Il-23 and TNF-alpha inhibitors for use in autoimmune and inflammatory disorders

Abstract

Disclosed herein is a method of treating an individual afflicted with moderate to severe ulcerative colitis, the method comprising: administering a local inhibitor of interleukin-23 (IL-23) and tumor necrosis factor alpha (TNF-alpha) to an intestine of the individual.

Description

WSGR Docket No. 60790-720.601IL-23 AND TNF- ALPHA INHIBITORS FOR USE IN AUTOIMMUNE AND INFLAMMATORY DISORDERSCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U. S. Provisional Application No. 63 / 761,814 filed February 21, 2025, U. S. Provisional Application No. 63 / 779,779 filed March 28, 2025, and U. S. Provisional Application No. 63 / 794,246 filed April 24, 2025, the contents of which are incorporated by reference in their entirety.BACKGROUND

[0002] Autoimmune and inflammatory diseases afflict at least 1 in 15 in the United States. Autoimmune and inflammatory conditions arise through dysregulation of the immune system leading to sustained activation of pro-inflammatory pathways, such as constitutive expression of cytokines and chemokines by immune cells, that cause tissue damage. Various autoimmune and inflammatory diseases, such as inflammatory bowel disease (IBD) which includes Crohn’s disease and ulcerative colitis (UC), affect the gastrointestinal (GI) tract. Several autoimmune and inflammatory conditions of the GI tract, such as IBD and celiac disease, are presently incurable which leaves afflicted patients with life-long symptoms. Cytokines have been discovered to play a critical role in the pathogenesis of autoimmune diseases and inflammatory conditions of the GI tract. In particular, interleukin (IL)-23 and tumor necrosis factor alpha (TNF-alpha) have been implicated as key drivers of autoimmune and inflammatory pathologies.SUMMARY

[0003] The clinical research described herein has shown that IL-23 and / or TNF-alpha blockade can be effective in ameliorating autoimmune and inflammatory conditions in the GI tract. The clinical data described herein also suggest that the treatment of IL-23 and / or TNF-alpha blockade induces various clinical responses and outcomes including, but are not limited to, decreasing T cell activation, decreasing inflammatory signaling, increasing endocrine cell differentiation, and / or inducing histological improvement. The treatment of IL-23 and / or TNF-alpha blockade can surprisingly induce intestinal epithelial stem cell maintenance and / or proliferation, and induce mucosal regeneration.

[0004] Described herein in one aspect is a method of treating an individual afflicted withWSGR Docket No. 60790-720.601moderate to severe ulcerative colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the individual is afflicted with moderate ulcerative colitis. In certain embodiments, the individual is afflicted with severe ulcerative colitis. In certain embodiments, the treating achieves symptomatic remission. In certain embodiments, the treating achieves histologic remission. In certain embodiments, the treating achieves remission by a Modified Mayo score of a Full Mayo score.

[0005] Described herein in another aspect is a method of inducing intestinal stem cell maintenance and / or proliferation in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the method induces intestinal stem cell maintenance. In certain embodiments, the method induces intestinal stem cell proliferation.

[0006] Described herein in another aspect is a method of inducing a reduction in the number of B-cells in an intestinal tissue of an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

[0007] Described herein in another aspect is a method of inducing a reduction in the number of naive B-cells in the intestinal tissues of an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

[0008] Described herein in another aspect is a method of increasing Myc signaling in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

[0009] Described herein in another aspect is a method of increasing endocrine cell differentiation in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the method induces enteroendocrine cell differentiation.

[0010] Described herein in another aspect is a method of increasing hormone secretion in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. Described herein in another aspect is a method of altering plasma cell and misfolded protein response. Described herein in another aspect is a method of altering alpha beta T cell activation. Described herein in another aspect is a method of altering myeloid inflammatory signaling. Described herein in another aspect is a method of altering TNF signaling. Described herein in another aspect is a method of altering inflammatory response. Described herein in another aspect is a method of altering ribosome and rRNA processing. Described herein in another aspect is a method of altering mature epithelial cells.WSGR Docket No. 60790-720.601Described herein in another aspect is a method of altering immature epithelial cells.

[0011] Described herein in another aspect is a method of reducing expression of consensus gene sets representing TNF-alpha or IL -23 pathways in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

[0012] Described herein in another aspect is a method of reducing a Robarts Histopathology Index (RHI) in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the RHI is reduced by 6 or more compared to baseline. In certain embodiments, the RHI is reduced by 7 or more compared to baseline. In certain embodiments, the RHI is reduced by 8 or more compared to baseline. In certain embodiments, the baseline comprises the RHI of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha.

[0013] Described herein in another aspect is a method of reducing a Modified Mayo score in an individual afflicted with ulcerative colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the Modified Mayo score is reduced by 2 or more compared to baseline. In certain embodiments, the Modified Mayo score is reduced by 3 or more compared to baseline. In certain embodiments, the Modified Mayo score is reduced by 4 or more compared to baseline. In certain embodiments, the baseline comprises the Modified Mayo score of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha. In certain embodiments, the Modified Mayo score is reduced to 2 or less. In certain embodiments, the Modified Mayo score is reduced to 1 or less.

[0014] Described herein in another aspect is a method of reducing a Full Mayo score in an individual afflicted with ulcerative colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the Full Mayo score is reduced by 2 or more compared to baseline. In certain embodiments, the Full Mayo score is reduced by 3 or more compared to baseline. In certain embodiments, the Full Mayo score is reduced by 4 or more compared to baseline. In certain embodiments, the Full Mayo score is reduced by 5 or more compared to baseline. In certain embodiments, the baseline comprises the Modified Mayo score of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha. In certain embodiments, the Full Mayo score is reduced to 2 or less, and no individual sub score is greater than 1. In certain embodiments, the Full Mayo score is reduced to 1 or less.

[0015] Described herein in another aspect is a method of reducing a UC-100 score in an individual afflicted with ulcerative colitis, the method comprising: administering a local inhibitorWSGR Docket No. 60790-720.601of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the UC-100 score is reduced by 10 or more compared to baseline. In certain embodiments, the UC-100 score is reduced by 20 or more compared to baseline. In certain embodiments, the UC-100 score is reduced by 30 or more compared to baseline. In certain embodiments, the UC-100 score is reduced by 40 or more compared to baseline. In certain embodiments, the baseline comprises the UC-100 score of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha. In certain embodiments, the UC-100 score is reduced to 25 or less. In certain embodiments, the UC-100 score is reduced to 10 or less.

[0016] Described herein in another aspect is a method of inducing histological improvement in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the administering reduces neutrophils in the epithelium of the intestine of the individual to about less than 5%. In certain embodiments, the histological improvement is selected from the group consisting of: no crypt destruction in the intestine of the individual, no erosions in the intestine of the individual, no ulcerations in the intestine of the individual, and no granulation tissue in the intestine of the individual.

[0017] Described herein in another aspect is a method of inducing endoscopic response in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the method further comprises reducing a Mayo endoscopic score in the individual. In certain embodiments, the Mayo endoscopic score is reduced by 1 or more compared to baseline. In certain embodiments, the baseline comprises the Mayo endoscopic score of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha. In certain embodiments, the Mayo endoscopic score is 0 or 1.

[0018] Described herein in another aspect is a method of inducing mucosal regeneration in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

[0019] Described herein in another aspect is a method of inducing symptomatic remission in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. Described herein in another aspect is a method of inducing symptomatic improvement in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual. In certain embodiments, the method further comprises reducing a symptom stool frequency subscore (SFS) in the individual. In certain embodiments, the SFS score is reduced byWSGR Docket No. 60790-720.6011 or more compared to baseline. In certain embodiments, the SFS score is reduced by 2 or more compared to baseline. In certain embodiments, the SFS score is reduced by 3 or more compared to baseline. In certain embodiments, the method further comprises comprising reducing a rectal bleeding subscore (RBS) in the individual. In certain embodiments, the RBS score is reduced by 1 or more compared to baseline. In certain embodiments, the RBS score is reduced by 2 or more compared to baseline. In certain embodiments, the method results in a SFS=0 or 1 without worsening and a RBS=0.

[0020] In certain embodiments, the colitis is an inflammatory bowel disease. In certain embodiments, the inflammatory bowel disease is Crohn’s disease. In certain embodiments, the inflammatory bowel disease is ulcerative colitis. In certain embodiments, the ulcerative colitis is moderate to severe ulcerative colitis.

[0021] In certain embodiments, the local inhibitor is an oral inhibitor. In certain embodiments, the local inhibitor is orally dosed.

[0022] In certain embodiments, the individual is refractory to a previous treatment for colitis. In certain embodiments, in a serum cytokine level of the individual is not altered by administering the local inhibitor of IL-23 and TNF-alpha to the individual. In certain embodiments, the cytokine is selected from the list consisting of: fFNy, IL-17A, IL-1β, IL-6, TNFa, IL-10 and combinations thereof. In certain embodiments, in a tissue cytokine level of the individual is altered by administering the local inhibitor of IL-23 and TNF-alpha to the individual. In certain embodiments, the tissue is selected from the group consisting of intestinal tissue, colonic tissue, rectal tissue, and esophageal tissue.

[0023] In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is dosed twice daily. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is dosed twice daily for I, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 weeks. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is dosed once daily. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is dosed once daily for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 weeks. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is dosed for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, II, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36,37,38,39,40, 41, 42, 43, 44, 45, 46, 47,48,49,50,51, or 52 weeks or more. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is dosed until the individual achieves a clinical outcome and / or endpoint described herein.

[0024] In certain embodiments, the local inhibitor of IL-23 and TNF-alpha comprises a multispecific polypeptide comprising a first antigen binding domain that binds to TNF-alpha and a second antigen binding domain that binds to IL-23. In certain embodiments, the first antigenWSGR Docket No. 60790-720.601binding domain that binds to TNF-alpha comprises a VHH. In certain embodiments, the second antigen binding domain that binds to IL-23 comprises a VHH. In certain embodiments, the first antigen binding domain that binds to TNF-alpha and the second antigen binding domain that binds to IL-23 are coupled by a linker. In certain embodiments, the linker is a protease cleavable linker. In certain embodiments, the linker is a trypsin cleavable linker. In certain embodiments, the linker is a polypeptide linker. In certain embodiments, the linker is a polypeptide linker comprising an amino acid sequence as set forth in SEQ ID NO: 302. In certain embodiments, the first antigen binding domain that binds to TNF-alpha comprises: (a) a complementarity determining region 1 (CDR1) as set forth in SEQ ID NOs: 101, 107, 108, or 119; (b) a complementarity determining region 2 (CDR2) as set forth in SEQ ID NOs: 102, 109 to 117, or 120; and / or (c) a complementarity determining region 3 (CDR3) as set forth in SEQ ID NOs: 103 to 106 or 121 to 124. In certain embodiments, the first antigen binding domain that binds to TNF-alpha comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 118 or 125. In certain embodiments, the second antigen binding domain that binds to IL-23 comprises: (a) a complementarity determining region 1 (CDR1) as set forth in SEQ ID NOs: 1, 4, or 7; (b) a complementarity determining region 2 (CDR2) as set forth in SEQ ID NOs: 2, 5, or 8; and / or (c) a complementarity determining region 3 (CDR3) as set forth in SEQ ID NOs: 3, 6, or 9. In certain embodiments, the second antigen binding domain that binds to IL-23 comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10 to 12. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 201. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 202.

