and IL17. In some embodiments, the methods are used to monitor disease progression for patients with inflammatory disorders. In some embodiments, inflammatory disease progression or pharmacologic responses to an

intervention are monitored using the biomarkers. The biomarkers can also be used to monitor surrogate activities such as target modulation, disease progression, and treatment efficacy marker.

[0015] Also disclosed herein, in certain embodiments, are methods of screening populations for subjects with inflammatory disorders susceptible to treatment by antagonizing or inhibiting IL-1 β and IL17 (e.g., using one of the DVD-lg binding proteins discussed above), diagnosing patients with

inflammatory disorders who may be suitable for treatment by antagonizing or inhibiting IL-1 β and IL17, monitoring a patient during treatment with antagonists or inhibitors of IL-1 β and IL17 for treatment effectiveness, and a method of calibrating an effective dose of antagonists or inhibitors of IL-1 β and IL17. In some embodiments, the methods comprise determining the level of one or more biomarkers in a biological sample from a subject, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, ΜΙΡ-Ια, TNF (preferably TN Fa), CD11b, IL-6, IL-10, IL- 15, and IL-13Ra2, and comparing to a level in a reference sample, wherein an elevated level indicates subjects suitable for treatment by antagonizing or inhibiting IL-Ιβ and IL17. Such methods may help reduce the cost of treatment by targeting IL-1 β and IL17 therapy to those more likely to respond to treatment.

[0016] In some embodiments, disclosed herein are methods of screening, diagnosing, evaluating, monitoring and/or treating rheumatoid arthritis (RA) using the biomarkers discussed above. In some embodiments, the methods and biomarkers relate to treatment of RA by disrupting IL-1 β and IL-17, e.g., using one of the DVD-lg binding proteins discussed above.

[0017] The methods disclosed herein also include methods of evaluating the effectiveness of a treatment for an inflammatory disorder. In certain aspects, the methods comprise administering to the subject one or more therapeutic agents that antagonize IL-1 β and IL-17, collecting a biological sample from the subject after the administration of the therapeutic agents, measuring the level of one or more biomarkers in the sample, and comparing the levels of the biomarkers in the sample to reference levels. In some embodiments, the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF

(preferably TNFa), CD11b, IL-6, IL-10, IL-15, and IL-13Ra2, and a level equal to or less than the reference level indicates an effective treatment.

[0018] The methods disclosed herein also include methods of screening a subject or population of subjects to identify a subject suitable for treatment of an inflammatory disorder by administering one or more therapeutic agents that antagonize IL-1 β and IL-17. In certain aspects, the method comprises collecting a biological sample from the subject, measuring the level of one or more biomarkers in the sample, comparing the levels of the

biomarkers in the sample to reference levels; the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1, TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11b, IL-6, IL-10, IL-15, and IL- 13Ra2, and identifying a subject suitable for treatment with one or more therapeutic agents that antagonize IL-1 β and IL-17 if the levels of the one or more biomarkers are elevated as compared to the reference level. Such a method of screening could potentially reduce testing costs.

[0019] In some embodiments, the methods disclosed herein can also be used to select and monitor a subject for participation in a clinical trial using one or more therapeutic agents that antagonize IL-1 β and IL-17. For instance, if expression levels of a biomarker are elevated in the subject, they may be suitable for inclusion in a study. Also, if levels decrease after experimental treatment, this may indicate that the effectiveness of the treatment in the subject.

BRIEF DESCRIPTION OF THE DRAWINGS

[0020] FIG. 1 A and Fig. 1 B are schematic representations of a Dual

Variable Domain (DVD) binding protein construct.

[0021] FIG. 2 is a schematic representation of the rheumatoid arthritis

(RA) QSP Platform, showing the inflammatory network dynamics and impact of the network modulation on bone and cartilage turnover in a joint with

established chronic inflammation and rheumatoid arthritis disease.

[0022] FIG. 3 is a bar graph comparing different clinically approved rheumatoid arthritis therapies with respect to the mean improvement in ACR score extracted from clinical trials to the QSP predicted ACR-N score of the reference virtual patient. Tested treatments include Humira (anti-TNF), Tocilizumab (anti-IL-6R), Anakinra (anti-IL-1R), Secukinumab (anti-IL17A), and Abatacept (CTLA-4-lgG1 fusion).

[0023] FIG. 4 is a bar graph showing histopathology scores in DBA-1

CIA mice (vertical axis) after administration of PBS, anti-TNFa, anti-IL17, anti- 1L-1 p, and the combination of anti-IL-Ιβ and anti-IL-17. Measurements were taken from inflammation, cartilage, and bone. Anti-TNFa was the positive control.

[0024] FIG. 5 is a bar graph from QSP analysis showing the percent change for different proteins (x-axis) as a result of treatment with Secukinumab, Anakinra, or a combination of Secukinumab and Anakinra.

[0025] FIG. 6 is a plot showing the gene expression profiles generated from mouse paw gene array data of Aggrecan, TRAP-5b, and TIMP- 1 in naive mice or DBA-1 CIA mice treated either with vehicle, anti-IL17, anti- f L1 β, or an anti-IL17/anti-IL1 β combination ("combo"). The experiment had a positive control of anti-TNFa.

[0026] FIG. 7 is a schematic representation of the mediators that play a role in regulating chondrocyte production of the key biomarkers modulated by the anti-ILi p/anti-IL17 DVD-lg therapy, and the postulated effect of this DVD-lg therapy on cartilage health.

DESCRIPTION OF CERTAIN EXEMPLARY EMBODIMENTS

[0027] Reference will now be made in detail to certain exemplary embodiments according to the present disclosure, certain examples of which are illustrated in the accompanying drawings. Wherever possible, the same reference numbers will be used throughout the drawings to refer to the same or like parts.

[0028] In this application, the use of the singular includes the plural unless specifically stated otherwise. In this application, the use of "or" means "and/or" unless stated otherwise. Furthermore, the use of the term "including," as well as other forms, such as "includes" and "included," is not limiting. Any range described herein will be understood to include the endpoints and all values between the endpoints.

[0029] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described. All documents, or portions of documents, cited in this application, including but not limited to patents, patent applications, articles, books, and treatises, are hereby expressly incorporated by reference in their entirety for any purpose. To the extent anything in those references contradicts anything disclosed herein, the statements disclosed herein will control.

[0030] The term "therapeutic agents that antagonizes IL-1 β" includes agents that have the effect of binding to or neutralizing, inhibiting, reducing, or negatively modulating the activity of lnterleukin-1 beta (IL-1 β). The term "anti-IL- 1 β" is used interchangeably with "therapeutic agents that antagonizes IL-Ι β." In an embodiment, the therapeutic agent that antagonizes IL-1 β comprises an anti- IL-1 β binding protein. In another example, the anti-IL-1 β binding protein comprises a fusion protein, e.g., Rilonacept, or an IL-1 β-binding fragment thereof. In an embodiment, the therapeutic agents that antagonizes IL-1 β can comprise an anti- IL-1 β antibody, or an antigen binding fragment thereof. In an embodiment, an antibody is a murine antibody, a human antibody, a humanized antibody, a bispecific antibody, a chimeric antibody, a Fab, a Fab', a F(ab')₂, an ScFv, an SMIP, an affibody, an avimer, a versabody, a nanobody, a domain antibody, and an antigen binding fragment of any of the foregoing.

[0031] In an embodiment, the anti- IL-1 β antibody comprises a human anti- IL-1 β antibody, e.g., a human anti- IL-1 β antibody, e.g., Anakinra,

Canakinumab, or an antigen binding fragment thereof. In another embodiment, the anti- IL-1 β antibody comprises a humanized anti- IL-1 β antibody, e.g., Gevokizumab, or an antigen binding fragment thereof.

[0032] In an embodiment, the therapeutic agents that antagonizes IL-

1 β comprises a multispecific binding protein. In an embodiment, the

multispecific binding protein comprises a dual variable domain immunoglobulin (DVD-lg™) molecule, a half-body DVD-lg (hDVD-lg) molecule, a triple variable domain immunoglobulin (tDVD-lg) molecule, a receptor variable domain immunoglobulin (rDVD-lg) molecule, a polyvalent DVD-lg (pDVD-lg) molecule, a monobody DVD-lg (mDVD-lg) molecule, a cross over (coDVD-lg) molecule, a blood brain barrier (bbbDVD-lg) molecule, a cleavable linker DVD-lg (cIDVD-lg) molecule, or a redirected cytotoxicity DVD-lg (rcDVD-lg) molecule.

[0033] The term "therapeutic agents that antagonizes IL-17" includes agents that have the effect of binding to or neutralizing, inhibiting, reducing, or negatively modulating the activity of interleukin 17 (IL-17). The term "anti-IL-17" is used interchangeably with "therapeutic agents that antagonizes IL-17". In an embodiment, the therapeutic agent that antagonizes IL-17 comprises an anti-IL- 17 binding protein. In another example, the anti-IL-17 binding protein comprises a fusion protein. In an embodiment, the therapeutic agents that antagonizes IL- 17 comprises an anti-IL-17 antibody, or an antigen binding fragment thereof. In an embodiment, the anti-IL-17 antibody comprises a human antibody, e.g., Secukinumab and RG7624, or an antigen binding fragment thereof. In an embodiment, the anti-IL17 antibody comprises a humanized antibody, for example ixekizumab, 10F7, B6-17, or an antigen binding fragment thereof. In other embodiments, the therapeutic agents that antagonizes IL-17 comprises methotrexate, an analog thereof, or a pharmaceutically acceptable salt thereof. In an embodiment, the anti-IL-17 can include a multispecific binding protein, such as the dual variable domain immunoglobulin molecules described above for IL-Ι β, and as described in more detail below.

[0034] The phrase "inflammatory disorder" refers to a disease or disorder characterized by chronic or acute inflammation. Numerous

inflammatory disorders are known in the art, such as arthritis, including rheumatoid arthritis, osteoarthritis, psoriatic arthritis, juvenile idiopathic arthritis; Alzheimer's disease; ankylosing spondylitis; asthma; autoimmune diseases; angina; acne vulgaris; appendicitis; atherosclerosis; Behcet's disease; bursitis; blepharitis; Crohn's disease; colitis; cystitis; chronic obstructive pulmonary disease (COPD); cystic fibrosis; celiac disease; chronic prostatitis; dermatitis; diverticulitis; emphysema; ectopic eczema; fibromyalgia; gastroenteritis;

glomerulonephritis; hay fever; hepatitis; inflammatory bowel diseases (IBD); insulin dependent diabetes mellitus; intestinal flupelvic inflammatory disease (PID); interstitial cystitis; keratoconjunctivitis sicca; keratitis; multiple sclerosis; nephritis; necrotizing enterocolitis (NEC); periodontitis; pleurisy; pyelitis;

pharyngitis; Parkinson's disease; psoriasis; pelvic inflammatory disease; plaque psoriasis; phlebitishypersensitivities; rubor; rhinitis; reperfusion injury; sore throat; sarcoidosis; spondylarthritis; systemic lupus erythematosus (SLE); stomach flu; transplant rejection; tendonitis; tonsillitis; uveitis; urticarial; urinary tract infection; ulcerative colitis; vasculitis; and some cancers (e.g., gallbladder carcinoma).

[0035] The terms "marker" and "biomarker" are used interchangeably herein to mean a substance that is used as an indicator of a biologic state, e.g., genes, messenger RNAs (mRNAs, microRNAs (miRNAs)); heterogeneous nuclear RNAs (hnRNAs), and proteins, or portions thereof.

[0036] Reference to a gene encompasses naturally occurring or endogenous versions of the gene, including wild type, polymorphic or allelic variants or mutants (e.g., germline mutation, somatic mutation) of the gene, which can be found in a subject. In an embodiment, the sequence of

the biomarker gene is at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% identical to a wild type sequence. Sequence identity can be determined, e.g., by comparing sequences using NCBI BLAST (e.g., Megablast with default parameters).

[0037] The "level of expression" or "expression level" refers to a quantitative or qualitative indication of the expression of one or more markers or biomarkers in a subject, such as in comparison to a standard, a reference value, or a control.