[0025] In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 600 mg to about 3200 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 600 mg to about 1600 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1200 mg to about 3200 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1200 mg to about 1600 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1800 mg to about 2400 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2400 mg to about 3200 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total dailyWSGR Docket No. 60790-720.601dose of about 600 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 800 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1200 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1300 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1400 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 1300 mg to about 1400 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1360 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1600 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1800 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 2000 mg to about 2800 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 2000 mg to about 2800 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2040 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2720 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2400 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 3200 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered twice daily at a dose of about 600 mg to achieve a total daily dose of about 1200 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered twice daily at a dose of about 900 mg to achieve a total daily dose of about 1800 mg. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered twice daily at a dose of about 1200 mg to achieve a total daily dose of about 2400 mg.

[0026] In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered for 8, 12 or 16 weeks followed by a maintenance dose. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered for 12 weeks followed by a maintenance dose. In certain embodiments, the maintenance dose is from about 1300 to about 1400 milligrams. In certain embodiments, the maintenance dose is about 1360 milligrams. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered for another period of 12 weeks if the individual is a non-responder.WSGR Docket No. 60790-720.601BRIEF DESCRIPTION OF THE DRAWINGS

[0027] The novel features described herein are set forth with particularity in the appended claims. A better understanding of the features and advantages of the features described herein will be obtained by reference to the following detailed description that sets forth illustrative examples, in which the principles of the features described herein are utilized, and the accompanying drawings of which:

[0028] FIGURE 1 shows logarithm base 2 of the fold change (Log2FC) of module expression levels of gene set variation analysis (GVSA) of modules associated with aP T cells activation in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor administered once a day (QD), and TNF-alpha / IL-23 dual inhibitor administered twice a day (BID).

[0029] FIGURE 2 shows change in expression levels of GVSA of modules associated with aP T cells activation in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0030] FIGURE 3 shows Log2FC of module expression levels in GVSA of modules associated with myeloid inflammatory TNF signaling in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0031] FIGURE 4 shows change in expression levels of GVSA of modules associated with myeloid inflammatory TNF signaling in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0032] FIGURE 5 shows Log2FC of module expression levels in GVSA of modules associated with inflammatory response in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0033] FIGURE 6 shows change in expression levels of GVSA of modules associated with inflammatory response in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0034] FIGURE 7 shows Log2FC of module expression levels in GVSA of modulesWSGR Docket No. 60790-720.601associated with intestinal stem cell maintenance in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0035] FIGURE 8 shows change in expression levels of GVSA of modules associated with intestinal stem cell maintenance in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0036] FIGURE 9 shows Log2FC of module expression levels in GVSA of modules associated with Myc signaling in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0037] FIGURE 10 shows change in expression levels of GVSA of modules associated with Myc signaling in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0038] FIGURE 11 shows Log2FC of module expression levels in GVSA of modules associated with B cells in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0039] FIGURE 12 shows change in expression levels of GVSA of modules associated with B cells in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0040] FIGURE 13 shows Log2FC of module expression levels in GVSA of modules associated with plasma cell misfolded protein response in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0041] FIGURE 14 shows change in expression levels of GVSA of modules associated with plasma cell misfolded protein response in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0042] FIGURE 15 shows Log2FC of module expression levels in GVSA of modules associated with endocrine cell differentiation in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0043] FIGURE 16 shows change in expression levels of GVSA of modules associated withWSGR Docket No. 60790-720.601endocrine cell differentiation in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0044] FIGURE 17 shows Log2FC of module expression levels in GVSA of modules associated with ribosome and rRNA processing in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0045] FIGURE 18 shows change in expression levels of GVSA of modules associated with ribosome and rRNA processing in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0046] FIGURE 19 shows Log2FC of module expression levels in GVSA of modules associated with mature and immature epithelial cells in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0047] FIGURE 20 shows change in expression levels of GVSA of modules associated with mature and immature epithelial cells in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0048] FIGURE 21 shows Log2FC of module expression levels in GVSA of modules associated with hormone secretion in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0049] FIGURE 22 shows change in expression levels of GVSA of modules associated with hormone secretion in paired samples in Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0050] FIGURE 23 shows change in mean combined ulcerative colitis (UC) symptom stool frequency subscore (SFS) and rectal bleeding subscore (RBS) in a per protocol (completer) population between Visit 2 (Baseline) and Visit 8 (Week 6) of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0051] FIGURE 24 shows improvement of UC-100 Score in patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID in an intention to treat (ITT; enrolled but did not complete treatment) population and a per protocol (completer) population of patients.WSGR Docket No. 60790-720.601

[0052] FIGURE 25 shows end of treatment (EOT) effects across multiple endpoints in a per protocol (completer) population of patient groups receiving of TNF-alpha / IL-23 dual inhibitor BID, where grey denotes improvement and black denotes no change or worsening.

[0053] FIGURE 26 shows change in endoscopic response in a per protocol (completer) population between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0054] FIGURE 27 shows percentage number of patients with endoscopic improvement (e.g., Mayo endoscopic score of 0 or 1 at day 42) in an intention-to-treat (ITT; enrolled but did not complete treatment) population of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0055] FIGURE 28 shows percentage number of patients with endoscopic improvement (e.g., Mayo endoscopic score of 0 or 1 at day 42) in a completer (per protocol) population of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0056] FIGURE 29 shows change in Robarts Histopathology Index (RHI) histological improvement in a completer population between Visit 2 (Baseline) and Visit 8 of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0057] FIGURE 30 shows improvement of Mayo Score in patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID in an intention-to-treat (ITT; enrolled but did not complete treatment) population and a per protocol (completer) population of patients.

[0058] FIGURE 31 shows improvement of Modified Mayo Score in patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID in an intention-to-treat (ITT; enrolled but did not complete treatment) population and a per protocol (completer) population of patients.

[0059] FIGURE 32 shows percentage of responders with improved Mayo Score, Modified Mayo Score, or symptom remission in an intention-to-treat (ITT; enrolled but did not complete treatment) population of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0060] FIGURE 33 shows percentage of responders with improved Mayo Score, Modified Mayo Score, or symptom remission in a per protocol (completer) population of patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.WSGR Docket No. 60790-720.601

[0061] FIGURE 34 shows Log2FC of pro-inflammatory cytokine IL-lbeta between Visit 2 (Baseline) and Visit 8 in paired colonic tissue in patient groups receiving of placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0062] FIGURE 35 shows Log2FC of pro-inflammatory cytokine IL-6 between Visit 2 (Baseline) and Visit 8 in paired colonic tissue in patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0063] FIGURE 36 shows Log2FC of pro-inflammatory cytokine TNF-alpha between Visit 2 (Baseline) and Visit 8 in paired colonic tissue in patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0064] FIGURE 37 shows Log2FC of pro-inflammatory cytokine IFN-gamma between Visit 2 (Baseline) and Visit 8 in paired colonic tissue in patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0065] FIGURE 38 shows Log2FC of pro-inflammatory cytokine IL-17A between Visit 2 (Baseline) and Visit 8 in paired colonic tissue in patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0066] FIGURE 39 shows Log2FC of anti-inflammatory cytokine IL-10 between Visit 2 (Baseline) and Visit 8 in paired colonic tissue in patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0067] FIGURE 40 shows change of cytokine IL-lbeta in Visit 2 (Baseline) and Visit 8 (Week 6) in paired colonic tissue of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0068] FIGURE 41 shows change of cytokine IL-6 in Visit 2 (Baseline) and Visit 8 (Week 6) in paired colonic tissue of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0069] FIGURE 42 shows change of cytokine TNF-alpha in Visit 2 (Baseline) and Visit 8 (Week 6) in paired colonic tissue of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0070] FIGURE 43 shows change of cytokine IFN-gamma in Visit 2 (Baseline) and Visit 8 (Week 6) in paired colonic tissue of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0071] FIGURE 44 shows change of cytokine IL-17A in Visit 2 (Baseline) and Visit 8 (Week 6) in paired colonic tissue of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0072] FIGURE 45 shows change of cytokine IL-10 in Visit 2 (Baseline) and Visit 8 (WeekWSGR Docket No. 60790-720.6016) in paired colonic tissue of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0073] FIGURE 46 shows an exemplary schematic dosing regimen for TNF-alpha / IL-23 dual inhibitor.

[0074] FIGURE 47 shows change of C-reactive protein (CRP) concentration in Visit 2 (Baseline) and Visit 8 (Week 6) of intention to treat (ITT; enrolled but did not complete treatment) patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0075] FIGURE 48 shows change of C-reactive protein (CRP) concentration in Visit 2 (Baseline) and Visit 8 (Week 6) of completer (per protocol) patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0076] FIGURE 49 shows change of fecal calprotectin concentration in Visit 2 (Baseline) and Visit 8 (Week 6) of intention to treat (ITT; enrolled but did not complete treatment) patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0077] FIGURE 50 shows change of fecal calprotectin concentration in Visit 2 (Baseline) and Visit 8 (Week 6) of completer (per protocol) patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0078] FIGURE 51 shows histological improvement in completer (per protocol) patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0079] FIGURE 52 shows symptomatic remission of intention to treat (ITT; enrolled but did not complete treatment) patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0080] FIGURE 53 shows symptomatic remission of completer (per protocol) patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0081] FIGURE 54 shows the log2FC change from Baseline to Weeks 1, 2, 3, 4, and 6 of pro-inflammatory cytokine IL-lbeta in paired serum samples of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0082] FIGURE 55 shows the log2FC change from Baseline to Weeks 1, 2, 3, 4, and 6 of pro-inflammatory cytokine IL-6 in paired serum samples of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0083] FIGURE 56 shows the log2FC change from Baseline to Weeks 1, 2, 3, 4, and 6 of pro-inflammatory cytokine TNF-alpha in paired serum samples of patient groups receivingWSGR Docket No. 60790-720.601placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0084] FIGURE 57 shows the log2FC change from Baseline to Weeks 1, 2, 3, 4, and 6 of pro-inflammatory cytokine IFN-gamma in paired serum samples of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0085] FIGURE 58 shows the log2FC change from Baseline to Weeks 1, 2, 3, 4, and 6 of pro-inflammatory cytokine IL-17A in paired serum samples of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0086] FIGURE 59 shows the log2FC change from Baseline to Weeks 1, 2, 3, 4, and 6 of pro-inflammatory cytokine IL-22 in paired serum samples of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0087] FIGURE 60 shows the log2FC change from Baseline to Weeks 1, 2, 3, 4, and 6 of anti-inflammatory cytokine IL- 10 in paired serum samples of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0088] FIGURE 61 shows Part 3 clinical remission rates in patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID in an intention to treat (ITT; includes all patients enrolled, including those that did not complete treatment) population and a per protocol (completer) population of patients. Clinical remission rates were determined based on the Mayo Clinic Score (MCS) and the modified MCS, where both MCS and modified MCS clinical remission rates were the same.

[0089] FIGURE 62 shows Part 3 pharmacokinetics of TNF-alpha / IL-23 dual inhibitor and its corresponding liberated anti-TNF-alpha and anti-IL-23 monomer arms in fecal samples collected from per protocol (completer) patient groups receiving TNF-alpha / IL-23 dual inhibitor QD.

[0090] FIGURE 63 shows Part 3 pharmacokinetics of TNF-alpha / IL-23 dual inhibitor and its corresponding liberated anti-TNF-alpha and anti-IL-23 monomer arms in fecal samples collected from per protocol (completer) patient groups receiving TNF-alpha / IL-23 dual inhibitor BID.

[0091] FIGURE 64 shows Part 3 colonic tissue concentrations of TNF-alpha / IL-23 dual inhibitor and its corresponding liberated anti-TNF-alpha and anti-IL-23 monomer arms using ELISA-based assays. Lines denote median values of positive samples. Open symbols correspond to patient A. Left-filled symbols correspond to patient B.

[0092] FIGURE 65 shows Part 3 colonic tissue concentrations of TNF-alpha / IL-23 dual inhibitor and its corresponding liberated anti-TNF-alpha and anti-IL-23 monomer arms using Mass Spectrometry (MS). Open symbols indicate not detected. Peptides sequences detectedWSGR Docket No. 60790-720.601shown in square brackets.

[0093] FIGURE 66 shows Part 3 colonic tissue levels of liberated anti-TNF-alpha and anti-IL-23 monomer arms using ELISA-based assays and Mass Spectrometry. Individual patients are represented by unique shapes; closed shading = QD, open shading = BID.

[0094] FIGURE 67 shows Part 3 Log2FC gene expression of inflammatory tissue cytokines IL-1B, IL-6, and TNF between Baseline and Week 6 in paired colonic tissue in patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0095] FIGURE 68 shows Part 3 Log2FC gene expression of T-cell associated tissue cytokines IL12B, IL23A, IL17A, IL22, IFNG, and IL10 between Baseline and Week 6 in paired colonic tissue in patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID.

[0096] FIGURE 69 shows logarithm base 2 of the fold change (Log2FC) of consensus gene sets representing TNF-alpha (2 gene sets: “Hallmark_TNFA_ Signaling_Via_NFKB”, “REACTOME_TNF_ SIGNALING”) or IL-23 (1 gene set: “PID_IL23_Pathway”) between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor administered once a day (QD), and TNF-alpha / IL-23 dual inhibitor administered twice a day (BID).

[0097] FIGURE 70 shows an exemplary schematic dosing regimen and for TNF-alpha / IL-23 dual inhibitor.DETAILED DESCRIPTIONCertain Definitions

[0098] In the following description, certain specific details are set forth in order to provide a thorough understanding of various embodiments. However, one skilled in the art will understand that the embodiments provided may be practiced without these details. Unless the context requires otherwise, throughout the specification and claims which follow, the word “comprise” and variations thereof, such as, “comprises” and “comprising” are to be construed in an open, inclusive sense, that is, as “including, but not limited to.” As used in this specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the content clearly dictates otherwise. It should also be noted that the term “or” is generally employed in its sense including “and / or” unless the content clearly dictates otherwise. Further, headings provided herein are for convenience only and do not interpret the scope or meaning of the claimed embodiments.WSGR Docket No. 60790-720.601

[0099] As used herein the term “about” refers to an amount that is near the stated amount by 10% or less.

[0100] As used herein the term “individual,” “patient,” or “subject” refers to individuals diagnosed with, suspected of being afflicted with, or at-risk of developing at least one disease for which the described compositions and method are useful for treating. In certain embodiments the individual is a mammal. In certain embodiments, the mammal is a mouse, rat, rabbit, dog, cat, horse, cow, sheep, pig, goat, llama, alpaca, or yak. In certain embodiments, the individual is a human.

[0101] “ Treat,” “treatment,” or “treating,” as used herein refers to, e.g., a deliberate intervention to a physiological disease state resulting in the reduction in severity of a disease or condition; the reduction in the duration of a condition; the amelioration or elimination of one or more symptoms associated with a disease or condition; the maintenance of a therapeutic benefit, or the provision of beneficial effects to a subject with a disease or condition. Treatment does not require curing the underlying disease or condition.

[0102] A “therapeutically effective amount”, “effective dose”, “effective amount”, or “therapeutically effective dosage” of a drug or therapeutic agent is any amount of the drug that, when used alone or in combination with another therapeutic agent, protects a subject against the onset of a disease or promotes disease regression evidenced by a decrease in severity of disease symptoms, an increase in frequency and duration of disease symptom-free periods, or a prevention of impairment or disability due to the disease affliction. The ability of a therapeutic agent to promote disease regression can be evaluated using a variety of methods known to the skilled practitioner, such as in human subjects during clinical trials, in animal model systems predictive of efficacy in humans, or by assaying the activity of the agent in in vitro assays.

[0103] The terms “polypeptide” and “protein” are used interchangeably to refer to a polymer of amino acid residues, and are not limited to a minimum length. Polypeptides, including the provided antibodies and antibody chains and other peptides, e.g., linkers and binding peptides, may include amino acid residues including natural and / or non-natural amino acid residues. The terms also include post-expression modifications of the polypeptide, for example, glycosylation, sialylation, acetylation, phosphorylation, and the like. In some aspects, the polypeptides may contain modifications with respect to a native or natural sequence, as long as the protein maintains the desired activity.

[0104] The term “refractory” as used herein refers to individuals that do not exhibit improvement in symptoms, Mayo score or improved Mayo Score after a course of treatment with a previous therapeutic intervention. Those who discontinue treatment due to side effects or onlyWSGR Docket No. 60790-720.601show minimal improvement less than what would be considered a clinical response (e.g., improvement in Mayo score of less than 30% from baseline) can also be classified as refractory.

[0105] The term “antibody” herein is used in the broadest sense and includes monoclonal antibodies, bispecific antibodies, and includes intact antibodies and functional (antigen-binding) antibody fragments thereof, including fragment antigen binding (Fab) fragments, F(ab')2 fragments, Fab' fragments, Fv fragments, recombinant IgG (rlgG) fragments, single chain antibody fragments, including single chain variable fragments (sFv or scFv), VHHs, VNARs, vorabodies, immunoglobulin chain variable domains, and single domain antibodies (e.g., sdAb, sdFv, nanobody) fragments. The term encompasses genetically engineered and / or otherwise modified forms of immunoglobulins, such as intrabodies, peptibodies, chimeric antibodies, fully human antibodies, humanized antibodies, and heteroconjugate antibodies, multi specific, e.g., bispecific, antibodies, diabodies, triabodies, and tetrabodies, tandem di-scFv, tandem tri-scFv. Unless otherwise stated, the term “antibody” should be understood to encompass functional antibody fragments thereof. The term also encompasses intact or full- length antibodies, including antibodies of any class or sub-class, including IgG and sub-classes thereof, IgM, IgE, IgA, and IgD. The antibody can comprise a human IgGl constant region. The antibody can comprise a human IgG4 constant region.

[0106] The term “small molecule” typically refers to organic, inorganic or organometallic compounds having a molecular weight of less than about 2000 Daltons.

[0107] As used herein, “pharmaceutically acceptable” with reference to a carrier” “excipient” or “diluent” includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible. In some aspects, the carrier is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, epidermal, oral, rectal, or intranasal administration (e.g., by injection, infusion, ingestion, insertion, or inhalation). Depending on the route of administration, the active compound, i.e., antibody, can be coated in a material to protect the compound from the action of acids and other natural conditions that can inactivate the compound.

[0108] The pharmaceutical compounds described herein can include one or more pharmaceutically acceptable salts. A “pharmaceutically acceptable salt” refers to a salt that retains the desired biological activity of the parent compound and does not impart any undesired toxicological effects (see e.g., Berge, S. M., et al. (1977) J. Pharm. Sci. 66: 1-19). Examples of such salts include acid addition salts and base addition salts. Acid addition salts include those derived from nontoxic inorganic acids, such as hydrochloric, nitric, phosphoric, sulfuric, hydrobromic, hydroiodic, phosphorous and the like, as well as from nontoxic organic acids suchWSGR Docket No. 60790-720.601as aliphatic mono- and di carboxylic acids, phenyl- substituted alkanoic acids, hydroxy alkanoic acids, aromatic acids, aliphatic and aromatic sulfonic acids and the like. Base addition salts include those derived from alkaline earth metals, such as sodium, potassium, magnesium, calcium and the like, as well as from nontoxic organic amines, such as N, N'-dibenzylethylenediamine, N-methylglucamine, chloroprocaine, choline, diethanolamine, ethylenediamine, procaine and the like.

[0109] Percent (%) sequence identity with respect to a reference polypeptide sequence is the percentage of amino acid residues in a candidate sequence that are identical with the amino acid residues in the reference polypeptide sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be achieved in various ways that are known for instance, using publicly available computer software such as BLAST, BLAST-2, ALIGN or Megalign (DNASTAR) software. Appropriate parameters for aligning sequences are able to be determined, including algorithms needed to achieve maximal alignment over the full length of the sequences being compared. For purposes herein, however, % amino acid sequence identity values are generated using the sequence comparison computer program ALIGN-2. The ALIGN-2 sequence comparison computer program was authored by Genentech, Inc., and the source code has been filed with user documentation in the U. S. Copyright Office, Washington D. C., 20559, where it is registered under U. S. Copyright Registration No. TXU510087. The ALIGN-2 program is publicly available from Genentech, Inc., South San Francisco, Calif., or may be compiled from the source code. The ALIGN-2 program should be compiled for use on a UNIX operating system, including digital UNIX V4.0D. All sequence comparison parameters are set by the ALIGN-2 program and do not vary.

[0110] For the purposes of comparing two closely-related polypeptide sequences, the “% sequence identity" between a first polypeptide sequence and a second polypeptide sequence may be calculated using NCBI BLAST v2.0, using standard settings for polypeptide sequences (BLASTP). For the purposes of comparing two closely-related polynucleotide sequences, the “% sequence identity" between a first nucleotide sequence and a second nucleotide sequence may be calculated using NCBI BLAST v2.0, using standard settings for nucleotide sequences (BLASTN). The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses as defaults a wordlength (W) of 11, an expectation (E) or 10, M=5, N=-4 and a comparison of both strands. For amino acid sequences, the BLASTP program uses as defaults a wordlength of 3, andWSGR Docket No. 60790-720.601expectation (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff & Henikoff, Proc. Natl. Acad. Sci. USA 89:10915 (1989)) alignments (B) of 50, expectation (E) of 10, M=5, N=-4, and a comparison of both strands. The BLAST algorithm also performs a statistical analysis of the similarity between two sequences (see, e.g., Karlin & Altschul, Proc. Nat’l. Acad. Sci. USA 90:5873-5787 (1993)). One measure of similarity provided by the BLAST algorithm is the smallest sum probability (P(N)), which provides an indication of the probability by which a match between two nucleotide or amino acid sequences would occur by chance. For example, a nucleic acid is considered similar to a reference sequence if the smallest sum probability in a comparison of the test nucleic acid to the reference nucleic acid is less than about 0.2, more suitably less than about 0.01, and most suitably less than about 0.001.