[0038] As used herein, the terms "subject" and "patient" refers to human, and to non-human animals, e.g., veterinary patients. The term "non- human animal" includes vertebrates, e.g., mammals, such as non-human primates, mice, rodents, rabbits, sheep, dogs, cats, horses, cows, ovine, canine, feline, equine or bovine species. In an embodiment, the subject is a human (e.g., a human with an inflammatory disease, e.g., RA).

[0039] The term "biological sample" refers to a quantity of a

substance from a living thing or formerly living thing. Such substances include, but are not limited to, blood, (e.g., whole blood, dried blood spot), tissue, plasma, serum, urine, cell, bone, fluid, organ, or blood vessel. In one

embodiment, the cell is an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a plasma cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a chondrocyte, a Kupffer cell, or a mastocyte. In one embodiment, the tissue or cell is removed from the subject. In another embodiment, the tissue or cell is present within the subject. In an embodiment, the fluid comprises amniotic fluid, aqueous humor, vitreous humor, bile, blood, breast milk, cerebrospinal fluid, cerumen, chyle, cystic fluid, endolymph, feces, gastric acid, gastric juice, lymph, mucus, nipple aspirates, pericardial fluid, perilymph, peritoneal fluid, plasma, pleural fluid, pus, saliva, sebum, semen, sweat, serum, sputum, synovial fluid, tears, urine, vaginal secretions, fluid collected from a biopsy, or other fluid from space within a joint cavity. In one embodiment, the sample contains protein (e.g., proteins or peptides) from the subject. In another embodiment, the sample contains RNA (e.g., mRNA) from the subject or DNA (e.g., genomic DNA molecules) from the subject.

[0040] As used herein, the term "therapeutically effective amount" means an amount of a treatment, e.g., an anti- IL-Ι β treatment and/or an anti- IL-17 treatment, which is capable of treating, e.g., reducing a symptom, ameliorating, lessoning, etc., one or more phenotypic aspect of an inflammatory disease (e.g., RA). The dose of a therapy to be administered may also be determined in light of the particular circumstances of the patient to be treated, including, for example, the therapy administered, the route of administration, condition and particular features of the patient, and the pathological condition being treated, for example, the severity of the RA in the subject. [0041] For administration to a subject, a combination therapy may be formulated into a pharmaceutical composition comprising one or more therapeutic agents that antagonize IL-1 β and IL-17 and a pharmaceutically acceptable carrier. Therapeutic compositions typically may be sterile and adequately stable under the conditions of manufacture and storage.

[0042] As used herein, a "pharmaceutically acceptable carrier" includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible. Preferably, the carrier is suitable for parenteral (e.g., intravenous, intramuscular, subcutaneous, intrathecal) administration (e.g., by injection or infusion). Depending on the route of administration, the active compound may be coated in a material to protect the compound from the action of acids and other natural conditions that may inactivate the compound.

Methods of screening, diagnosing, evaluating, monitoring and/or Treating

[0043] Disclosed herein are methods of screening, diagnosing, evaluating, monitoring and/or treating inflammatory disorders, such as rheumatoid arthritis. An "inflammatory disorder" can encompass any disorder associated with the development of dysregulated or persistent inflammation. Many inflammatory disorders are known, including inflammatory bowel disease (IBD), Crohn's disease, ulcerative colitis, arthritis, juvenile idiopathic arthritis, osteoarthritis, systemic lupus erythematosus (SLE), keratoconjunctivitis sicca, blepharitis, keratitis, uveitis, insulin dependent diabetes mellitus,

spondyloarthropathy, chronic obstructive pulmonary disease (COPD), cystic fibrosis, urticaria, eczema, ectopic eczema psoriasis, plaque psoriasis, psoriatic arthritis, multiple sclerosis, ankylosing spondylitis, asthma, and rheumatoid arthritis.

[0044] The methods disclosed herein can be used, in some

embodiments, to monitor the long-term efficacy of a course of treatment for an inflammatory disorder. For example, if expression levels of a biomarker do not decrease over the course of treatment, this can indicate an ineffective treatment or a need to increase the dose of treatment being administered. In contrast, if levels decrease relative to levels prior to treatment, this can be used to indicate an effective treatment, and subsequent treatment can be given at the same dosage, or a reduced dosage, or treatment can be discontinued.

[0045] In some embodiments, the therapeutic agent is not

administered, or a further dose is not administered, if the biomarker level is less than or equal to the reference level. In some embodiments, a reduced concentration dose is administered if the biomarker level is less than or equal to the reference level.

[0046] In some embodiments, the methods disclosed herein can comprise treating a human subject having an inflammatory disorder. In certain embodiments, the methods can comprise determining the level of one or more biomarkers in a biological sample from a subject. In some embodiments, the biomarkers include one of the following: collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TIM Fa), CD11b, IL-6, IL-10, IL-15, and IL-13Ra2. The marker may be a gene or gene product, such as RNA or protein. In some embodiments, biomarker levels are determined before treatment. In some embodiments, biomarker levels are determined after administering a first dose of treatment, and/or after administering a subsequent dose. In some embodiments, biomarker levels are determined before and after administration.

[0047] In some embodiments, treatment comprises administering one or more agents to antagonize or inhibit IL-1 β and IL-17 (simultaneously or sequentially) if the levels of biomarkers measured in a patient sample are elevated relative to a reference sample, and/or continuing to administer treatment if the markers are elevated after a first dose of treatment. Where sequential administration is used, the separation in time between administering the agents is selected such that both the therapeutic agent that antagonizes IL- 1β and the therapeutic agent that antagonizes IL-17 act concomitantly and/or achieve a synergistic effect. In some embodiments, an elevated level of a biomarker in the biological sample is a level greater than a reference level by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 350%, 400%, 450%, 500%, or more (or any value in between). In some embodiments, the biomarker comprises at least one of: aggrecan, TIMP-1 , and TRAP-5b.

[0048] In some embodiments, a lower level of expression of at least one of the biomarkers after the subject is treated with one or more therapeutic agents that antagonize IL-1 β and IL-17, as compared to a reference level (e.g., the level of expression of the biomarkers in a sample from the patient before the treatment, or an arbitrary value), indicates that the treatment has been effective in treating the subject. In some embodiments, treatment is administered, or dosage is increased, until a lower level of expression of one or more biomarkers is achieved. In some embodiments, a lower level of expression" means that the expression level of a biomarker in the biological sample is lower than the reference level by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 350%, 400%, 450%, 500%, or more (or any value in between). In some embodimens, the "lower level of expression" means that the expression level of a biomarker in the biological sample is lower than the reference level by at least about 25%, 50%, or 75% (or any value in between). In some embodiments, the biomarker comprises one or more of: TNF, IL-10, GM-CSF, MIP1a, IL-6, CD11b, and IL-13Ra2. In some embodiments, the biomarker comprises one or more of: TNF, IL-10, GM-CSF, MIP1a, and IL-6. In some embodiments, the biomarker comprises at least one of: CD11b and IL-13Ra2. In some embodiments, the biomarker comprises at least one of: aggrecan, TIMP-1, and TRAP-5b.

[0049] In some embodiments, antagonizing or inhibiting IL-1 β and IL-

17 (simultaneously or sequentially) includes, but is not limited to, a partial reduction or complete elimination of measured protein jn a sample, a partial reduction or complete elimination of gene expression measured in a sample, and/or a partial reduction or complete elimination in the measured activity of a protein or gene in a sample (e.g., ability to activate a signaling pathway, induce inflammation, or other known functions of IL-Ιβ and/or IL-17). "Partial" encompasses a reduction of at least about 5%, such as at least about 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 85, 97, 98, 99, or 99.5%. "Partial" protein or gene expression may refer to a change in the activity of the protein or gene (such as transcription of an RNA, mRNA, rRNA, tRNA from the target gene and/or translation of the protein product encoded by it). [0050] In some embodiments, the partial reduction is measured by assessing IL-1 β and/or IL-17 activity In a functional assay. In some

embodiments, the partial reduction of the protein or gene may be determined using, for example, an immunological detection assay. Moreover, in some embodiments, biochemical techniques such as of fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high-throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked

immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof may be used to analyze the partial reduction in protein quantity or gene expression.

[0051] In some embodiments, the method of treating a human subject having an inflammatory disorder comprises maintaining, decreasing dosage, or discontinuing administration of the therapeutic agent(s) if the level of the biomarker in the subject is determined to be less than or equal to the reference level. In some embodiments, the method of treating a human subject having an inflammatory disorder comprises maintaining or increasing dosage of the therapeutic agent(s) if the level of the biomarker in the subject is determined to be greater than the reference level. In certain embodiments, the dosage is calibrated to reduce the level of measured biomarkers to the level in a reference sample, but not lower. In other embodiments, the dosage is calibrated to reduce the level of measured biomarkers below that in a reference sample. In some embodiments, treatment is discontinued when he level of measured biomarkers reaches or goes below the level in the reference sample.

[0052] In some embodiments, the therapeutic agent(s) are

administered intravenously, intradermally, subcutaneously, or intramuscularly. In some embodiments, a DVD-lg binding protein capable of binding IL-1 β and IL-17 is administered intravenously, intradermally, subcutaneously, or intramuscularly.

[0053] In some embodiments, each successive administration of the therapeutic agent(s) is administered one, two, three, four, five, six, or seven days, or one, two, three, or four weeks after the previous dose. In certain embodiments, each administered dose is repeated at least twice. In some embodiments, these steps are repeated until the level of the biomarker collected from the biological sample is equal to or less than the reference level.

[0054] In certain embodiments, a biomarker is considered elevated in a biological sample when comparing to a reference level. In certain

embodiments, an elevated biomarker is at a protein or nucleic acid level that is least 5% greater than the reference level.

[0055] In some embodiments, the administering of one or more therapeutic agents to a subject having an inflammatory disorder comprises simultaneously or sequentially administering an anti-IL-1 β antagonist and an anti-IL-17 antagonist. In other embodiments, the anti-IL-1 β antagonist is an anti-IL-1 β antibody and the anti-IL17 antagonist is an anti-IL-17 antibody, e.g., antibodies having variable domains selected from those in Table 1.

[0056] In some embodiments, the treatment is a bispecific antibody capable of binding IL-1 β and IL-17, or another multispecific construct capable of binding both targets, in certain embodiments, the treatment is with bispecific antibodies that have been produced by quadroma technology (Milstein and Cuello (1983) Nature 305(5934): 537-40), by chemical conjugation of two different monoclonal antibodies (Staerz et al. (1985) Nature 314(6012): 628-31), or by knob-into-hole or similar approaches which introduces mutations in the Fc region (Holliger et al. (1993) Proc. Natl. Acad. Sci. USA 90(14): 6444-6448). In some embodiments, the multispecific binding protein is a dual variable domain immunoglobulin (DVD-lg). In some embodiments, the binding protein is a DVD- Ig disclosed in U.S. Patent Publication 2014/0271458 and International

Application WO 2014/144280 (incorporated herein by reference in their entirety). In an embodiment, the treatment is a DVD-lg targeting IL-1 β and IL- 17, such as any of the DVD-lg constructs listed in tables 1 or 2. In some embodiments, the DVD-lg binding protein is capable of binding IL-1 β at the VD1 position and IL-17 at the VD2 position. In some embodiments, the binding protein is capable of binding IL-17 at the VD1 position and IL-1 β at the VD2 position.

[0057] In some embodiments, the DVD-lg binding protein capable of binding IL-1 β and IL-17 comprises first and second polypeptide chains, each independently comprising VD1-(X1)n-VD2-C-X2, wherein VD1 is a first variable domain; VD2 is a second variable domain; C is a constant domain; X1 is a linker; X2 is an Fc region that is either present or absent; and n is 0 or 1 ;

wherein the VD1 domains on the first and second polypeptide chains form a first functional target binding site and the VD2 domains on the first and second polypeptide chains form a second functional target binding site, and wherein the binding protein is capable of binding IL-1 β and IL-17. In an embodiment, the DVD-lg binding protein comprises two first and two second polypeptide chains.

[0058] In an embodiment, the variable domains of the DVD-lg binding protein that form a functional target binding site for IL-1 β comprise paired heavy and light chain CDRs 1-3 from any of the paired IL-1 β CDR sets listed in Table 1. In an embodiment, the variable domains that form a functional target binding site for IL-1 β comprise paired heavy and light variable domain sequences from any of those listed in Table 1. In an embodiment, the variable domains that form a functional target binding site for IL-17 comprise paired heavy and light chain CDRs 1-3 from any of the paired IL-17 CDR sets listed in Table 1. In an embodiment, the variable domains that form a functional target binding site for IL-17 comprise paired heavy and light variable domain sequences from any of those listed in Table 1. In an embodiment, the variable domains that form a functional target binding site for IL-1 β comprise: CDRs 1-3 from SEQ ID NO: 1 and CDRs 1-3 from SEQ ID NO: 2, CDRs 1-3 from SEQ ID NO: 3 and CDRs 1- 3 from SEQ ID NO: 4, CDRs 1-3 from SEQ ID NO: 5 and CDRs 1-3 from SEQ ID NO: 6, CDRs 1-3 from SEQ ID NO: 7 and CDRs 1-3 from SEQ ID NO: 8, or CDRs 1-3 from SEQ ID NO: 9 and CDRs 1-3 from SEQ ID NO: 10; and the variable domains that form a functional target binding site for IL-17 comprise CDRs 1-3 from SEQ ID NO: 13and CDRs 1-3 from SEQ ID NO:14. In an embodiment, the DVD-lg binding protein comprises DVD3418 (comprising a first polypeptide chain of SEQ ID NO: 19 and a second polypeptide chain of SEQ ID NO: 20. The CDR sequences of the variable domains in Table 1 are underlined. Table 1 : List of Amino Acid Sequences of VH and VL Regions of

Antibodies for Generating Binding Proteins, Including Multivalent Binding

Proteins

[0059] Detailed descriptions of exemplary DVD-lg binding proteins capable of binding ΙΙ_-1 β and IL-17, and methods of making the same, are provided in U.S. Patent Publication 2014/0271458 and International Application WO 2014/144280 (incorporated herein by reference in their entirety).

Table 2: Exemplary DVD-lg Binding Protein That Bind IL-Ιβ and IL-17

[0060] In certain embodiments, the X1 linker on the first polypeptide chain comprises SEQ ID NO: 21 and the X1 linker on the second polypeptide chain comprises SEQ ID NO: 22. In an embodiment, the binding protein comprises a first polypeptide chain comprising SEQ ID NO: 19 and a second polypeptide chain comprising SEQ ID NO: 20. In an embodiment, the binding protein comprises a first polypeptide chain comprising an outer variable domain of SEQ ID NO:3, a linker between the outer and inner variable domains of SEQ ID NO: 21 , and an inner variable domain of SEQ ID NO: 13; and/or the binding protein comprises a second polypeptide chain comprising an outer variable domain of SEQ ID NO: 4, a linker between the outer and inner variable domains of SEQ ID NO: 22, and an inner variable domain of SEQ ID NO: 14. In an embodiment, the binding protein comprises a second polypeptide chain comprising an outer variable domain of SEQ ID NO: 3, a linker between the outer and inner variable domains of SEQ ID NO: 21, and an inner variable domain of SEQ ID NO: 13; and/or the binding protein comprises a first polypeptide chain comprising an outer variable domain of SEQ ID NO: 4, a linker between the outer and inner variable domains of SEQ ID NO: 30, and an inner variable domain of SEQ ID NO: 14. In certain embodiments, the binding protein comprises a first polypeptide chain comprising SEQ ID NO: 19 and a second polypeptide chain comprising SEQ ID NO: 20. Any of the heavy chain, light chain, two chain, or four chain embodiments can include at least one X1 linker comprising GGGGSGGGGS (SEQ ID NO: 21) or GGSGGGGSG (SEQ ID NO: 22). In an embodiment, the linker is GGGGSGGGGS (SEQ ID NO: 21) on the first chain and/or GGSGGGGSG (SEQ ID NO: 22) on the second chain. In an embodiment, the linker is GGSGGGGSG (SEQ ID NO: 22) on the first chain and/or GGGGSGGGGS (SEQ ID NO: 21) on the second chain.

[0061] In some embodiments, the DVD-lg binding protein comprises the full-length first and second polypeptide chains shown in Table 3 below. The CDR sequences in the variable domains are underlined in Table 3, linkers are shown in bold, and constant domains are in italics.

Table 3

[0062] There are numerous types of anti-inflammatory approaches that may be used in conjunction with one or more therapeutic agents that antagonize IL-1 β and IL-17. Any suitable additional anti-inflammatory treatment may be used with the methods disclosed herein. These include, for example, nonsteroidal anti-inflammatory drugs (NSAIDs), steroids, disease-modifying antirheumatic drugs (DMARDs) including methotrexate (Trexall), leflunomide (Arava), hydroxychloroquine (Plaquenil), sulfasalazine (Azulfidine) and minocycline (Dynacin, Minocin), and/or immunosuppressants including azathioprine (Imuran, Azasan), cyclosporine (Neoral, Sandimmune, Gengraf) and cyclophosphamide (Cytoxan).

[0063] In yet another aspect, disclosed herein are methods of diagnosing an inflammatory disorder in a subject. The method includes the steps of (a) collecting a biological sample from the subject; (b) measuring the level of one or more biomarkers in the sample, wherein the biomarkers are collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, CD4, CD40, IL- 15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL-10, and IL-13Ra2; (c) comparing the level of the biomarker in the sample to a reference level; and (e) diagnosing an inflammatory disorder if the level of the biomarker is greater than the reference level. In some embodiments, the expression level of a biomarker in the biological sample is greater than a reference level by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 350%, 400%, 450%, 500%, or more (or any value in between). In some embodiments, the biomarker comprises at least one of: TNF, IL-10, GM-CSF, MIP1a and IL-6. In some embodiments, the biomarker comprises at least one of: CD11b and IL-13Ra2. In some embodiments, the biomarker comprises at least one of: aggrecan, TIMP-1, and TRAP-5b.

[0064] In another aspect, the present disclosure provides a method of evaluating the effectiveness of a treatment for an inflammatory disorder. The method includes the steps of (a) administering to the subject one or more therapeutic agents that antagonize IL-1 β and IL-17; (b) collecting a biological sample from the subject after the administration of the therapeutic agents; and (c) measuring the level of one or more biomarkers in the sample, and comparing the biomarker levels in the sample to a reference level. The biomarkers may be one or more of collagen (preferably collagen II), aggrecan, TI P-1 , TRAP-5b, CD4, CD40, IL-15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL- 10, and IL-13Ra2, wherein a level equal to or less than the reference level indicates an effective treatment. In some embodiments, the expression level of a biomarker in the biological sample is less than a reference level by at least about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 350%, 400%, 450%, 500%, or more (or any value in between). In some embodiments, the biomarker comprises at least one of: TNF, IL-10, GM-CSF, MIP1a and IL-6. In some embodiments, In some embodiments, the biomarker comprises at least one of: CD11b and IL-13Ra2. In some embodiments, the biomarker comprises at least one of: aggrecan, TIMP-1 , and TRAP-5b.

[0065] In still another aspect, the present disclosure provides a method of screening a subject or population of subjects to identify a subject suitable for treatment of an inflammatory disorder by administering one or more therapeutic agents that antagonize IL-1 β and IL-17. The method may include the steps of (a) collecting a biological sample from the subject; (b) measuring the level of one or more biomarkers in the sample; (c) comparing the levels of the biomarkers in the sample to reference levels, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, CD4, CD40, IL-15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL-10, and IL-13Ra2; and (d) identifying a subject suitable for treatment with one or more therapeutic agents that antagonize IL-1 β and IL-17 if the levels of the one or more biomarkers are elevated as compared to the reference level. A higher level of expression of at least one of the biomarkers, as compared to the level of expression of the reference level, indicates that the subject is suitable for an anti- IL-1 β treatment or an anti-IL-17 treatment alone or in combination. In some embodiments, the biomarker comprises at least one of: TNF, IL-10, GM-CSF, MIP1a and IL-6. In some embodiments, In some embodiments, the biomarker comprises at least one of: CD11b and IL-13Ra2. In some embodiments, the biomarker comprises at least one of: aggrecan, TIMP-1 , and TRAP-5b.

[0066] In another aspect, the present disclosure provides a method of selecting a subject for participation in a clinical trial for treatment of an inflammatory disease using one or more therapeutic agents that antagonize IL- 1β and IL-17. The method includes the steps of (a) collecting a biological sample from the subject; (b) measuring the level of one or more biomarkers in the sample, wherein the biomarkers are collagen II, aggrecan, TIMP-1 , TRAP- 5b, CD4, CD40, IL-15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL-10, and IL- 13Ra2; and (c) comparing the level of the biomarker in the sample to a reference level. A higher level of expression of at least one of the biomarkers, as compared to the level of expression of the reference level, indicates that the subject is suitable for participation in the clinical trial. In some embodiments, the biomarker comprises at least one of: TNF, IL-10, GM-CSF, MIP1a and IL-6. In some embodiments, In some embodiments, the biomarker comprises at least one of: CD11b and IL-13Ra2. In some embodiments, the biomarker comprises at least one of: aggrecan, TIMP-1, and TRAP-5b.

[0067] In any of the above embodiments, the inflammatory disorder may be Alzheimer's disease, juvenile idiopathic arthritis, psoriasis, plaque psoriasis, keratoconjunctivitis sicca, blepharitis, keratitis, ankylosing spondylitis, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, asthma, atherosclerosis, Crohn's disease, colitis, inflammatory bowel disease (IBD), insulin dependent diabetes mellitus, chronic obstructive pulmonary disease (COPD), cystic fibrosis, urticaria, ectopic eczema, multiple sclerosis, dermatitis, diverticulitis, fibromyalgia, hepatitis, irritable bowel syndrome (IBS), systemic lupus erythematous (SLE), nephritis, Parkinson's disease, or ulcerative colitis.

Biomarkers

[0068] In certain embodiments, the biomarkers measured in any of the methods disclosed herein comprise one or a combination of the following biomarkers: collagen (preferably collagen II), aggrecan, TIMP-1, TRAP-5b, CD4, CD40, IL-15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL-10, and IL-13Ra2. In some embodiments, the biomarkers comprise one or a combination of:

Aggrecan, TIMP-1 , CD40 (preferably CD40 levels measured on macrophages), IL-15, IL-1 , IL-17, CD4 T cell adhesion molecule (preferably T cell surface expression levels), collagen (preferably collagen II), and TRAP-5b (preferably TRAP-5b levels in synovium or synovial fluid). In certain embodiments, the biomarkers measured herein comprise one or a combination of the following biomarkers: Aggrecan, TIMP-1, collagen II, and TRAP-5b.

[0069] In certain embodiments, at least one of the biomarkers is aggrecan. Aggrecan ("cartilage-specific proteoglycan core protein," "CSPCP," or "chondroitin sulfate proteoglycan Γ) is encoded by the ACAN gene.

Aggrecan is a large proteoglycan mainly found in the cartilage where it provides resilience and toughness. Aggrecan is also present in tensile portions of tendons and, in slightly different biochemical form, in fibrocartilage. Aggrecan has a role in the physiology and biomechanical function of tendons, ligaments and cardiovascular system through involvement in regulation of assembly and maintenance of extracellular matrix. Aggrecan participates in cell proliferation through interactions with growth factors.

[0070] In certain embodiments, at least one of the biomarkers is a tissue inhibitor of metalloproteinases 1 (TIMP-1). ΤΊΜΡ-1 is a glycoprotein a member of the TIMP family, and is a natural inhibitor of the matrix

metalloproteinases (MMPs), a group of peptidases involved in degradation of the extracellular matrix. In addition to its inhibitory role against most of the known MMPs, the encoded protein is able to promote cell proliferation in a wide range of cell types, and may also have an anti-apoptotic function.

[0071] In certain embodiments, at least one of the biomarkers is interleukin-1 (IL-1). IL-1 is known to have multiple and varied biological functions. T helper 17 (TH17) cells require IL-1 for their differentiation from naive T cells and the subsequent maintenance of their phenotype. IL-1 is also known as a proinflammatory cytokine and through expression of integrins on leukocytes and endothelial cells, initiates and regulates inflammatory

responses. The IL-1 family comprises 11 members: IL-1a, IL-Ιβ, IL- 1 receptor antagonist (IL-1 Ra), IL-18, IL-33 and IL- 1 F5-IL-1 F10.