[0111] Polypeptide or polynucleotide sequences are said to be the same as or identical to other polypeptide or polynucleotide sequences, if they share 100% sequence identity over their entire length. Residues in sequences are numbered from left to right, i.e. from N- to C- terminus for polypeptides; from 5’ to 3’ terminus for polynucleotides.

[0112] A “difference” between sequences refers to an insertion, deletion or substitution of a single amino acid residue in a position of the second sequence, compared to the first sequence. Two polypeptide sequences can contain one, two or more such amino acid differences.Insertions, deletions or substitutions in a second sequence which is otherwise identical (100% sequence identity) to a first sequence result in reduced % sequence identity. For example, if the identical sequences are 9 amino acid residues long, one substitution in the second sequence results in a sequence identity of 88.9%. If the identical sequences are 17 amino acid residues long, two substitutions in the second sequence results in a sequence identity of 88.2%. If the identical sequences are 7 amino acid residues long, three substitutions in the second sequence results in a sequence identity of 57.1%. If first and second polypeptide sequences are 9 amino acid residues long and share 6 identical residues, the first and second polypeptide sequences share greater than 66% identity (the first and second polypeptide sequences share 66.7% identity). If first and second polypeptide sequences are 17 amino acid residues long and share 16 identical residues, the first and second polypeptide sequences share greater than 94% identity (the first and second polypeptide sequences share 94.1% identity). If first and second polypeptide sequences are 7 amino acid residues long and share 3 identical residues, the first and second polypeptide sequences share greater than 42% identity (the first and second polypeptide sequences share 42.9% identity).

[0113] Alternatively, for the purposes of comparing a first, reference polypeptide sequence to a second, comparison polypeptide sequence, the number of additions, substitutions and / orWSGR Docket No. 60790-720.601deletions made to the first sequence to produce the second sequence may be ascertained. An addition is the addition of one amino acid residue into the sequence of the first polypeptide (including addition at either terminus of the first polypeptide). A substitution is the substitution of one amino acid residue in the sequence of the first polypeptide with one different amino acid residue. A deletion is the deletion of one amino acid residue from the sequence of the first polypeptide (including deletion at either terminus of the first polypeptide).

[0114] For the purposes of comparing a first, reference polynucleotide sequence to a second, comparison polynucleotide sequence, the number of additions, substitutions and / or deletions made to the first sequence to produce the second sequence may be ascertained. An addition is the addition of one nucleotide residue into the sequence of the first polynucleotide (including addition at either terminus of the first polynucleotide). A substitution is the substitution of one nucleotide residue in the sequence of the first polynucleotide with one different nucleotide residue. A deletion is the deletion of one nucleotide residue from the sequence of the first polynucleotide (including deletion at either terminus of the first polynucleotide).

[0115] Numerical identifiers with respect to polypeptides or binding regions used herein, such as “first” or “second”, do not imply structural order. For instance, a fusion protein described as comprising a first domain and a second domain does not imply that the first domain is at an N-terminal position relative to the second domain.Antibodies and antibody-derived binding moieties

[0116] The antibodies or antibody-derived binding moieties described herein can be encoded by a nucleic acid. A nucleic acid is a type of polynucleotide comprising two or more nucleotide bases. In certain embodiments, the nucleic acid is a component of a vector that can be used to transfer the polypeptide encoding polynucleotide into a cell. As used herein, the term “vector” refers to a nucleic acid molecule capable of transporting another nucleic acid to which it has been linked. Immunoglobulin chain variable domains of the invention may, for example, be obtained by preparing a nucleic acid encoding an immunoglobulin chain variable domain using techniques for nucleic acid synthesis, followed by expression of the nucleic acid thus obtained.

[0117] Among the provided antibodies are antibody fragments. An “antibody fragment” refers to a molecule other than an intact antibody that comprises a portion of an intact antibody that binds the antigen to which the intact antibody binds. Examples of antibody fragments include, but are not limited to, Fv, Fab, Fab’, Fab’-SH, F(ab’)2; diabodies; linear antibodies; single-chain antibody molecules (e.g. scFv or sFv); and multispecific antibodies formed from antibody fragments. In particular embodiments, the antibodies are single-chain antibodyWSGR Docket No. 60790-720.601fragments comprising a variable heavy chain region and / or a variable light chain region, such as scFvs.

[0118] Herein a molecule, peptide, polypeptide, antibody, or antibody fragment can be referred to as “bispecific” or “dual-specific” including grammatical equivalents. A bispecific molecule possesses the ability to specifically bind to at least two structurally distinct targets. The specific binding may be the result of two distinct binding moieties that are structurally distinct at the molecular level, including but not limited to distinct non-identical amino acid sequences; or a single binding moiety that is able to specifically bind to two structurally distinct targets with high affinity (e.g., with a dissociation constant (KD) less than about IxlO'6). A molecule, peptide, polypeptide, antibody, or antibody fragment referred to as “multi-specific” refers to a molecule that possesses the ability to specifically bind to at least three structurally distinct targets. A “bispecific antibody” including grammatical equivalents refers to a bispecific molecule that preserves at least one fragment of an antibody able to specifically bind a target, for example, a variable region, heavy or light chain, or one or more complementarity determining regions from an antibody molecule. A “multi-specific antibody” including grammatical equivalents refers to a multi-specific molecule that preserves at least one fragment of an antibody able to specifically bind with a target, for example, a variable region, heavy or light chain, or complementarity determining region from an antibody molecule.

[0119] A “linker” herein is also referred to as “linker sequence” “spacer” “tethering sequence” or grammatical equivalents thereof. A “linker” as referred herein connects two distinct molecules that by themselves possess target binding, catalytic activity, or are naturally expressed and assembled as separate polypeptides, or comprise separate domains of the same polypeptide. For example, two distinct binding moieties or a heavy-chain / light-chain pair. A number of strategies may be used to covalently link molecules together. Linkers described herein may be utilized to join a light chain variable region and a heavy chain variable region in an scFv molecule; or may be used to tether an scFv or other antigen binding fragment on the N- or C-terminus of an antibody heavy chain; or the N- or C- terminus of a light chain to create a bispecific or multispecific binding molecule. These include but are not limited to polypeptide linkages between N- and C-termini of proteins or protein domains, linkage via disulfide bonds, and linkage via chemical cross-linking reagents. In one aspect of this embodiment, the linker is a peptide bond, generated by recombinant techniques or peptide synthesis. The linker peptide may predominantly include the following amino acid residues: Gly, Ser, Ala, or Thr. The linker peptide should have a length that is adequate to link two molecules in such a way that they assume the correct conformation relative to one another so that they retain the desired activity. InWSGR Docket No. 60790-720.601one embodiment, the linker is from about 1 to 50 amino acids in length, or about 1 to 30 amino acids in length. In one embodiment, linkers of 1 to 20 amino acids in length may be used. Useful linkers include glycine-serine polymers, including for example (GS)n, (GSGGS)n, (GGGGS)n, and (GGGS)n, where n is an integer of at least one, glycine-alanine polymers, alanine-serine polymers, and other flexible linkers. Exemplary, linkers for linking antibody fragments, single chain variable fragments, or VHHs can include AAEPKSS, AAEPKSSDKTHTCPPCP, GGGG, or GGGGDKTHTCPPCP. Alternatively, a variety of non-proteinaceous polymers, including but not limited to polyethylene glycol (PEG), polypropylene glycol, polyoxyalkylenes, or copolymers of polyethylene glycol and polypropylene glycol, may find use as linkers. In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 301 (GGGGSKGGGGS). In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 302 (GGGGSGGGGSKGGGGSGGGGS).

[0120] The terms “complementarity determining region,” and “CDR,” which are synonymous with “hypervariable region” or “HVR,” are known in the art to refer to noncontiguous sequences of amino acids within antibody variable regions, which confer antigen specificity and / or binding affinity. In general, there are three CDRs in each heavy chain variable region (CDR-H1, CDR-H2, CDR-H3) and three CDRs in each light chain variable region (CDR-Ll, CDR-L2, CDR-L3). “Framework regions” and “FR” are known in the art to refer to the non-CDR portions of the variable regions of the heavy and light chains. In general, there are four FRs in each full-length heavy chain variable region (FR-H1, FR-H2, FR-H3, and FR-H4), and four FRs in each full-length light chain variable region (FR-L1, FR-L2, FR-L3, and FR-L4). The precise amino acid sequence boundaries of a given CDR or FR can be readily determined using any of a number of well-known schemes, including those described by Kabat et al. (1991), “Sequences of Proteins of Immunological Interest,” 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD (“Kabat” numbering scheme), Al-Lazikani et al., (1997) JMB 273,927-948 (“Chothia” numbering scheme); MacCallum et al., J. Mol. Biol. 262:732-745 (1996), “Antibody-antigen interactions: Contact analysis and binding site topography,” J. Mol. Biol. 262, 732-745.” (“Contact” numbering scheme); Lefranc MP et al., “IMGT unique numbering for immunoglobulin and T cell receptor variable domains and Ig superfamily V-like domains,” Dev Comp Immunol, 2003 Jan;27(l):55-77 (“IMGT” numbering scheme); Honegger A and Pliickthun A, “Yet another numbering scheme for immunoglobulin variable domains: an automatic modeling and analysis tool,” J Mol Biol, 2001 Jun 8;309(3):657-70, (“Aho” numbering scheme); and Whitelegg NR and Rees AR, “WAM: an improved algorithm for modelling antibodies on the WEB,” Protein Eng. 2000 Dec;13(12):819-24 (“AbM” numbering scheme. InWSGR Docket No. 60790-720.601certain embodiments, the CDRs of the antibodies described herein can be defined by a method selected from Kabat, Chothia, IMGT, Aho, AbM, or combinations thereof.

[0121] The boundaries of a given CDR or FR may vary depending on the scheme used for identification. For example, the Kabat scheme is based on structural alignments, while the Chothia scheme is based on structural information. Numbering for both the Kabat and Chothia schemes is based upon the most common antibody region sequence lengths, with insertions accommodated by insertion letters, for example, “30a,” and deletions appearing in some antibodies. The two schemes place certain insertions and deletions (“indels”) at different positions, resulting in differential numbering. The Contact scheme is based on analysis of complex crystal structures and is similar in many respects to the Chothia numbering scheme.

[0122] Specific binding or binding of antibody molecules described herein refers to binding mediated by one or more CDR portions of the antibody. Not all CDRs may be required for specific binding. Specific binding can be demonstrated for example by an ELISA against a specific recited target or antigen that shows significant increase in binding compared to an isotype control antibody.