[0072] In certain embodiments, at least one of the biomarkers is interleukin-17 (IL-17). IL-17 is a cytokine that acts as a potent mediator in delayed-type reactions by increasing chemokine production in various tissues to recruit monocytes and neutrophils to the site of inflammation, similar to

Interferon gamma (IFN-γ). IL-17 is produced by T-helper cells and is induced by IL-23 which results in destructive tissue damage in delayed-type reactions. Interleukin 17 as a family functions as a proinflammatory cytokine that responds to the invasion of the immune system by extracellular pathogens and induces destruction of the pathogen's cellular matrix. Interleukin 17 acts synergistically with tumor necrosis factor and interleukin-1.

[0073] In certain embodiments, at least one of the biomarkers is IL-6.

IL-6 ("Interleukin 6") is a cytokine encoded by the IL6 gene. IL-6 is involved in both inflammation and infection responses and in the regulation of metabolic, regenerative, and neural processes. [Scheller et al., Biochimica et Biophysica Acta 2011, 1813:878-888.]

[0074] In certain embodiments, at least one of the biomarkers is TNF, preferably TNFa. TNF ("tumor necrosis factor," "tumor necrosis factor-alpha," "TNF-a", or "cachectin") is a cytokine encoded by the TNF gene. TNF is involved in the regulation of immune responses.

[0075] In certain embodiments, at least one of the biomarkers is

CD11 b. CD11 b ("cluster of differentiation molecule 11 B," "integrin alpha M," "ITGAM," or "CR3A") is encoded by the ITGAM gene. CD11b is the alpha subunit of the integrin alpha-M beta-2 (α_Μβ₂) molecule, c^ is expressed in many cells types, including monocytes, granulocytes, macrophages, and natural killer cells. It mediates inflammation by regulating leukocyte adhesion and migration. [Solovjov D et al., J. Biol. Chem. 2005, 280 (2): 1336-45.]

[0076] In certain embodiments, at least one of the biomarkers is tartrate-resistant acid phosphatase 5b (TRAP -5b), also known as type-5 acid phosphatase and purple acid phosphatase. TRAP-5b is a glycosylated monomeric metalloprotein enzyme encoded by the TRAP gene. [0077] In certain embodiments, at least one of the biomarkers is collagen. In an embodiment, the collagen is free collagen. Collagen is known as a product of cartilage metabolism and is the most prevalent protein

component of cartilage. As the main component of connective tissue, it is one of the most abundant proteins in mammals. In an embodiment, the marker is collagen II ("Type II Collagen"). Collagen II is synthesized as procollagen alpha Col2A1 chains. Type II collagen contains three polypeptide chains of the alpha- 1(11) type, encoded by COL2A1 gene.

[0078] In certain embodiments, at least one of the biomarkers is

CD40, particularly CD40 levels on macrophages. CD40 ("cluster of

differentiation 40") is a transmembrane receptor of the tumor necrosis factor gene superfamily encoded by the CD40 gene. CD40 is a co-stimulatory molecule and is expressed on macrophages. [Suttles and Stout, Semin

Immunol. 2009, 21(5):257-64.]

[0079] In certain embodiments, at least one of the biomarkers is IL-

10. IL-10 ("Interleukin 10," "human cytokine synthesis inhibitory factor," or "CSIF") is a cytokine encoded by the IL10 gene. IL-10 has a central role in infection by inhibiting the activity of Th1 cells, NK cells, and macrophages, and thereby preventing tissue damage to the host. [Riley et al., The Journal of Immunology 2008, 180(9): 5771-5777.]

[0080] In certain embodiments, at least one of the biomarkers is IL-

13Ra2. IL-13Ra2 ("lnterleukin-13 receptor subunit alpha-2," "cluster of differentiation 213A2," or "CD213A2") is a membrane bound protein encoded by the IL13RA2 gene. IL-13Ra2 binds IL-13 with high affinity and internalizes, but it does not mediate signal transduction. [Kawakami K et al. Blood 2001 ; 97:2673- 9.]

[0081] In certain embodiments, at least one of the biomarkers is interleukin-15 (IL-15), particularly IL-15 levels on macrophages. IL-15 is a cytokine encoded by the IL15 gene. IL-15 mimics the stimulatory activity of IL-2 by binding to and signaling through a complex composed of IL-2/IL-15 receptor beta chain (CD122) and the common gamma chain (gamma-C, CD132).

Cytokines such as IL-15 contribute to regulation of the phenotype of effector and regulatory T cells in the synovium.

[0082] In certain embodiments, at least one of the biomarkers is GM-

CSF. GM-CSF ("granulocyte-macrophage colony-stimulating factor," "colony stimulating factor 2," or "CSF2") is a cytokine encoded by the CSF2 gene. GM- CSF effects on the inflammatory response influence by inducing differentiation, proliferation and activation of macrophages and dendritic cells. [Francisco-Cruz et al. Medical Oncology 2013, 31 :774.]

[0083] In certain embodiments, at least one of the biomarkers is MIP-

1a. MIP-1a ("macrophage inflammatory protein 1a," "chemokine (C-C motif) ligand 3," or "CCL3") is a cytokine encoded by the CCL3 gene. MIP-1a is produced by many cells types, including macrophages, dendritic cells, and lymphocytes. It has pro-inflammatory effects and can also promote

homoeostasis. [Maurer M and von Stebut E. The International Journal of Biochemistry and Cell Biology, 2004, 36(10): 1882-1886.]

[0084] In certain embodiments, at least one of the biomarkers is CD4

T cell adhesion molecule, preferably T cell-surface expression levels. CD4 ("cluster of differentiation 4" or "T cell surface protein T4") is a glycoprotein encoded by the CD4 gene. It is found on the surface of immune cells such as T helper cells, monocytes, macrophages, and dendritic cells. It has been implicated in the regulation of T-cell activation and in the associative recognition of class II antigens of the major histocompatibility complex. [Rudd et al. Proc Natl Acad Sci U S A. 1988, 85(14): 5190-5194.]

[0085] In certain embodiments, biomarkers are identified using the

Quantitative Systems Pharmacology (QSP) mathematical modeling approach. For instance, in some embodiments the Entelos® RA PhysioLab QSP model is used to identify suitable biomarkers and the extent to which those markers are expected to exhibit altered expression in an inflammatory disorder, relative to a level in a reference sample.

Biological Samples

[0086] A biological sample may be obtained, directly or indirectly, from any cell, tissue, organ, or fluid in a subject. In some embodiments, the biological sample can include a cell, tissue, fluid, organ, blood vessel, or bone taken from a subject. In other embodiments the biological sample is cartilage, bone, synovial membrane, synovial fluid, or another fluid from space within a joint cavity. In certain embodiments, the cell includes one of the following: an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a plasma cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a chondrocyte, a Kupffer cell, or a mastocyte. In other embodiments, the biological sample is whole blood, a dried blood spot or serum.

[0087] For example, the sample may be any fluid or component thereof, such as a fraction or extract, e.g., blood, plasma, lymph, synovial fluid, cystic fluid, urine, nipple aspirates, or fluids collected from a biopsy, amniotic fluid, aqueous humor, vitreous humor, bile, blood, breast milk, cerebrospinal fluid, cerumen, chyle, cystic fluid, endolymph, feces, gastric acid, gastric juice, mucus, pericardial fluid, perilymph, peritoneal fluid, plasma, pleural fluid, pus, saliva, sebum, semen, sweat, serum, sputum, synovial fluid, joint tissue or fluid, tears, or vaginal secretions obtained from the subject. In one embodiment, the fluid may be blood, or a component thereof, obtained from the subject, including whole blood or components thereof, including, plasma, serum, and blood cells, such as red blood cells, white blood cells and platelets. In another embodiment, the fluid may be synovial fluid, joint tissue or fluid, or any other sample reflective of an inflammatory disease (e.g., RA). The sample may also be any tissue or component thereof, connective tissue, lymph tissue or muscle tissue obtained from the subject.

[0088] Techniques or methods for obtaining samples from a subject are well known in the art and include, for example, obtaining samples by a mouth swab or a mouth wash; drawing blood; obtaining a biopsy; or obtaining synovial fluid or other sample from a subject suffering from inflammatory disease (e.g., skin, as in the case of psoriasis or psoriatic arthritis). Isolating components of fluid or tissue samples (e.g., cells or RNA or DNA) may be accomplished using a variety of techniques. After the sample is obtained, it may be further processed.

[0089] In various embodiments, the biomarker level from a patient sample is compared to the level in a reference sample. The reference sample may be any of the biological sample types mentioned above. It may be the same sample type collected from the patient, or a different sample type. [0090] In certain embodiments, the reference level is the level in a reference sample from a control subject that does not have an inflammatory disorder. In some embodiments, the reference level is the average level in samples from at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 subjects who do not have the inflammatory disorder. In other embodiments, the reference level is from one or more subjects who do not have the inflammatory disorder and are healthy adults.

[0091] In certain embodiments, the reference level comprises the level of the biomarker in a sample from one or more subjects who have the inflammatory disorder. In some embodiments, the reference level is the average level in samples from at least ten subjects who have the inflammatory disorder. In certain embodiments, the reference may be a sample from a patient with the inflammatory disorder prior to or during treatment (e.g., the same patient that is subsequently given a treatment following measurement, or the patient being monitored during treatment).

[0092] In some embodiments, the reference level is from a subject who has already received treatment. In other embodiments, the reference level is from a non-inflammatory tissue of a subject diagnosed with an inflammatory disorder.

[0093] In some embodiments, the reference level is normalized against another anti-inflammatory cytokine, such as IL-4, interferon-alpha (IFN- a) or transforming growth factor-beta (TGF-β).

Methods for Detecting the Expression Level of a Biomarker

[0094] The level of expression of a biomarker in a sample obtained from a subject or in a reference sample may be assayed by any of a wide variety of techniques and methods. In some embodiments, the techniques used to determine the level of one or more biomarkers in a sample include one or more of fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high- throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof. In some embodiments, the technique is a flow cytometry cell surface marker assay. In other embodiments, the technique is a multiplex cytokine or multiple chemokine assay. In some embodiments the technique is an ex vivo analysis of biomarkers following stimulation and the stimulation is by LPS, anti-CD3, anti-CD28 and combinations thereof. One of skill in the art recognizes the rapid development of new technology to detect a difference in levels between molecules. The methods disclosed herein encompass such alternate methods for measuring and determining the levels of biomarkers in a sample.

[0095] In one embodiment, the expression level of a biomarker in a sample can be assessed at the protein level. Methods of measuring protein levels include, but are not limited to, electrophoresis, capillary electrophoresis, high performance liquid chromatography (HPLC), thin layer chromatography (TLC), hyperdiffusion chromatography, and the like, or various immunological methods, using a detection reagent that detects the protein biomarker, directly or indirectly, such as fluid or gel precipitation reactions, immunohistochemistry, binding assay, immunodiffusion (single or double), immunoprecipitations, immunoelectrophoresis, radioimmunoassay (RIA), enzyme-linked immunosorbent assays (ELISAs), immunofluorescent assays, fluorescence- activated cell sorting (FACS) analysis, and Western blotting. Other suitable methods of measuring protein levels analysis include, but are not limited to, electron microscopy, mass spectrometry, e.g., MALDI-TOF and SELDI-TOF.

[0096] In one embodiment, the expression level of a biomarker in a sample can be determined at the nucleic acid level by detecting a transcribed polynucleotide, or portion thereof, e.g., mRNA, miRNA, hnRNA, or cDNA, of the biomarker gene. Methods of measuring polynucleotide levels include, but are not limited to, nuclear run-on assays, reverse-transcriptase PCR (RT- PCR)analysis, quantitative PCR analysis, real time PCR analysis, RNase protection assays, Northern blotting, Southern hybridizations, in situ

hybridization, high-throughput sequencing, and next generation sequencing, and RNA sequencing. Other suitable systems for mRNA sample analysis include, but are not limited to, microarray analysis.