[0123] The term “variable region” or “variable domain” refers to the domain of an antibody heavy or light chain that is involved in binding the antibody to antigen. The variable domains of the heavy chain and light chain (VH and VL, respectively) of a native antibody generally have similar structures, with each domain comprising four conserved framework regions (FRs) and three CDRs (See e.g, Kindt et al. Kuby Immunology, 6th ed., W. H. Freeman and Co., page 91(2007)). A single VH or VL domain may be sufficient to confer antigen-binding specificity. Furthermore, antibodies that bind a particular antigen may be isolated using a VH or VL domain from an antibody that binds the antigen to screen a library of complementary VL or VH domains, respectively (See e.g., Portolano et al., J. Immunol. 150:880-887 (1993); Clarkson et al., Nature 352:624-628 (1991)).

[0124] Substituting at least one amino acid residue in the framework region of a non-human immunoglobulin variable domain with the corresponding residue from a human variable domain is humanization. Humanization of a variable domain may reduce immunogenicity in humans.

[0125] An exception to conventional antibody structure can found in sera of camelids and cartilaginous fish (e.g., sharks). In addition to conventional antibodies, these sera possess special IgG antibodies. These IgG antibodies, known as heavy-chain antibodies (HCAbs), are devoid of the L chain polypeptide and lack the first constant domain (CHI). At its N-terminal region, the H chain of the homodimeric protein contains a dedicated immunoglobulin chain variable domain, referred to as the VHH for camelids and vNAR for sharks, which serves to associate with itsWSGR Docket No. 60790-720.601cognate antigen (Muyldermans 2013 Annu Rev Biochem 82:775-797, Hamers-Casterman et al.1993 Nature 363(6428):446-448, Muyldermans et al. 1994 Protein Eng 7(9): 1129-1135, Konning et al. 2017 Curr Opin Struct Biol 45:10-16 herein incorporated by reference in their entirety).

[0126] VHHs are single-domain antibodies that comprise a single heavy chain variable domain. Similar to the variable domains from conventional antibodies, VHHs comprise four FRs and three CDRs. Unlike conventional antibodies, VHHs do not have nor require a light chain variable domain to pair with its heavy chain variable domain for antigen binding to occur. In some embodiments, the VHHs described herein are stable throughout recombinant production and in vivo delivery. In some embodiments, the VHHs described herein are resistant to cleavage by recombinant host organism (e.g., yeast) proteases and by proteases present in the GI tract. In some embodiments, the total number of amino acid residues in a VHH or VH may be in the region of 110-130.

[0127] VHHs can be used to construct multivalent and / or multispecific polypeptides. In some embodiments, a polypeptide described herein can comprise at least two identical VHHs. In some embodiments, a polypeptide described herein can comprise at least two distinct VHHs. A multivalent polypeptide comprises two or more binding polypeptides (e.g., VHHs) which presents two or more sites at which binding to one or more antigens can occur. A multispecific polypeptide comprises two or more distinct binding polypeptides (e.g., VHHs) which present two or more sites at which either (a) binding to two or more distinct antigens can occur or (b) binding to two or more distinct epitopes on the same antigen can occur.

[0128] Multivalent and / or multispecific polypeptides described herein can incorporate linkers to covalently join functional domains (e.g., VHHs) together to act as one molecule throughout the recombinant production process and / or in vivo delivery. In some embodiments, the linkers described herein can be stable and resistant to cleavage by recombinant host organism (e.g., yeast) proteases as well as proteases present in the GI tract. In some embodiments, the linkers described herein can be labile. In some embodiments, the labile peptide linker can be engineered such that it resists cleavage by proteases to a desired extent and / or is only cleaved upon exposure to a specific area of the intestinal tract. This can be achieved according to one embodiment of the invention by incorporating shielding residues into the labile peptide linker flanking the labile site(s). Shielding residues flank the labile site(s) of the labile peptide linker and reduce the lability thereof. Cleavage resistance can also be increased by positioning the labile site(s) closer to or at the periphery of the labile peptide linker. This concept is referred to as a “shielded labile site” and provides controlled lability. In a further embodiment of the invention, the labile peptideWSGR Docket No. 60790-720.601linker can be engineered such that it is highly labile to cleavage by intestinal tract proteases, thereby quickly releasing the constituent first and second polypeptides of the construct after oral administration. This is achieved by incorporating one or more labile sites into the labile peptide linker such that the labile site is exposed for proteolysis, for example by positioning the labile site(s) substantially centrally in the labile peptide linker and / or by the labile site not being shielded substantially by flanking residues. This concept is referred to as a “non-shielded labile site”. In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 301 (GGGGSKGGGGS). In certain embodiments, the linker comprises or consists of the amino acid sequence set forth in SEQ ID NO: 302 (GGGGSGGGGSKGGGGSGGGGS).

[0129] As used herein a “Full Mayo Clinical Score” or “Full Mayo” scores consists of a rectal bleeding sub score (RBS) from 0 to 3, stool frequency (SF) from 0 to 3, physician’s global assessment from 0 to 3, and an endoscopy sub score from 0 to 3 based on central review, score ranges from 0 to 12, with a higher score indicating more severe disease. See e.g., Schroeder KW, Tremaine WJ, Ilstrup DM. “Coated oral 5-aminosalicylic acid therapy for mildly to moderately active ulcerative colitis. Arandomized study.” N Engl J Med. 1987;317:1625-1629.

[0130] As used herein a “Modified Mayo Clinical Score” or “Modified Mayo” scores consists of a Rectal bleeding sub score (RBS) from 0 to 3, stool frequency (SF) from 0 to 3 and an endoscopy sub score from 0 to 3 based on central review, score ranges from 0 to 9, with a higher score indicating more severe disease. See e.g, Scherl EJ, Pruitt R, Gordon GL, et al. “Safety and efficacy of a new 3.3 g b.i.d. tablet formulation in patients with mild-to-moderately-active ulcerative colitis: a multi center, randomized, double-blind, placebo-controlled study.” Am J Gastroenterol. 2009; 104:1452-1459.

[0131] As used herein a “clinical response” refers to, with respect to the Full Mayo Clinical Score, a decrease from baseline in the full MCS >30% and >3 points with either a decrease in RBS >1 or RBS of 0 or 1. With respect to the modified Mayo Score, a decrease from study baseline modified Mayo score of >2 points and >30%, plus a decrease in rectal bleeding subscore of >1 point or an absolute rectal bleeding subscore of 0 or 1. Responders are those patients or individuals that exhibit a clinical response.

[0132] As used herein “clinical remission” or “remission” refers to, with respect to a full Mayo clinical score, a score of less than or equal to 2 with no individual sub score greater than 1. With respect to a modified Mayo clinical score, a score of 0 on the rectal bleeding sub score, a stool frequency sub score of 0 or 1 with no increase from baseline, a endoscopy sub score of 0 or 1, and symptomatic remission.WSGR Docket No. 60790-720.601

[0133] “Symptomatic remission” refers to Stool Frequency score of 0 or 1 with no increase from baseline, and a rectal bleeding score of 0.

[0134] “Histologic Remission” as used herein refers to absence of neutrophils from the mucosa (both lamina propria and epithelium), no crypt destruction, and no erosions, ulcerations, or granulation tissue according to the Geboes grading system. See e.g., Geboes K, Riddell R, Ost A, et al. “A reproducible grading scale for histological assessment of inflammation in ulcerative colitis. Gut 2000;47:404-9.

[0135] “Endoscopic Improvement” as described herein refers to a Mayo endoscopy sub score of 0 or 1 with no friability.

[0136] “Endoscopic Normalization” as described herein refers to a Mayo endoscopy sub score of 0.

[0137] “Moderate Ulcerative Colitis” as used herein refers to a full mayo score of 6, 7, 8, 9, or 10.

[0138] “ Severe Ulcerative Colitis” as used herein refers to a full mayo score of 11, or 12.Specificity., affinity, avidity and cross-reactivity

[0139] Specificity refers to the number of different types of antigens or antigenic determinants to which a particular antigen-binding polypeptide can bind. The specificity of an antigen-binding polypeptide is the ability of the antigen-binding polypeptide to recognize a particular antigen as a unique molecular entity and distinguish it from another.

[0140] Affinity, represented by the equilibrium constant for the dissociation of an antigen with an antigen-binding polypeptide (KD), is a measure of the binding strength between an antigenic determinant and an antigen-binding site on the antigen-binding polypeptide: the lesser the value of the KD, the stronger the binding strength between an antigenic determinant and the antigen-binding polypeptide (alternatively, the affinity can also be expressed as the affinity constant (Ka), which is 1 / KD). Affinity can be determined by known methods, depending on the specific antigen of interest. Suitably, affinity is determined using a dynamically switchable biosurface (e.g. “switch SENSE®”, see Knezevic et al. 2012 Journal of the American Chemical Society 134(37):15225-15228) or by surface plasmon resonance.

[0141] Avidity is the measure of the strength of binding between an antigen-binding polypeptide and the pertinent antigen. Avidity is related to both the affinity between an antigenic determinant and its antigen binding site on the antigen-binding polypeptide and the number of pertinent binding sites present on the antigen-binding polypeptide.

[0142] In some embodiments, an antibody or antibody-derived binding moiety provided hereinWSGR Docket No. 60790-720.601has a KDof about 1 pM, 100 nM, 50 nM, 40 nM, 30 nM, 20 nM, 10 nM, 5 nM, 2 nM, 1 nM, 0.5 nM, 0.1 nM, 0.05 nM, 0.01 nM, or 1 pM or less (e.g., 108M or less, e.g., from 108M to 1012M, e.g., from 109M to 1012M) for the antibody target. In some embodiments, an antibody or antibody-derived binding moiety provided herein has a KD of about 100 nM, 50 nM, 40 nM, 30 nM, 20 nM, 10 nM, 5 nM, 2 nM, 1 nM, 0.5 nM, 0.1 nM, 0.05 nM, 0.01 nM, or 1 pM or greater (e.g., 1012M or greater, e.g., from 108M to 1012M, e.g., from 109M to 1012M) for the antibody target. KD can be measured by any suitable assay. In certain embodiments, KD can be measured using surface plasmon resonance assays e.g., using a BIACORE®-2000, a BIACORE®-3000 or Octet).

[0143] The antibody or antibody-derived binding moiety can be an IL-23 or an IL-23 receptor targeting antibody or VHH. In some embodiments, the IL-23-specific polypeptide of the invention can bind to IL-23 with a KD of 106M or less, 10'7M or less, 10'8M or less, 10'9M or less, 10'10M or less, 10'11M or less, 10'12M or less, or 10'13M or less. In some embodiments, the polypeptide of the invention can bind to IL-23 with a KD of 106M or greater, 10'7M or greater, 10'8M or greater, 10'9M or greater, 10'10M or greater, 10'11M or greater, 10'12M or greater, or 10'13M or greater. In some embodiments, the polypeptide of the invention can bind to IL-23 with aKDof 106M to 1013M, IO’7to 10’13M, IO’8to IO’13M, or IO’9to IO’13M.