[0097] In one embodiment, the expression level of a biomarker is measured using a flow cytometry cell surface marker assay. In one

embodiment, the expression level of a biomarker is measured using a multiplex cytokine or multiple chemokine assay. In one embodiment, the expression level of a biomarker is measured using an ex vivo analysis of biomarkers following stimulation. In one embodiment, the stimulation is by LPS, anti-CD3, anti-CD28 and combinations thereof.

[0098] In some embodiments, the expression level of a subset of biomarkers in the biological sample is measured. In one embodiment, the level of expression of collagen II measures the level of expression of free collagen II. In one embodiment, the level of expression of aggrecan measures the level of expression of free aggrecan. In one embodiment, the level of expression of TIMP-1 measures the level of expression of cartilage TIMP-1. In one

embodiment, the level of expression of CD40 measures the level of expression of macrophage surface CD40 (mac surface CD40). In one embodiment, the level of expression of TRAP-5b measures the level of expression of synovial TRAP-5b.

[0099] In general, where a difference in the level of expression of a biomarker and the reference level is to be detected, although this difference can be as small as the limit of detection of the method for determining the level of expression, it is preferred that the difference be greater than the limit of detection of the method or greater than the standard error of the assessment method. In some embodiments, a difference of at least about 2-, about 3-, about 4-, about 5-, about 6-, about 7-, about 8-, about 9-, about 10-, about 15-, about 20-, about 25-, about 100-, about 500-, 1000-fold greater than the standard error of the assessment method is detected. Alternatively, the difference may be greater than the limit of detection of the method or greater than the standard error of the assessment method, and preferably a difference of at least about 2-, about 3-, about 4-, about 5-, about 6-, about 7-, about 8-, about 9-, about 10-, about 15-, about 20-, about 25-, about 100-, about 500-, 1000-fold less than the standard error of the assessment method.

[00100] Any suitable sample obtained from a subject having an inflammatory disease (e.g., RA) may be used to assess the level of expression, including a lack of expression, of the biomarker. The biomarker may be, for example, collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, CD4, CD40, IL-15, GM-CSF, MIP-1a, TNF, CD11b, IL-6, IL-10, and IL-13Ra2.

[00101] In some embodiments, the activity level of a marker, or of IL-1 β and/or IL-17 is measured. Suitable methods for measuring activity levels of the proteins described herein are known in the art and can be used.

Kits

[00102] In still yet another aspect, the present disclosure provides a kit comprising means to measure the level of one or more biomarkers in a patient sample, and instructions for using the kit to measure the levels to the one or more biomarkers in a sample from a patient with an inflammatory disorder, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, CD4, CD40, IL-15, GM-CSF, ΜΙΡ-Ια, TNF, CD11b, IL-6, IL-10, and IL-13Ra2. The kits of the invention can optionally comprise additional components useful for performing the methods of the invention. In one embodiment, comprising one or more therapeutic agents that antagonize IL-1 β and IL-17, and/or instructions to administer the agents before or after measuring the one or more biomarkers.

Examples

[00103] The following examples serve to illustrate, and in no way limit, the present disclosure.

Example 1: Summary Diagram

[00104] Starting with the Entelos® RA PhysioLab (IEE Proc.-Syst.

Biol., Vol. 152, No. 4, December 2005), further qualification of the reference virtual patient was added using clinical readout from approved RA therapies, specifically Humira, Rituximab, Tocilizumab, Secukinumab, and Abatacept, generated the RA QSP model (RA QSP Model, Figure 2). The RA QSP

Platform (FIG. 2) is focused on the inflammatory network dynamics and impact of the network modulation on bone and cartilage turnover in a joint with established chronic inflammation and RA disease. In addition, the life cycle of multiple cell types, thought to play a role in RA pathogenesis, as well as their products and interactions is represented. Secukinumab PKPD structure is represented, including direct inhibition of "Effective Synovial IL-17". Abatacept activity was represented as a direct impact on CD8 TCR stimulation, PB CD8 TCR stimulation, and CD4 TCR-dependent stimulation. The reference virtual patient was qualified for the two aforementioned therapies as well as for Methotrexate (MTX), Humira, Anakinra, Rituximab, and Tocilizumab.

Qualification of the virtual patient was confirmed by matching the clinical and biomarkers response from the corresponding in vivo testing data (see below). Example 2: Virtual Patient Qualification

[00105] Qualification of the Reference Virtual Patient (VP) response to the key RA therapies was conducted in sequence. The Reference VP was qualified by comparing the in silico clinical impact of the implemented therapies with in vivo data from RA animal testing. The Virtual Patient Qualification was confirmed by comparing the mean improvement in ACR score extracted from testing data with the simulated ACR-N (numerical ACR) score of the reference virtual patient to the corresponding therapies (FIG. 3). The simulated ACR-N is a model output that is based on the percentage change in overall tissue cellularity. Example 3: Mouse CIA Study

[00106] Activity of anti-mlL17, anti-mlL-1 β, and the combination was evaluated in the mouse cartilage induced arthritis (CIA) mouse model.

Significant improvement in the inflammatory, bone, and cartilage markers were observed with the combination therapy compared to the parental monotherapies (FIG. 4). Significant effects on histopathology scores of inflammation, cartilage, and bone were observed in the anti-IL17/anti-IL1 β combination group compared to anti-IL17 or anti-ILip treated DBA-1 CIA mice. Anti-TNFa was the positive control. RA animal model data suggests that the anti-IL17/anti-IL1 β

combination is superior to parental monotherapies, especially on cartilage and bone health.

Example 4: QSP Analysis

[00107] The QSP analysis identified intact free collagen, aggrecan,

TRAP-5b, and TIMP1 as biomarkers modulated in a synergistic fashion by IL- 1 β/ILI 7 combination therapy larger than the additive modulation by the parental monotherapies in RA (FIG 5). Other markers and their predicted effect in the QSP prediction, and their measured effect during ex vivo stimulation of normal cynomolgus monkey blood, are shown in the table below. Key disease modification biomarkers were identified for further experimental confirmation in mouse paw testing.

[00108] Table 4 summarizes results of the stimulation experiments and the QSP projected effect. The QSP analysis projected impact on the listed biomarkers was comparable to the observations from the ex vivo stimulation of normal cynomolgus monkey blood in a tolerability study (described in Example 6).

Example 5: Gene Expression Profiles

[00109] The results from Example 4 guided mining of the mouse paw gene array data, which confirmed and linked these biomarkers to RA disease modification (FIG. 6). The gene expression profiles of aggrecan, TRAP-5b, and TIMP-1 in nafve mice or DBA-1 CIA mice treated with vehicle, anti-IL.17, anti- ΙΙ_1β, or anti-IL17/anti-IL1 β combination therapy experimentally confirmed the QSP predictions. Anti-TNFa was the positive control. The QSP findings were verified experimentally at the gene expression level. FIG. 6 shows mRNA expression levels for the measured markers in the different treatment conditions.

Example 6: Cynomolgus Monkey Tolerability Study

[00110] 1 female, 1 male/group were administered DVD 3418 (anti-

IL1p/anti-IL17 DVD-lg) at 20 and 100 mg/kg iv, 20 mg/kg sc once per week for 6 weeks. Blood samples for ex vivo analysis were collected at: predose, D22 pre- dose, D43 (1 week post dose), and 6 week recovery. Ex vivo stimulation was achieved either by LPS (primarily monocyte) or CD3/28 (primary cells). Two separate bio-assays were utilized; a flow cytometry 9 cell surface marker assay was used to evaluate markers on B cell, T cell, monocyte & granulocytes, and a Multiplex cytokine/chemokine analysis (13 analytes) bioassay was used to determine the level of the target soluble mediators.

[00111] The Table in Example 4 above summarizes results of the stimulation experiments and the associated QSP projected effect. Even with the limited animal numbers and high dose levels utilized in toxicity studies, decreases in cell surface biomarkers CD11b and IL-13Ra2, as well as decreases in TNFa, GM-CSF, IL-6, MIP-1a and IL-10, were observed following ex vivo stimulations of samples after treatment with DVD3418.

Example 7: Schematic Depiction of Mediators and Biomarkers Modulated by the DVD-lg therapy

[00112] Tracing back the mechanism of action of the anti-IL1 β/anti-

IL17 combination generated further insights regarding bone protection activity of the therapy rather than stimulation of bone anabolism (FIG. 7). FIG. 7 shows a schematic depiction of the mediators that play a role in regulating chondrocyte production, the key biomarkers modulated by the anti-IL1 p/anti-IL17 therapy, and the postulated effect of the combination therapy on cartilage health as represented in the QSP RA platform.

Incorporation by Reference

[00113] The contents of all cited references (including literature references, patents, patent applications, and websites) that maybe cited throughout this application are hereby expressly incorporated by reference in their entirety for any purpose, as are the references cited therein. Equivalents

[00114] The disclosure may be embodied in other specific forms without departing from the spirit or essential characteristics thereof. The foregoing embodiments are therefore to be considered in all respects illustrative rather than limiting of the disclosure. Scope of the disclosure is indicated by the appended claims rather than by the foregoing description, and all changes that come within the meaning and range of equivalency of the claims are therefore intended to be embraced herein.

Claims

WHAT IS CLAIMED IS

1. A method of treating a human subject having an inflammatory disorder, comprising:

(a) determining the level of one or more biomarkers in a biological sample from a subject, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11 b, IL-6, IL-10, IL-15, and IL-13Ra2; and

(b) administering one or more therapeutic agents that antagonize IL-1 β and IL-17 if the subject has a level of the biomarker that is greater than a reference level.

2. A method of treating a human subject having an inflammatory disorder, comprising:

(a) measuring the level of one or more biomarkers in the subject, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan. TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11b, IL-6, IL-10, IL-15, and IL-13Ra2;

(b) determining whether the one or more biomarkers are elevated relative to a reference level; and

(c) administering one or more therapeutic agents that antagonize IL-1 β and IL-17 if the subject has a level of the biomarker that is greater than a reference level.

3. The method of claim 1 , further comprising repeating the process of steps (a)- (b) until the level of the biomarker collected from the biological sample is equal to or less than the reference level.

4. The method of any of claims 1-3, wherein determining the level of the biomarker in the subject comprises:

(a) collecting the biological sample from the subject;

(b) measuring the level of one or more of the biomarkers in the biological sample; and (b) comparing the level of the biomarker in the sample to the reference level.

5. The method of any one of claims 1-4, further comprising maintaining, decreasing, or discontinuing administration of the therapeutic agent if the level of the biomarker in the subject is determined to be less than or equal to the reference level.

6. The method of any one of claims 1-5, wherein each successive

administration of the therapeutic agent is administered one, two, three, four, five, six, or seven days, or one, two, three, or four weeks after the previous dose.

7. The method of any one of claims 1-6, wherein each administration of the therapeutic agent therapy is administered at least twice.

8. The method of any one of claims 1-7, wherein the biological sample is a cell, tissue, fluid, organ, blood vessel, or bone.

9. The method of claim 8, wherein the cell is at least one of: an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a plasma cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a

chondrocyte, a Kupffer cell, or a mastocyte.

10. The method of claim 8, wherein the biological sample is cartilage, bone, synovial membrane, synovial fluid, or another fluid from space within a joint cavity.

11. The method of claim 8, wherein the biological sample is whole blood,

12. The method of claim 11 , wherein the whole blood is a dried blood spot.

13. The method of claim 8, wherein the biological sample is serum.

14. The method of claim 1 or 2, wherein the biomarker comprises at least one of: aggrecan, TIMP-1 , and TRAP-5b.

15. The method of any one of claims 1-7, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who do not have the inflammatory disorder.

16. The method of claim 15, wherein the reference level is the average level in samples from at least ten subjects who do not have the inflammatory disorder.

17. The method of claim 15 or 16, wherein the subjects who do not have the inflammatory disorder are healthy adults.

18. The method of any one of claims 1-7, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who have the inflammatory disorder.

19. The method of claim 18, wherein the reference level is the average level in samples from at least ten subjects who have the inflammatory disorder.

20. The method of claim 18 or 19, wherein the subjects who have the inflammatory disorder are healthy adults.

21. The method of any of one of claims 1-7, wherein administering the one or more therapeutic agents comprises simultaneously or sequentially

administering an anti-IL-1 β antagonist and an anti-IL-17 antagonist.