[0144] The antibody or antibody-derived binding moiety can be a TNF-alpha or a TNF receptor targeting antibody or VHH. In some embodiments, the polypeptide of the invention can bind to TNF-alpha with a KD of 106M or less, 10'7M or less, 10'8M or less, 10'9M or less, 10'10M or less, 10'11M or less, 10'12M or less, or 10'13M or less. In some embodiments, the polypeptide of the invention can bind to TNF-alpha with a KD of 106M or greater, 10'7M or greater, 10'8M or greater, 10'9M or greater, 10'10M or greater, 10-11M or greater, 10'12M or greater, or 10'13M or greater. In some embodiments, the polypeptide of the invention can bind to TNF-alpha with a KD of 106M to 1013M, IO’7to 10’13M, IO’8to IO’13M, or IO’9to IO’13M.

[0145] A KD value less than 10'6M can be considered to indicate binding. Specific binding of an antigen-binding polypeptide to an antigen or antigenic determinant can be determined in any suitable known manner, including, for example, Scatchard analysis and / or competitive binding assays, surface plasmon resonance such as radioimmunoassays (RIA), enzyme immunoassays (EIA) and sandwich competition assays, and the different variants thereof known in the art.Stability

[0146] In some embodiments, the polypeptide or construct of the present inventionWSGR Docket No. 60790-720.601substantially retains neutralization ability and / or potency when delivered orally and after exposure to the intestinal tract (for example, after exposure to proteases of the small and / or large intestine and / or IBD inflammatory proteases). Such proteases include enteropeptidase, trypsin, chymotrypsin, and irritable bowel disease inflammatory proteases (such as MMP3, MMP12 and cathepsin). Proteases of, or produced in, the small and / or large intestine include proteases sourced from intestinal commensal microflora and / or pathogenic bacteria, for example wherein the proteases are cell membrane-attached proteases, excreted proteases and proteases released on cell lysis). In some embodiments, the proteases are trypsin and chymotrypsin.

[0147] In some embodiments, the intestinal tract is the intestinal tract of a dog, pig, human, cynomolgus monkey or mouse. In some embodiments, the small intestine can consist of the duodenum jejunum and ileum. In some embodiments, the large intestine can consist of the cecum, colon, rectum and anal canal. In some embodiments, the intestinal tract, as opposed to the gastrointestinal tract, can consist of only the small intestine and the large intestine.

[0148] In some embodiments, the polypeptide or construct of the invention substantially retains neutralization ability after exposure to proteases present in the small and / or large intestine and / or IBD inflammatory proteases for, for example, up to at least 1, more suitably up to at least 2, more suitably up to at least 3, more suitably up to at least 4, more suitably up to at least 7, more suitably up to at least 16 hours at 37 °C.Autoimmune and / or inflammatory diseases

[0149] Autoimmune diseases develop when the immune system responds adversely to normal body tissues and / or commensal microflora and / or pathogenic microflora. Autoimmune disorders may result in damage to body tissues, abnormal organ growth and / or changes in organ function. The disorder may affect only one organ or tissue type or may affect multiple organs and tissues. Organs and tissues commonly affected by autoimmune disorders include blood components such as red blood cells, blood vessels, connective tissues, endocrine glands such as the thyroid or pancreas, muscles, joints and skin An inflammatory disease is a disease characterized by inflammation. Many autoimmune diseases are also considered inflammatory diseases.Autoimmune diseases and / or inflammatory diseases of the GI tract

[0150] A non-limiting list of other autoimmune and inflammatory diseases afflicting the GI tract include GI cancers, colitis, microscopic colitis, celiac disease, mucositis, pouchitis, orWSGR Docket No. 60790-720.601gastritis. A non-limiting list of GI cancers include anal cancer, bile duct cancer, colon cancer, esophageal cancer, small intestine cancer, and gastric cancer. The chronic inflammatory bowel diseases, Crohn’s disease and ulcerative colitis, are examples of autoimmune and inflammatory diseases of the GI tract (Hendrickson et al. 2002 Clin Microbiol Rev 15(l):79-94, herein incorporated by reference in its entirety). Ulcerative colitis is a condition where the inflammatory response and morphologic changes remain confined to the colon. The rectum is involved in 95% of patients. Inflammation is largely limited to the mucosa and consists of continuous involvement of variable severity with ulceration, edema, and hemorrhage along the length of the colon (Hendrickson et al. 2002 Clin. Microbiol Rev 15(l):79-94, herein incorporated by reference in its entirety). Ulcerative colitis is usually manifested by the presence of blood and mucus mixed with stool, along with lower abdominal cramping which is most severe during the passage of bowel movements. Clinically, the presence of diarrhea with blood and mucus differentiates ulcerative colitis from irritable bowel syndrome, in which blood is absent.

[0151] Crohn's disease, also known as Crohn syndrome and regional enteritis, is a type of inflammatory bowel disease causing a wide variety of symptoms. It primarily causes abdominal pain, diarrhea, vomiting and / or weight loss but may also cause complications outside GI tract such as anemia, skin rashes, arthritis, inflammation of the eye, tiredness, and lack of concentration (Baumgart et al. 2012 The Lancet 380(9853): 1590-605, herein incorporated by reference in its entirety). Crohn’s disease is a presently incurable life-long GI disease that is difficult to control with conventional therapies. Unlike ulcerative colitis, the presentation of Crohn’s disease is usually subtle, which leads to a later diagnosis. Factors such as the location, extent, and severity of involvement determine the extent of GI symptoms. Patients who have ileocolonic involvement usually have postprandial abdominal pain, with tenderness in the right lower quadrant and an occasional inflammatory mass. Symptoms associated with gastroduodenal Crohn’s disease include early satiety, nausea, emesis, epigastric pain, or dysphagia. Perianal disease is common, along with anal tags, deep anal fissures, and fistulae (Hendrickson et al. 2002 Clin Microbiol Rev 15(l):79-94, herein incorporated by reference in its entirety).TNF-alpha and IL-23 in autoimmune diseases and / or inflammatory diseases

[0152] Autoimmune and / or inflammatory diseases are characterized by aberrant activation of pro-inflammatory pathways. While the root pathology of autoimmune and / or inflammatory diseases remains to be fully elucidated, therapeutic strategies that aim to dampen pro-inflammatory pathways can provide sustained relief or induce remission to patients suffering from autoimmune and / or inflammatory diseases.WSGR Docket No. 60790-720.601

[0153] TNF-alpha (UniProt ID: P01375, www.uniprot.org / uniprotkb / P01375 / entry) is a pleiotropic cytokine that is involved in a variety of pro-inflammatory cellular processes and is also implicated in the pathogenesis of autoimmune and / or inflammatory diseases. TNF alpha is a homo-trimeric protein that is primarily secreted by immune cell types, including natural killer (NK) cells, T cells, macrophages, and monocytes; and TNF-alpha can also be expressed as a transmembrane protein. Both transmembrane and soluble forms can bind to tumor necrosis factor receptor 1 (TNFR1, also known as p55) and tumor necrosis factor receptor 2 (TNFR2, also known as p75) to induce downstream signaling pathways, including nuclear factor kappa-light-chain-enhancer of activated B cells (NF-KB) and / or mitogen-activated protein kinase (MAPK) signaling.

[0154] IL-23 is a heterodimeric pro-inflammatory cytokine, comprising an IL-23 A subunit (also known as IL-23pl9; UniProt ID: Q9NPF7, www.uniprot.org / uniprotkb / Q9NPF7 / entry) that shares an IL-12B subunit (also known as IL-12 / 23p40, UniProt ID: P29460, www.uniprot.org / uniprotkb / P29460 / entry) with IL-12, that is implicated in the pathogenesis of autoimmune and / or inflammatory diseases. IL-23 is primarily secreted by immune cell types, such as activated macrophages, monocytes, or dendritic cells (DCs), can promote recruitment and activation of immune cells, such as macrophages and granulocytes, and is implicated in the maintenance and expansion of a pro-inflammatory T helper 17 (Thl7) cells. IL-23 signaling is mediated by binding of the IL-23 A subunit (also known as IL-23pl9) to the IL-23 receptor (IL-23R) and the IL-12B subunit to IL- 12 receptor beta 1 (IL-12RP1), thereby inducing JAK-mediated signal transducer and activator of transcription (STAT) signaling. In particular, IL-23 binding can recruit of JAK2 and TYK2 for the induction of STAT3 and / or STAT4 signaling pathways. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered for 8, 12 or 16 weeks followed by a maintenance dose. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered for 12 weeks followed by a maintenance dose. In certain embodiments, the maintenance dose is from about 1300 to about 1400 milligrams. In certain embodiments, the maintenance dose is about 1360 milligrams. In certain embodiments, the local inhibitor of IL-23 and TNF-alpha is administered for another period of 12 weeks if the individual is a non-responder.Inhibitors of IL-23

[0155] Inhibitors of IL-23 described herein can antagonize IL-23-induced cellular processes, which include pro-inflammatory pathways. In some embodiments, inhibitors of IL-23 describedWSGR Docket No. 60790-720.601herein are local inhibitors of IL-23. In some embodiments, inhibitors of IL-23 described herein can bind to human IL-23. In some embodiments, inhibitors of IL-23 described herein can bind to both human and at least one additional primate IL-23 selected from the group consisting of baboon IL-23, marmoset IL-23, cynomolgus IL-23, and rhesus IL-23. In some embodiments, inhibitors of IL-23 described herein can bind to the IL-23 A subunit (also known as IL-23pl9) of IL-23. In some embodiments, inhibitors of IL-23 described herein may bind to a linear or conformational epitope on IL-23. In some embodiments, inhibitors of IL-23 described herein can bind to IL-23R. In some embodiments, inhibitors of IL-23 described herein can block binding of the IL-23 A subunit to IL-23R. In some embodiments, inhibitors of IL-23 described herein can block binding of IL-23 to IL-23R. In some embodiments, inhibitors of IL-23 described herein can bind to the IL-12B subunit (also known as IL-12 / 23p40) of IL-23. In some embodiments, inhibitors of IL-23 described herein can bind to IL-12RPL In some embodiments, inhibitors of IL-23 described herein can block binding of the IL-12B subunit to IL-12RPL In some embodiments, inhibitors of IL-23 described herein can block binding of IL-23 to IL-12RPL In some embodiments, inhibitors of IL-23 described herein can block binding of IL- 12 to IL-12RPL

[0156] In some embodiments, inhibitors of IL-23 described herein can be directed against epitopes on IL-23 that lie in and / or form part of the receptor binding site(s) of IL-23, such that said inhibitors of IL-23, upon binding to IL-23, is capable inhibiting or reducing the IL-23 receptor crosslinking that is mediated by said IL-23 and / or the signal transduction that is mediated by such receptor crosslinking.

[0157] In some embodiments, inhibitors of IL-23 described herein can block signaling pathways activated in response to IL-12. In some embodiments, inhibitors of IL-23 described herein can block signaling pathways activated in response to IL-23. In some embodiments, inhibitors of IL-23 described herein can inhibit JAK / STAT signaling. In some embodiments, inhibitors of IL-23 described herein can inhibit STAT3 signaling, STAT4 signaling, or combinations thereof. In some embodiments, inhibitors of IL-23 described herein can inhibit cellular processes downstream of STAT3 signaling, STAT4 signaling, or combinations thereof. In some embodiments, inhibitors of IL-23 described herein can inhibit expression of pro-inflammatory genes.

[0158] In some embodiments, the inhibitor of IL-23 comprises an antibody that binds IL-23 or an IL-23 binding fragment thereof. In some embodiments, the antibody that binds IL-23 or an IL-23 binding fragment thereof comprises risankizumab, guselkumab, tildrakizumab, briakinumab, brazikumab, mirikizumab, ustekinumab, or combinations thereof.

[0159] In some embodiments, the antibody that binds to IL-23 or an IL-23 binding fragmentWSGR Docket No. 60790-720.601thereof comprises a VHH. In some embodiments, the IL-23 binding VHH comprises: a) a CDR1 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 1, 4, or 7; b) a CDR2 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 2, 5, or 8; and c) a CDR3 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 3, 6, or 9. In some embodiments, the IL-23 binding VHH comprises an amino acid sequence at least about 85%, 90%, 95%, 97%, 98%, or 99% identical to that set forth in any one of SEQ ID NOs: 10-12.

[0160] In some embodiments, inhibitors of IL-23 described herein can bind to one or more epitope(s) on IL-23. In one aspect of the invention there is provided an inhibitor of IL-23 which binds to the same epitope on IL-23 as ID-L253T, 12G1, 1E2, 10E2 or 10G10.Inhibitors of TNF-alpha

[0161] Inhibitors of TNF-alpha described herein can antagonize TNF-alpha-induced cellular processes, including pro-inflammatory pathways. In some embodiments, inhibitors of TNF-alpha described herein are local inhibitors of TNF-alpha. In some embodiments, inhibitors of TNF-alpha described herein can bind to the soluble form of TNF-alpha. In some embodiments, inhibitors of TNF-alpha described herein can bind to transmembrane forms of TNF-alpha. In some embodiments, inhibitors of TNF-alpha described herein can bind to both soluble forms of TNF-alpha, transmembrane forms of TNF-alpha, or combinations thereof. In some embodiments, inhibitors of TNF-alpha described herein can also bind to TNFR1. In some embodiments, inhibitors of TNF-alpha described herein can also bind to TNFR2. In some embodiments, inhibitors of TNF-alpha described herein can also bind to TNFR1, TNFR2, or combinations thereof. In some embodiments, inhibitors of TNF-alpha described herein can bind to both soluble and transmembrane forms of TNF-alpha. In some embodiments, inhibitors of TNF-alpha described herein can block binding of the soluble form of TNF-alpha to TNFR1. In some embodiments, inhibitors of TNF-alpha described herein can block binding of the soluble form of TNF-alpha to TNFR2. In some embodiments, inhibitors of TNF-alpha described herein can block binding of the transmembrane form of TNF-alpha to TNFR1. In some embodiments, inhibitors of TNF-alpha described herein can block binding of the transmembrane form of TNF-alpha to TNFR2. In some embodiments, inhibitors of TNF-alpha described herein can block binding of both the soluble and transmembrane form of TNF-alpha to TNFR1. In some embodiments, inhibitors of TNF-alpha described herein can block binding of both the soluble and transmembrane form of TNF-alpha to TNFR2. In some embodiments, inhibitors of TNF-alpha described herein can block binding of both the soluble and transmembrane form of TNF-alpha to TNFR1, TNRFR2, or combinations thereof.

[0162] In some embodiments, inhibitors of TNF-alpha described herein can be directed againstWSGR Docket No. 60790-720.601epitopes on TNF-alpha that lie in and / or form part of the receptor binding site(s) of TNF-alpha, such that said inhibitors of TNF-alpha, upon binding to TNF-alpha, is capable inhibiting or reducing the TNF-alpha receptor crosslinking that is mediated by said TNF-alpha and / or the signal transduction that is mediated by such receptor crosslinking. In some embodiments, inhibitors of TNF-alpha described herein can block signaling pathways activated in response to TNF-alpha. In some embodiments, inhibitors of TNF-alpha described herein can inhibit NF-KB and / or MAPK signaling. In some embodiments, inhibitors of TNF-alpha described herein can inhibit cellular processes downstream of NF-KB and / or MAPK signaling. In some embodiments, inhibitors of TNF-alpha described herein can inhibit expression of pro-inflammatory genes.

[0163] In some embodiments, the inhibitor of TNF-alpha comprises an antibody that binds TNF-alpha or a TNF-alpha binding fragment thereof. In some embodiments, the antibody that binds TNF-alpha or TNF-alpha binding fragment thereof comprises infliximab, adalimumab, certolizumab pegol, golimumab, or combinations thereof. In some embodiments, the inhibitor of TNF-alpha comprises a soluble TNF-alpha receptor. In some embodiments, the soluble TNF-alpha receptor can comprise etanercept.

[0164] In some embodiments, the antibody that binds to TNF-alpha or a TNF-alpha binding fragment thereof comprises a VHH. In some embodiments, the TNF-alpha binding VHH comprises: a) a CDR1 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 101, 107, 108, or 119; b) a CDR2 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 102, 109-117, or 120; and c) a CDR3 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 103-106 or 121-124. In some embodiments, the TNF-alpha binding VHH comprises an amino acid sequence at least about 85%, 90%, 95%, 97%, 98%, or 99% identical to that set forth in any one of SEQ ID NOs: 118 or 125.Dual inhibitors of IL-23 and TNF-alpha

[0165] Inhibitors described here can be an inhibitor of both IL-23 and TNF-alpha. In some embodiments, inhibitors of both IL-23 and TNF-alpha described herein are local inhibitors of both IL-23 and TNF-alpha. In some embodiments, the inhibitor of both IL-23 and TNF-alpha can comprise an antibody that binds IL-23 and an IL-23 binding fragment thereof. In some embodiments, the inhibitor of both IL-23 and TNF-alpha can comprise an antibody that binds TNF-alpha and a TNF-alpha binding fragment thereof. In some embodiments, the inhibitor of both IL-23 and TNF-alpha can comprise an antibody that binds IL-23, an IL-23 binding fragment thereof, an antibody that binds TNF-alpha, a TNF-alpha binding fragment thereof, orWSGR Docket No. 60790-720.601combinations thereof. In some embodiments, the inhibitor of both IL-23 and TNF-alpha can comprise a covalent fusion of an inhibitor of IL-23 and an inhibitor of TNF-alpha. In some embodiments, the inhibitor of both IL-23 and TNF-alpha can comprise a peptide linker.

[0166] In some embodiments, the inhibitor of both IL-23 and TNF-alpha comprises a combination therapeutic that is a co-formulated composition comprising both an IL-23 and a TNF-alpha inhibitor. In some embodiments, the inhibitor of both IL-23 and TNF-alpha comprises a polypeptide that binds to both IL-23 and TNF-alpha (e.g., an IL-23 and TNF-alpha bispecific antibody). In some embodiments, the polypeptide that binds to both IL-23 and TNF-alpha comprises a first binding region that binds to TNF-alpha and a second binding region that binds to TNF-alpha, wherein the first binding region that binds TNF-alpha comprises: a) a CDR1 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 101, 107, 108, or 119; b) a CDR2 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 102, 109-117, or 120; and c) a CDR3 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 103-106 or 121-124; wherein the second binding region that binds IL-23 comprises: a) a CDR1 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 1, 4, or 7; b) a CDR2 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 2, 5, or 8; and c) a CDR3 comprising an amino acid sequence as set forth in any one of SEQ ID NOs: 3, 6, or 9. In some embodiments, the first binding region that binds TNF-alpha comprises an amino acid sequence at least about 85%, 90%, 95%, 97%, 98%, or 99% identical to that set forth in any one of SEQ ID 118 or 125; and the second binding region that binds IL-23 comprises an amino acid sequence at least about 85%, 90%, 95%, 97%, 98%, or 99% identical to that set forth in any one of SEQ ID NOs: 10-12. In some embodiments, the first binding region and the second binding region are coupled by a linker. In some embodiments, the first binding region and the second binding region are coupled by a protease-labile linker. In some embodiments, the inhibitor of both IL-23 and TNF-alpha comprises an amino acid sequence at least about 85%, 90%, 95%, 97%, 98%, or 99% identical to that set forth in SEQ ID NO: 201. In some embodiments, the inhibitor of both IL-23 and TNF-alpha comprises an amino acid sequence at least about 85%, 90%, 95%, 97%, 98%, or 99% identical to that set forth in SEQ ID NO: 202.Therapeutic use and delivery

[0167] Described herein are methods of treating an autoimmune or inflammatory condition in an individual in need thereof. In some embodiments, the autoimmune or inflammatory conditionWSGR Docket No. 60790-720.601is an autoimmune or inflammatory condition of the GI tract. In some embodiments, the autoimmune or inflammatory condition of the GI tract comprises IBD. In some embodiments, the autoimmune or inflammatory condition of the GI tract comprises Crohn’s disease, ulcerative colitis, colitis, microscopic colitis, celiac disease, mucositis, pouchitis, or gastritis. In some embodiments, the autoimmune or inflammatory condition of the GI tract is Crohn’s disease. In some embodiments, the autoimmune or inflammatory condition of the GI tract is ulcerative colitis.Administration route

[0168] The methods of treatment described herein can encompass systemic administration. The methods of treatment described herein can encompass local administration. In certain embodiments the local administration is oral administration of a polypeptide, immunoglobulin, or VHH that does not achieve systemic availability by an oral route. The methods of treatment described herein can be administered to an individual in need thereof through parenteral administration or enteral administration. The enteral administration route encompasses administration involving the GI tract. In some embodiments, enteral administration encompasses administration to the alimentary canal. Non-limiting examples of enteral administration include oral administration, rectal administration, or direct administration to the alimentary canal or a portion thereof. In some embodiments, enteral administration can be provided as a suppository. In some embodiments, enteral administration is oral administration. In some embodiments, enteral administration is administered by gavage or intubation. In some embodiments, enteral administration can be provided orally as a liquid, as a nutraceutical, or as a powder. In some embodiments, enteral administration can be provided orally in the form of a pill.

[0169] In some embodiments, a method disclosed herein comprises a tissue-targeted therapy. In some embodiments, the tissue- targeted therapy is a local administration. In some embodiments, the tissue-targeted therapy induces one or more improvements in outcomes in the individual.

[0170] In some embodiments, the tissue-targeted therapy is administered to an individual, wherein the therapy has a serum concentration of less than about 500 ng / mL, less than about 450 ng / mL, less than about 400 ng / mL, less than about 350 ng / mL, less than about 300 ng / mL, less than about 250 ng / mL, less than about 200 ng / mL, less than about 150 ng / mL, less than about 100 ng / mL, less than about 90 ng / mL, less than about 80 ng / mL, less than about 70 ng / mL, less than about 60 ng / mL, less than about 50 ng / mL, less than about 40 ng / mL, less than about 30 ng / mL, less than about 20 ng / mL, less than about 10 ng / mL, less than about 5 ng / mL, or less thanWSGR Docket No. 60790-720.601about 1 ng / mL.