22. The method of claim 21 , wherein the anti-IL-1 β antagonist is an anti-IL-1 β antibody and the anti-IL17 antagonist is an anti-IL-17 antibody.

23. The method of claim 1 , wherein the therapeutic agent is a bispecific binding protein capable of binding IL-1 β and IL-17.

24. The method of claim 23, wherein the bispecific binding protein is a dual variable domain immunoglobulin (DVD-lg).

25. The method of claim 24, wherein the DVD-lg comprises

(i) a binding site for IL-1 β, comprising:

and

(ii) a binding site for IL-17, comprising:

(a) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 13, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 14.

26. The method of claim 24, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising:

and

(ii) variable domains that form a functional target binding site for IL-17 comprising:

SEQ ID NO: 13 and SEQ ID NO: 14.

27. The method of claim 24, wherein the DVD-lg comprises SEQ ID NO: 19 and SEQ ID NO: 20.

28. The method of claim 24, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising CDRs 1-3 from SEQ ID NO: 3 and CDRs 1-3 from SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising CDRs 1-3 from SEQ ID NO: 13 and CDRs 1-3 from SEQ ID NO: 14.

29. The method of claim 24, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising SEQ ID NO: 3 and SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising SEQ ID NO: 13 and SEQ ID NO: 14.

30. The method of any one of claims 1-29, wherein the therapeutic agent is administered intravenously, intradermally, subcutaneously, or intramuscularly,

31. The method of any one of claims 1-30, wherein the inflammatory disorder is Alzheimer's disease, juvenile idiopathic arthritis, psoriasis, plaque psoriasis, keratoconjunctivitis sicca, blepharitis, keratitis, ankylosing spondylitis, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, asthma, atherosclerosis, Crohn's disease, colitis, inflammatory bowel disease (IBD), insulin dependent diabetes mellitus, chronic obstructive pulmonary disease (COPD), cystic fibrosis, urticaria, ectopic eczema, multiple sclerosis, dermatitis, diverticulitis, fibromyalgia, hepatitis, irritable bowel syndrome (IBS), systemic lupus erythematous (SLE), nephritis, Parkinson's disease, or ulcerative colitis.

32. The method of claim 31 , wherein the inflammatory disorder is rheumatoid arthritis.

33. The method of any one of claims 1-32, wherein the determining of the level of one or more biomarkers in the biological sample from the subject comprises using a technique selected from the group consisting of fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high-throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked

immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof.

34. The method of claim 33, wherein the technique is a flow cytometry cell surface marker assay.

35. The method of claim 33, wherein the technique is a multiplex cytokine or multiple chemokine assay.

36. The method of claim 33, wherein the technique is an ex vivo analysis of biomarkers following stimulation.

37. The method of claim 36, wherein the stimulation is by LPS, anti-CD3, anti- CD28 and combinations thereof.

38. A method of treating a human subject having an inflammatory disorder, comprising:

(a) administering one or more therapeutic agents that antagonize IL-1 β and IL-17;

(b) determining the level of one or more biomarkers in a biological sample from the subject, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11 b, IL-6, IL-10, IL-15, and IL-13Ra2; (c) comparing the levels of the biomarkers in the sample to a reference level; and

(d) administering a further dose of the one or more therapeutic agents if the levels of the one or more biomarkers in the biological sample are greater than the reference level.

39. The method of claim 38, further comprising repeating steps (a)-(d) until the level of the biomarker is less than or equal to the reference level.

40. The method of claim 38 or 39, wherein administering the one or more therapeutic agents comprises simultaneously or sequentially administering an anti-IL-1 β antagonist and an anti-IL-17 antagonist.

41. The method of claim 40, wherein the anti-IL-1 β antagonist is an anti-IL-1 β antibody and the anti-IL17 antagonist is an anti-IL-17 antibody.

42. The method of claim 38, wherein the therapeutic agent is a bispecific binding protein capable of binding IL-1 β and IL-17.

43. The method of claim 42, wherein the bispecific binding protein is a dual variable domain immunoglobulin (DVD-lg).

44. The method of claim 43, wherein the DVD-lg comprises

(i) a binding site for IL-1 β, comprising:

CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 10;

and

(ii) a binding site for IL-17, comprising:

(a) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 13, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 14.

45. The method of claim 43, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising:

(1) SEQ ID NO: 1 and SEQ ID NO:2,

(2) SEQ ID NO: 3 and SEQ ID NO: 4,

(3) SEQ ID NO: 5 and SEQ ID NO: 6,

(4) SEQ ID NO: 7 and SEQ ID NO: 8, or

(5) SEQ ID NO: 9 and SEQ ID NO: 10;

and

(ii) variable domains that form a functional target binding site for IL-17 comprising:

SEQ ID NO: 13 and SEQ ID NO: 14.

46. The method of claim 43, wherein the DVD-lg comprises SEQ ID NO: 19and SEQ ID NO: 20.

47. The method of claim 43, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising CDRs 1-3 from SEQ ID NO: 3 and CDRs 1-3 from SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising CDRs 1-3 from SEQ ID NO: 13 and CDRs 1-3 from SEQ ID NO: 14.

48. The method of claim 43, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising SEQ ID NO: 3 and SEQ ID NO: 4, and (ii) variable domains that form a functional target binding site for IL-17 comprising SEQ ID NO: 13 and SEQ ID NO: 14.

49. The method of claim 38, further comprising maintaining, decreasing, or discontinuing administration of the therapeutic agent when the level of the biomarker in the subject is determined to be less than or equal to the reference level.

50. The method of claim 38 or 39, wherein each successive administration of the therapeutic agent is administered one, two, three, four, five, six, or seven days, or one, two, three, or four weeks after the previous dose.

51. The method of claim 38 or 39, wherein the biological sample is a cell, tissue, fluid, organ, blood vessel, or bone.

52. The method of claim 51 , wherein the cell is at least one of: an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a Plasma Cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a chondrocyte, a Kupffer cell, and a mastocyte.

53. The method of claim 51 , wherein the biological sample is cartilage, bone, synovial membrane, synovial fluid, or another fluid from space within a joint cavity.

54. The method of claim 51 , wherein the biological sample is whole blood,

55. The method of claim 54, wherein the whole blood is a dried blood spot.

56. The method of claim 51 , wherein the biological sample is serum.

57. The method of any one of claims 38-56, wherein the therapeutic agent is administered intravenously, intradermally, subcutaneously, or intramuscularly.

58. The method of any one of claims 38-57, wherein the biomarker comprises at least one of: aggrecan, TIMP-1 , and TRAP-5b.

59. The method of any one of claims 38-58, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who do not have the inflammatory disorder.

60. The method of claim 59, wherein the reference level is the average level in samples from at least ten subjects who do not have the inflammatory disorder.

61. The method of claim 59 or 60, wherein the subjects who do not have the inflammatory disorder are healthy adults.

62. The method of any one of claims 38-58, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who have the inflammatory disorder.

63. The method of claim 62, wherein the reference level is the average level in samples from at least ten subjects who have the inflammatory disorder.

64. The method of claim 62 or 63, wherein the subjects who have the inflammatory disorder are healthy adults.

65. A method of monitoring a subject during treatment of an inflammatory disorder, the method comprising administering to the subject a dose of one or more therapeutic agents that antagonize IL-1 β and IL-17, collecting a biological sample from the subject after the administration of the dose of therapeutic agent, measuring the level of one or more biomarkers in the sample, and comparing the levels of the biomarkers in the sample to reference levels, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD1 1b, IL-6, IL-10, IL-15, and IL-13Ra2.

66. The method of claim 65, wherein the inflammatory disorder is Alzheimer's disease, juvenile idiopathic arthritis, psoriasis, plaque psoriasis,

keratoconjunctivitis sicca, blepharitis, keratitis, ankylosing spondylitis, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, asthma, atherosclerosis, Crohn's disease, colitis, inflammatory bowel disease (IBD), insulin dependent diabetes mellitus, chronic obstructive pulmonary disease (COPD), cystic fibrosis, urticaria, ectopic eczema, multiple sclerosis, dermatitis, diverticulitis, fibromyalgia, hepatitis, irritable bowel syndrome (IBS), systemic lupus

erythematous (SLE), nephritis, Parkinson's disease, or ulcerative colitis.

67. The method of claim 66, wherein the inflammatory disorder is rheumatoid arthritis.

68. The method of any one of claims 65-67, wherein the determining of the level of one or more biomarkers in the biological sample from the subject comprises using a technique selected from the group consisting of

fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high-throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof.

69. The method of claim 68, wherein the technique is a flow cytometry cell surface marker assay.

70. The method of claim 68, wherein the technique is a multiplex cytokine or multiple chemokine assay.

71. The method of claim 68, wherein the technique is an ex vivo analysis of biomarkers following stimulation.

72. The method of claim 71 , wherein the stimulation is by LPS, anti-CD3, anti- CD28 and combinations thereof.

73. The method of claim 65, wherein administering the dose of one or more therapeutic agents comprises simultaneously or sequentially administering an anti-IL-1 β antagonist and an anti-IL-17 antagonist.

74. The method of claim 73, wherein the anti-IL-1 β antagonist is an anti-IL-1 β antibody and the anti-IL17 antagonist is an anti-IL-17 antibody.

75. The method of claim 65, wherein the therapeutic agent is a bispecific binding protein capable of binding IL-1 β and IL-17.

76. The method of claim 75, wherein the bispecific binding protein is a dual variable domain immunoglobulin (DVD-lg).

77. The method of claim 76, wherein the DVD-lg comprises

(i) a binding site for IL-1 β, comprising:

(a) CDR H1 CDR H2 and CDR H3 of SEQ ID NO: 1 and

and

(ii) a binding site for IL-17, comprising:

(a) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 13, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 14.

78. The method of claim 76, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-Ιβ comprising:

(1) SEQ ID NO: 1 and SEQ ID NO:2,

(2) SEQ ID NO: 3 and SEQ ID NO: 4,

(3) SEQ ID NO: 5 and SEQ ID NO: 6,

(4) SEQ ID NO: 7 and SEQ ID NO: 8, or

(5) SEQ ID NO: 9 and SEQ ID NO: 10;

and

(ii) variable domains that form a functional target binding site for IL-17 comprising:

SEQ ID NO: 13 and SEQ ID NO: 14.

79. The method of claim 76, wherein the DVD-lg comprises SEQ ID NO: 19 and SEQ ID NO: 20.

80. The method of claim 76, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising CDRs 1-3 from SEQ ID NO: 3 and CDRs 1-3 from SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising CDRs 1-3 from SEQ ID NO: 13 and CDRs 1-3 from SEQ ID NO: 14.

81. The method of claim 76, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising SEQ ID NO: 3 and SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising SEQ ID NO: 13 and SEQ ID NO: 14.

82. The method of any one of claims 65-81 , further comprising administering an additional dose of the one or more therapeutic agents if the an elevated level of the one or more biomarkers is detected relative to the reference level.

83. The method of claim 82, further comprising administering a reduced dose of the one or more therapeutic agents or discontinuing treatment if a reduced level of the one or more biomarkers is detected relative to the reference level.

84. The method of claim 65, wherein the biological sample is a cell, tissue, fluid, organ, blood vessel, or bone.

85. The method of claim 84, wherein the cell is at least one of: an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a Plasma Cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a chondrocyte, a Kupffer cell, or a mastocyte.

86. The method of claim 84, wherein the biological sample is cartilage, bone, synovial membrane, synovial fluid, or another fluid within the joint cavity.

87. The method of claim 84, wherein the biological sample is whole blood,

88. The method of claim 87, wherein the whole blood is a dried blood spot,

89. The method of claim 84, wherein the biological sample is serum.

90. The method of any one of claims 65-89, wherein the therapeutic agent is administered intravenously, intradermally, subcutaneously, or intramuscularly,

91. The method of any one of claims 65-90, wherein the biomarker comprises at least one of the following: aggrecan, TIMP-1 , TRAP-5b.

92. The method of any one of claims 65-91 , wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who do not have the inflammatory disorder.

93. The method of claim 92, wherein the reference level is the average level in samples from at least ten subjects who do not have the inflammatory disorder.

94. The method of claim 92 or 93, wherein the subjects who do not have the inflammatory disorder are healthy adults.

95. The method of any one of claims 65-91 , wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who have the inflammatory disorder.

96. The method of claim 95, wherein the reference level is the average level in samples from at least ten subjects who have the inflammatory disorder.

97. The method of claim 95 or 96, wherein the subjects who have the inflammatory disorder are healthy adults.

98. The method of one of claims 65-97, wherein the therapeutic agent is not administered if the biomarker level is less than or equal to the reference level.

99. A method of diagnosing an inflammatory disorder in a subject, comprising:

(a) collecting a biological sample from the subject;

(b) measuring the level of one or more biomarkers in the sample, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11 b, IL-6, IL-10, IL-15, and IL-13Ra2;

(c) comparing the level of the biomarker in the sample to a reference level; and

(e) diagnosing an inflammatory disorder if the level of the biomarker is greater than the reference level.

100. The method of claim 99, wherein the inflammatory disorder is Alzheimer's disease, juvenile idiopathic arthritis, psoriasis, plaque psoriasis,

keratoconjunctivitis sicca, blepharitis, keratitis, ankylosing spondylitis, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, asthma, atherosclerosis, Crohn's disease, colitis, inflammatory bowel disease (IBD), insulin dependent diabetes mellitus, chronic obstructive pulmonary disease (COPD), cystic fibrosis, urticaria, ectopic eczema, multiple sclerosis, dermatitis, diverticulitis, fibromyalgia, hepatitis, irritable bowel syndrome (IBS), systemic lupus

erythematous (SLE), nephritis, Parkinson's disease, or ulcerative colitis.

101. The method of claim 100, wherein the inflammatory disorder is rheumatoid arthritis.

102. The method of any one of claims 99-101 , wherein the determining of the level of one or more biomarkers in the biological sample from the subject comprises using a technique selected from the group consisting of

fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high-throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof.

103. The method of claim 102, wherein the technique is a flow cytometry cell surface marker assay.

104. The method of claim 102, wherein the technique is a multiplex cytokine or multiple chemokine assay.

105. The method of claim 102, wherein the technique is an ex vivo analysis of biomarkers following stimulation.

106. The method of claim 105, wherein the stimulation is by LPS, anti-CD3, anti-CD28 and combinations thereof.

107. The method of claim 99, wherein administering the dose of one or more therapeutic agents comprises simultaneously or sequentially administering an anti-IL-1 β antagonist and an anti-IL-17 antagonist.

108. The method of claim 107, wherein the anti-IL-1 β antagonist is an anti-IL- 1 β antibody and the anti-IL17 antagonist is an anti-IL-17 antibody.

109. The method of claim 99, wherein the therapeutic agent is a bispecific binding protein capable of binding IL-1 β and IL-17.

110. The method of claim 109, wherein the bispecific binding protein is a dual variable domain immunoglobulin (DVD-lg).

1 11. The method of claim 110, wherein the DVD-lg comprises

(i) a binding site for IL-1 β, comprising:

and

(ii) a binding site for IL-17, comprising:

(a) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 13, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 14.

112. The method of claim 110, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising:

and

(ii) variable domains that form a functional target binding site for IL-17 comprising:

SEQ ID NO: 13 and SEQ ID NO: 14.

113. The method of claim 110, wherein the DVD-lg comprises SEQ ID NO: 19 and SEQ ID NO: 20.

1 14. The method of claim 110, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising CDRs 1-3 from SEQ ID NO: 3 and CDRs 1-3 from SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising CDRs 1-3 from SEQ ID NO: 13 and CDRs 1-3 from SEQ ID NO: 14.

115. The method of claim 110, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising SEQ ID NO: 3 and SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising SEQ ID NO: 13 and SEQ ID NO: 14.

116. The method of any one of claims 99-115, further comprising administering an additional dose of the one or more therapeutic agents if the an elevated level of the one or more biomarkers is detected relative to the reference level.

117. The method of claim 116, further comprising administering a reduced dose of the one or more therapeutic agents or discontinuing treatment if a reduced level of the one or more biomarkers is detected relative to the reference level.

1 18. The method of claim 99, wherein the biological sample is a cell, tissue, fluid, organ, blood vessel, or bone.

119. The method of claim 118, wherein the cell is at least one of: an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a Plasma Cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a chondrocyte, a Kupffer cell, or a mastocyte.

120. The method of claim 118, wherein the biological sample is cartilage, bone, synovial membrane, synovial fluid, or another fluid within the joint cavity.

121. The method of claim 118, wherein the biological sample is whole blood,

122. The method of claim 121 , wherein the whole blood is a dried blood spot.

123. The method of claim 118, wherein the biological sample is serum,

124. The method of any one of claims 99-123, wherein the therapeutic agent is administered intravenously, intradermal^, subcutaneously, or intramuscularly.

125. The method of any one of claims 99-124, wherein the biomarker comprises at least one of the following: aggrecan, TIMP-1 , TRAP-5b.

126. The method of any one of claims 99-125, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who do not have the inflammatory disorder.

127. The method of claim 126, wherein the reference level is the average level in samples from at least ten subjects who do not have the inflammatory disorder.

128. The method of claim 126 or 127, wherein the subjects who do not have the inflammatory disorder are healthy adults.

129. The method of any one of claims 99-125, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who have the inflammatory disorder.

130. The method of claim 129, wherein the reference level is the average level in samples from at least ten subjects who have the inflammatory disorder.

131. The method of claim 129 or 130, wherein the subjects who have the inflammatory disorder are healthy adults.

132. The method of one of claims 99-131 , wherein the therapeutic agent is not administered if the biomarker level is less than or equal to the reference level.

133. A method of evaluating the effectiveness of a treatment for an

inflammatory disorder, comprising administering to the subject one or more therapeutic agents that antagonize IL-1 β and IL-17, collecting a biological sample from the subject after the administration of the therapeutic agents, measuring the level of one or more biomarkers in the sample, and comparing the levels of the biomarkers in the sample to reference levels, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, ΜΙΡ-Ια, TNF (preferably TNFa), CD11b, IL-6, IL-10, IL-15, and IL-13Ra2, and wherein a level equal to or less than the reference level indicates an effective treatment.

134. The method of claim 133, wherein the inflammatory disorder is

Alzheimer's disease, juvenile idiopathic arthritis, psoriasis, plaque psoriasis, keratoconjunctivitis sicca, blepharitis, keratitis, ankylosing spondylitis, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, asthma, atherosclerosis, Crohn's disease, colitis, inflammatory bowel disease (IBD), insulin dependent diabetes mellitus, chronic obstructive pulmonary disease (COPD), cystic fibrosis, urticaria, ectopic eczema, multiple sclerosis, dermatitis, diverticulitis, fibromyalgia, hepatitis, irritable bowel syndrome (IBS), systemic lupus

erythematous (SLE), nephritis, Parkinson's disease, or ulcerative colitis.

135. The method of claim 134, wherein the inflammatory disorder is rheumatoid arthritis.

136. The method of any one of claims 133-135, wherein the determining of the level of one or more biomarkers in the biological sample from the subject comprises using a technique selected from the group consisting of

fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high-throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof.

137. The method of claim 136, wherein the technique is a flow cytometry cell surface marker assay.

138. The method of claim 136, wherein the technique is a multiplex cytokine or multiple chemokine assay.

139. The method of claim 136, wherein the technique is an ex vivo analysis of biomarkers following stimulation.

140. The method of claim 139, wherein the stimulation is by LPS, anti-CD3, anti-CD28 and combinations thereof.

141. The method of claim 133, wherein administering the dose of one or more therapeutic agents comprises simultaneously or sequentially administering an anti-IL-1 β antagonist and an anti-IL-17 antagonist.

142. The method of claim 141 , wherein the anti-IL-1 β antagonist is an anti-IL- 1 β antibody and the anti-IL17 antagonist is an anti-IL-17 antibody.

143. The method of claim 133, wherein the therapeutic agent is a bispecific binding protein capable of binding IL-1 β and IL-17.

144. The method of claim 143, wherein the bispecific binding protein is a dual variable domain immunoglobulin (DVD-lg).

145. The method of claim 144, wherein the DVD-lg comprises

(i) a binding site for IL-1 β. comprising:

and

(ii) a binding site for IL-17, comprising:

(a) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 13, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 14.

146. The method of claim 144, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising: (1) SEQ ID NO: 1 and SEQ ID NO:2,

(2) SEQ ID NO: 3 and SEQ ID NO: 4,

(3) SEQ ID NO: 5 and SEQ ID NO: 6,

(4) SEQ ID NO: 7 and SEQ ID NO: 8, or

(5) SEQ ID NO: 9 and SEQ ID NO: 10;

and

(ii) variable domains that form a functional target binding site for IL-17 comprising:

SEQ ID NO: 13 and SEQ ID NO: 14.

147. The method of claim 144, wherein the DVD-lg comprises SEQ ID NO: 19 and SEQ ID NO: 20.

148. The method of claim 144, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising CDRs 1-3 from SEQ ID NO: 3 and CDRs 1-3 from SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising CDRs 1-3 from SEQ ID NO: 13 and CDRs 1-3 from SEQ ID NO: 14.

149. The method of claim 144, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising SEQ ID NO: 3 and SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising SEQ ID NO: 13 and SEQ ID NO: 14.

150. The method of any one of claims 133-149, further comprising

administering an additional dose of the one or more therapeutic agents if the an elevated level of the one or more biomarkers is detected relative to the reference level.

151. The method of claim 150, further comprising administering a reduced dose of the one or more therapeutic agents or discontinuing treatment if a reduced level of the one or more biomarkers is detected relative to the

reference level.

152. The method of claim 133, wherein the biological sample is a cell, tissue, fluid, organ, blood vessel, or bone.

153. The method of claim 152, wherein the cell is at least one of: an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a Plasma Cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a chondrocyte, a Kupffer cell, or a mastocyte.

154. The method of claim 152, wherein the biological sample is cartilage, bone, synovial membrane, synovial fluid, or another fluid within the joint cavity.

155. The method of claim 152, wherein the biological sample is whole blood.

156. The method of claim 155, wherein the whole blood is a dried blood spot,

157. The method of claim 152, wherein the biological sample is serum,

158. The method of any one of claims 133-157, wherein the therapeutic agent is administered intravenously, intradermal^, subcutaneously, or intramuscularly.

159. The method of any one of claims 133-158, wherein the biomarker comprises at least one of the following: aggrecan, TIMP-1 , TRAP^»5b.

160. The method of any one of claims 133-159, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who do not have the inflammatory disorder.

161. The method of claim 160, wherein the reference level is the average level in samples from at least ten subjects who do not have the inflammatory disorder.

162. The method of claim 160 or 161 , wherein the subjects who do not have the inflammatory disorder are healthy adults.

163. The method of any one of claims 133-159, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who have the inflammatory disorder.

164. The method of claim 163, wherein the reference level is the average level in samples from at least ten subjects who have the inflammatory disorder.

165. The method of claim 163 or 164, wherein the subjects who have the inflammatory disorder are healthy adults.

166. The method of one of claims 133-165, wherein the therapeutic agent is not administered if the biomarker level is less than or equal to the reference level.

167. A kit comprising means to measure the level of one or more biomarkers in a patient sample, and instructions for using the kit to measure the levels to the one or more biomarkers in a sample from a patient with an inflammatory disorder, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, MIP-1a, TNF (preferably TNFa), CD11 b, IL-6, IL-10, IL-15, and IL-13Ra2.

168. The kit of claim 167, further comprising one or more therapeutic agents that antagonize IL-1 β and IL-17.

169. A method of treating an inflammatory disorder in a subject, comprising (a) administering to the subject a first dose of one or more therapeutic agents that antagonize IL-1 β and IL-17,

(b) collecting a biological sample from the subject after the administration of the therapeutic agents,

(c) measuring the level of one or more biomarkers in the sample,

(d) comparing the levels of the biomarkers in the sample to reference levels, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, ΜΙΡ-Ια, TNF (preferably TNFa), CD1 1b, IL-6, IL-10, IL-15, and IL-13Ra2, and

(e) administering a higher concentration second dose if the levels of the biomarkers are greater than the reference levels, or administering a maintained or lower concentration second dose if the levels of the biomarkers are less than the reference levels.

170. The method of claim 169, wherein the inflammatory disorder is

Alzheimer's disease, juvenile idiopathic arthritis, psoriasis, plaque psoriasis, keratoconjunctivitis sicca, blepharitis, keratitis, ankylosing spondylitis, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, asthma, atherosclerosis, Crohn's disease, colitis, inflammatory bowel disease (IBD), insulin dependent diabetes mellitus, chronic obstructive pulmonary disease (COPD), cystic fibrosis, urticaria, ectopic eczema, multiple sclerosis, dermatitis, diverticulitis, fibromyalgia, hepatitis, irritable bowel syndrome (IBS), systemic lupus

erythematous (SLE), nephritis, Parkinson's disease, or ulcerative colitis.

171. The method of claim 170, wherein the inflammatory disorder is rheumatoid arthritis.

172. The method of any one of claims 169-171 , wherein the determining of the level of one or more biomarkers in the biological sample from the subject comprises using a technique selected from the group consisting of

fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high-throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof.

173. The method of claim 172, wherein the technique is a flow cytometry cell surface marker assay.

174. The method of claim 172, wherein the technique is a multiplex cytokine or multiple chemokine assay.

175. The method of claim 172, wherein the technique is an ex vivo analysis of biomarkers following stimulation.

176. The method of claim 175, wherein the stimulation is by LPS, anti-CD3, anti-CD28 and combinations thereof.

177. The method of claim 169, wherein administering the dose of one or more therapeutic agents comprises simultaneously or sequentially administering an anti-IL-Ιβ antagonist and an anti-IL-17 antagonist.

178. The method of claim 177, wherein the anti-IL-Ι β antagonist is an anti-IL- 1 β antibody and the anti-IL17 antagonist is an anti-IL-17 antibody.

179. The method of claim 169, wherein the therapeutic agent is a bispecific binding protein capable of binding IL-1 β and IL-17.

180. The method of claim 179, wherein the bispecific binding protein is a dual variable domain immunoglobulin (DVD-lg).

181. The method of claim 180, wherein the DVD-lg comprises

(i) a binding site for IL-1 β. comprising:

(a) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 1 , and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 2;

(b) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 3, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 4;

(c) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 5, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 6;

(d) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 7, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 8; or

(e) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 9, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 10;

and

(ii) a binding site for IL-17, comprising:

(a) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 13, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 14.

182. The method of claim 180, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising:

(1) SEQ ID NO: 1 and SEQ ID NO:2,

(2) SEQ ID NO: 3 and SEQ ID NO: 4,

(3) SEQ ID NO: 5 and SEQ ID NO: 6,

(4) SEQ ID NO: 7 and SEQ ID NO: 8, or

(5) SEQ ID NO: 9 and SEQ ID NO: 10;

and

(ii) variable domains that form a functional target binding site for IL-17 comprising:

SEQ ID NO: 13 and SEQ ID NO: 14.

183. The method of claim 180, wherein the DVD-lg comprises SEQ ID NO: 19 and SEQ ID NO: 20.

184. The method of claim 180, wherein the DVD-lg comprises (i) variable domains that form a functional target binding site for IL-1 β comprising CDRs 1-3 from SEQ ID NO: 3 and CDRs 1-3 from SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising CDRs 1-3 from SEQ ID NO: 13 and CDRs 1-3 from SEQ ID NO: 14.

185. The method of claim 180, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising SEQ ID NO: 3 and SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising SEQ ID NO: 13 and SEQ ID NO: 14.

186. The method of any one of claims 169-185, further comprising

administering an additional dose of the one or more therapeutic agents if the an elevated level of the one or more biomarkers is detected relative to the reference level.

187. The method of claim 186, further comprising administering a reduced dose of the one or more therapeutic agents or discontinuing treatment if a reduced level of the one or more biomarkers is detected relative to the reference level.

188. The method of claim 169, wherein the biological sample is a cell, tissue, fluid, organ, blood vessel, or bone.

189. The method of claim 188, wherein the cell is at least one of: an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a Plasma Cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a chondrocyte, a Kupffer cell, or a mastocyte.

190. The method of claim 188, wherein the biological sample is cartilage, bone, synovial membrane, synovial fluid, or another fluid within the joint cavity.

191. The method of claim 188, wherein the biological sample is whole blood;

192. The method of claim 191 , wherein the whole blood is a dried blood spot.

193. The method of claim 188, wherein the biological sample is serum.

194. The method of any one of claims 169-193, wherein the therapeutic agent is administered intravenously, intradermally, subcutaneously, or intramuscularly.

195. The method of any one of claims 169-194, wherein the biomarker comprises at least one of the following: aggrecan, TIMP-1 , TRAP-5b.

196. The method of any one of claims 169-195, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who do not have the inflammatory disorder.

197. The method of claim 196, wherein the reference level is the average level in samples from at least ten subjects who do not have the inflammatory disorder.

198. The method of claim 196 or 197, wherein the subjects who do not have the inflammatory disorder are healthy adults.

199. The method of any one of claims 169-198, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who have the inflammatory disorder.

200. The method of claim 199, wherein the reference level is the average level in samples from at least ten subjects who have the inflammatory disorder.

201. The method of claim 199 or 200, wherein the subjects who have the inflammatory disorder are healthy adults.

202. The method of one of claims 169-201 , wherein the therapeutic agent is not administered if the biomarker level is less than or equal to the reference level.

203. A method of screening a subject or population of subjects to identify a subject suitable for treatment of an inflammatory disorder by administering one or more therapeutic agents that antagonize IL-1 β and IL-17, the method comprising,

(a) collecting a biological sample from the subject,

(b) measuring the level of one or more biomarkers in the sample,

(c) comparing the levels of the biomarkers in the sample to reference levels, wherein the biomarkers are one or more of collagen (preferably collagen II), aggrecan, TIMP-1 , TRAP-5b, GM-CSF, CD4, CD40, ΜΙΡ-Ια, TNF (preferably TNFa), CD11b, IL-6, IL-10, IL-15, and IL-13Ra2, and

(d) identifying a subject suitable for treatment with one or more therapeutic agents that antagonize IL-1 β and IL-17 if the levels of the one or more biomarkers are elevated as compared to the reference level.

204. The method of claim 203, wherein the inflammatory disorder is

Alzheimer's disease, juvenile idiopathic arthritis, psoriasis, plaque psoriasis, keratoconjunctivitis sicca, blepharitis, keratitis, ankylosing spondylitis, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, asthma, atherosclerosis, Crohn's disease, colitis, inflammatory bowel disease (IBD), insulin dependent diabetes mellitus, chronic obstructive pulmonary disease (COPD), cystic fibrosis, urticaria, ectopic eczema, multiple sclerosis, dermatitis, diverticulitis, fibromyalgia, hepatitis, irritable bowel syndrome (IBS), systemic lupus erythematous (SLE), nephritis, Parkinson's disease, or ulcerative colitis.

205. The method of claim 204, wherein the inflammatory disorder is rheumatoid arthritis.

206. The method of any one of claims 203-205, wherein the determining of the level of one or more biomarkers in the biological sample from the subject comprises using a technique selected from the group consisting of

fluorescence-activated cell sorting (FACS), polymerase chain reaction (PCR), quantitative or real-time PCR (qPCR), gel electrophoresis, high-throughput sequencing, next generation sequencing, mass spectrometry, RNA sequencing, enzyme-linked immunosorbent spot (ELISpot), enzyme-linked immunosorbent assay (ELISA), microarray assay, quantitative biphasic calcium phosphate (BCP), Northern blot assay, Southern blot assay, Western blot assay, immunohistochemical assay, binding assay and combinations thereof.

207. The method of claim 206, wherein the technique is a flow cytometry cell surface marker assay.

208. The method of claim 206, wherein the technique is a multiplex cytokine or multiple chemokine assay.

209. The method of claim 206, wherein the technique is an ex vivo analysis of biomarkers following stimulation.

210. The method of claim 209, wherein the stimulation is by LPS, anti-CD3, anti-CD28 and combinations thereof.

21 1. The method of claim 203, wherein administering the dose of one or more therapeutic agents comprises simultaneously or sequentially administering an anti-IL-1 β antagonist and an anti-IL-17 antagonist.

212. The method of claim 211 , wherein the anti-IL-1 β antagonist is an anti-IL- 1β antibody and the anti-IL17 antagonist is an anti-IL-17 antibody.

213. The method of claim 203, wherein the therapeutic agent is a bispecific binding protein capable of binding IL-1 β and IL-17.

214. The method of claim 213, wherein the bispecific binding protein is a dual variable domain immunoglobulin (DVD-lg).

215. The method of claim 214, wherein the DVD-lg comprises

(i) a binding site for IL-Ι β, comprising:

( ) CDR H1 CDR H2 d CDR H3 f SEQ ID NO 1 d

and

(ii) a binding site for IL-17, comprising:

(a) CDR-H1 , CDR-H2, and CDR-H3 of SEQ ID NO: 13, and CDR-L1 , CDR-L2, and CDR-L3 of SEQ ID NO: 14.

216. The method of claim 214, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising:

and

(ii) variable domains that form a functional target binding site for IL-17 comprising:

SEQ ID NO: 13 and SEQ ID NO: 14.

217. The method of claim 214, wherein the DVD-lg comprises SEQ ID NO: 19 and SEQ ID NO: 20.

218. The method of claim 214, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising CDRs 1-3 from SEQ ID NO: 3 and CDRs 1-3 from SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising CDRs 1-3 from SEQ ID NO: 13 and CDRs 1-3 from SEQ ID NO: 14.

219. The method of claim 214, wherein the DVD-lg comprises

(i) variable domains that form a functional target binding site for IL-1 β comprising SEQ ID NO: 3 and SEQ ID NO: 4, and

(ii) variable domains that form a functional target binding site for IL-17 comprising SEQ ID NO: 13 and SEQ ID NO: 14.

220. The method of any one of claims 203-219, further comprising

administering an additional dose of the one or more therapeutic agents if the an elevated level of the one or more biomarkers is detected relative to the reference level.

221. The method of claim 220, further comprising administering a reduced dose of the one or more therapeutic agents or discontinuing treatment if a reduced level of the one or more biomarkers is detected relative to the reference level.

222. The method of claim 203, wherein the biological sample is a cell, tissue, fluid, organ, blood vessel, or bone.

223. The method of claim 222, wherein the cell is at least one of: an endothelial cell, a leukocyte, a macrophage, a CD4 T cell, a CD8 T cell, a B cell, a Plasma Cell, a neutrophil, a natural killer (NK) cell, a granulocyte, a monocyte, a chondrocyte, a Kupffer cell, or a mastocyte.

224. The method of claim 222, wherein the biological sample is cartilage, bone, synovial membrane, synovial fluid, or another fluid within the joint cavity.

225. The method of claim 222, wherein the biological sample is whole blood.

226. The method of claim 225, wherein the whole blood is a dried blood spot.

227. The method of claim 224, wherein the biological sample is serum.

228. The method of any one of claims 203-227, wherein the therapeutic agent is administered intravenously, intradermally, subcutaneously, or intramuscularly.

229. The method of any one of claims 203-228, wherein the biomarker comprises at least one of the following: aggrecan, TIMP-1 , TRAP-5b.

230. The method of any one of claims 203-229, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who do not have the inflammatory disorder.

231. The method of claim 230, wherein the reference level is the average level in samples from at least ten subjects who do not have the inflammatory disorder.

232. The method of claim 230 or 231 , wherein the subjects who do not have the inflammatory disorder are healthy adults.

233. The method of any one of claims 203-229, wherein the reference level comprises the level of the biomarker in a sample from one or more subjects who have the inflammatory disorder.

234. The method of claim 233, wherein the reference level is the average level in samples from at least ten subjects who have the inflammatory disorder.

235. The method of claim 233 or 234, wherein the subjects who have the inflammatory disorder are healthy adults.

236. The method of one of claims 203-235, wherein the therapeutic agent is not administered if the biomarker level is less than or equal to the reference level.