[0171] In some embodiments, the tissue-targeted therapy is administered having a serum concentration of less than about 500 ng / mL. In some embodiments, the tissue-targeted therapy is administered to an individual, wherein the therapy has a serum concentration of less than about 200 ng / mL. In some embodiments, the tissue-targeted therapy is administered to an individual, wherein the therapy has a serum concentration of less than about 1 ug / mL.Formulations

[0172] A pharmaceutical composition of the invention may suitably be formulated for oral, intramuscular, subcutaneous or intravenous delivery. The pharmaceutical compositions of the invention may be in a variety of forms. These include, for example, liquid, semi-solid and solid dosage forms, such as liquid solutions (e.g., injectable and infusible solutions), dispersions or suspensions, tablets, pills, powders, liposomes and suppositories. In some embodiments, solid dosage forms are preferred. The inhibitors of IL-23 and TNF-alpha described herein may be incorporated with excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like.

[0173] Typically, the pharmaceutical composition comprises a polypeptide or construct that is an inhibitor of IL-23 and TNF-alpha, and a pharmaceutically acceptable excipient, diluent, or carrier. Examples of pharmaceutically acceptable carriers include one or more of water, saline, phosphate buffered saline, dextrose, glycerol, ethanol and the like, as well as combinations thereof. Pharmaceutically acceptable carriers may further comprise minor amounts of auxiliary substances such as wetting or emulsifying agents, preservatives or buffers, which enhance the shelflife or effectiveness of the polypeptide or construct of the invention. Pharmaceutical compositions may include antiadherents, binders, coatings, disintegrants, flavors, colors, lubricants, sorbents, preservatives, sweeteners, freeze dry excipients (including lyoprotectants) or compression aids.

[0174] In certain embodiments, the inhibitors of IL-23 and TNF-alpha described herein is administered orally. A key problem with oral delivery is ensuring that a sufficient amount of the inhibitors of IL-23 and TNF-alpha reach the area of the intestinal tract where it is required.Factors which may prevent inhibitors of IL-23 and TNF-alpha of the invention from reaching the area of the intestinal tract where it is required may include the presence of proteases in digestive secretions which may degrade a polypeptide, pharmaceutical composition or construct of the invention. In some embodiments, the inhibitors of IL-23 and TNF-alpha described herein are substantially stable in the presence of one or more of such proteases by virtue of the inherentWSGR Docket No. 60790-720.601properties of the inhibitors of IL-23 and TNF-alpha itself. In some embodiments, the inhibitors of IL-23 and TNF-alpha described herein can be lyophilized before being incorporated into a pharmaceutical composition.

[0175] Inhibitors of IL-23 and TNF-alpha of the invention may also be provided with an enteric coating. An enteric coating is a polymer barrier applied on oral medication which helps to protect the polypeptide from the low pH of the stomach. Materials used for enteric coatings include fatty acids, waxes, shellac, plastics, and plant fibers. Suitable enteric coating components include methyl acrylate-methacrylic acid copolymers, cellulose acetate succinate, hydroxy propyl methyl cellulose phthalate, hydroxy propyl methyl cellulose acetate succinate (hypromellose acetate succinate), polyvinyl acetate phthalate (PVAP), methyl methacrylate-methacrylic acid copolymers, sodium alginate and stearic acid. Suitable enteric coatings include pH-dependent release polymers. These are polymers which are insoluble at the highly acidic pH found in the stomach, but which dissolve rapidly at a less acidic pH. Thus, suitably, the enteric coating will not dissolve in the acidic juices of the stomach (pH ~3), but will do so in the higher pH environment present in the small intestine (pH above 6) or in the colon (pH above 7.0). The pH-dependent release polymer is selected such that the polypeptide or construct of the invention will be released at about the time that the dosage reaches the small intestine.

[0176] A lyoprotectant may also be added in order to protect the polypeptide or construct of the invention against destabilizing conditions during the lyophilization process. For example, known lyoprotectants include sugars (including glucose, sucrose, mannose and trehalose); polyols (including mannitol, sorbitol and glycerol); and amino acids (including alanine, glycine and glutamic acid). Lyoprotectants can be included in an amount of about 10 mM to 500 mM.Therapeutic dosage

[0177] Therapeutic dosage can impact the efficacy and durability in disease treatment. The therapeutic dosage can be defined through several metrics. In some instances, therapeutic dosage can by mass (e.g., mg). In some instances, therapeutic dosage can be defined by mass normalized to the weight of an individual in need thereof (e.g., mg / kg). In some instances, therapeutic dosage can be defined by mass normalized to the height of an individual in need thereof (e.g., mg / m). In some instances, therapeutic dosage can be defined by mass normalized to the body surface area of an individual in need thereof (e.g., mg / m2).

[0178] The dosage ranges for administration of the inhibitors of IL-23 and TNF-alpha described herein are those to produce the desired therapeutic effect. The dosage range required depends on the precise nature of the inhibitor of IL-23 and TNF-alpha, the route ofWSGR Docket No. 60790-720.601administration, the nature of the formulation, the age of the patient, the weight, height and body surface are of the patient, the nature, extent or severity of the patient's condition, contraindications, if any, and the judgement of the attending physician. Variations in these dosage levels can be adjusted using standard empirical routines for optimization.

[0179] In some embodiments, the therapeutic can be an inhibitor of IL-23, an inhibitor of TNF-alpha, or an inhibitor of both IL-23 and TNF-alpha. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 100 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 135 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 270 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 405 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 540 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 675 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 810 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 945 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 1,080 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 1,215 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 1,350 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 2,025 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 2,700 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 3,645 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 5,400 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 6,750 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 8,100 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 10,935 mg.

[0180] In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 100 mg to about 10,935 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 100 mg to about 135 mg, about 100 mg to about 270 mg, about 100 mg to about 405 mg, about 100 mg to about 540 mg, about 100 mg to about 675 mg, about 100 mg to about 1,215 mg, about 100 mg to about 2,700 mg,WSGR Docket No. 60790-720.601about 100 mg to about 3,645 mg, about 100 mg to about 6,750 mg, about 100 mg to about 8,100 mg, about 100 mg to about 10,935 mg, about 135 mg to about 270 mg, about 135 mg to about 405 mg, about 135 mg to about 540 mg, about 135 mg to about 675 mg, about 135 mg to about 1,215 mg, about 135 mg to about 2,700 mg, about 135 mg to about 3,645 mg, about 135 mg to about 6,750 mg, about 135 mg to about 8,100 mg, about 135 mg to about 10,935 mg, about 270 mg to about 405 mg, about 270 mg to about 540 mg, about 270 mg to about 675 mg, about 270 mg to about 1,215 mg, about 270 mg to about 2,700 mg, about 270 mg to about 3,645 mg, about 270 mg to about 6,750 mg, about 270 mg to about 8,100 mg, about 270 mg to about 10,935 mg, about 405 mg to about 540 mg, about 405 mg to about 675 mg, about 405 mg to about 1,215 mg, about 405 mg to about 2,700 mg, about 405 mg to about 3,645 mg, about 405 mg to about 6,750 mg, about 405 mg to about 8,100 mg, about 405 mg to about 10,935 mg, about 540 mg to about 675 mg, about 540 mg to about 1,215 mg, about 540 mg to about 2,700 mg, about 540 mg to about 3,645 mg, about 540 mg to about 6,750 mg, about 540 mg to about 8,100 mg, about 540 mg to about 10,935 mg, about 675 mg to about 1,215 mg, about 675 mg to about 2,700 mg, about 675 mg to about 3,645 mg, about 675 mg to about 6,750 mg, about 675 mg to about 8,100 mg, about 675 mg to about 10,935 mg, about 1,215 mg to about 2,700 mg, about 1,215 mg to about 3,645 mg, about 1,215 mg to about 6,750 mg, about 1,215 mg to about 8,100 mg, about 1,215 mg to about 10,935 mg, about 2,700 mg to about 3,645 mg, about 2,700 mg to about 6,750 mg, about 2,700 mg to about 8,100 mg, about 2,700 mg to about 10,935 mg, about 3,645 mg to about 6,750 mg, about 3,645 mg to about 8,100 mg, about 3,645 mg to about 10,935 mg, about 6,750 mg to about 8,100 mg, about 6,750 mg to about 10,935 mg, or about 8,100 mg to about 10,935 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is about 100 mg, about 135 mg, about 270 mg, about 405 mg, about 540 mg, about 675 mg, about 1,215 mg, about 2,700 mg, about 3,645 mg, about 6,750 mg, about 8,100 mg, or about 10,935 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is at least about 100 mg, about 135 mg, about 270 mg, about 405 mg, about 540 mg, about 675 mg, about 1,215 mg, about 2,700 mg, about 3,645 mg, about 6,750 mg, or about 8,100 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is at most about 135 mg, about 270 mg, about 405 mg, about 540 mg, about 675 mg, about 1,215 mg, about 2,700 mg, about 3,645 mg, about 6,750 mg, about 8,100 mg, or about 10,935 mg.

[0181] In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is a total daily dose of about 600 mg to about 3,200 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is a total daily doseWSGR Docket No. 60790-720.601of about 600 mg to about 1,600 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is a total daily dose of about 1,200 mg to about 3,200 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is a total daily dose of about 1,200 mg to about 1,600 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is a total daily dose of about 1,800 mg to about 2,400 mg. In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is a total daily dose of about 2,400 mg to about 3,200 mg.

[0182] In some embodiments, the inhibitor of IL-23 and TNF-alpha is dosed once daily. In some embodiments, the inhibitor of IL-23 and TNF-alpha is dosed once daily for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 weeks. In certain embodiments, the inhibitor of IL-23 and TNF-alpha is dosed once daily for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36,37,38,39,40, 41, 42, 43, 44, 45, 46,47,48,49,50,51, or 52 weeks or more. In some embodiments, the inhibitor of IL-23 and TNF-alpha is dosed twice daily. In some embodiments, the inhibitor of IL-23 and TNF-alpha is dosed twice daily for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 weeks. In certain embodiments, the inhibitor of IL-23 and TNF-alpha is dosed twice daily for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36,37,38,39,40, 41, 42, 43, 44, 45, 46, 47,48,49,50,51, or 52 weeks or more. In certain embodiments, the inhibitor of IL-23 and TNF-alpha is dosed until the individual achieves a clinical outcome and / or endpoint described herein.

[0183] In some embodiments, the inhibitor of IL-23 and TNF-alpha can be administered twice daily at a dose of about 600 mg to achieve a total daily dose of about 1200 mg. In some embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1300 mg. In some embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1400 mg. In some embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 1300 mg to about 1400 mg. In some embodiments, the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1360 mg. In some embodiments, the inhibitor of IL-23 and TNF-alpha can be administered twice daily at a dose of about 900 mg to achieve a total daily dose of about 1800 mg. In some embodiments, the inhibitor of IL-23 and TNF-alpha can be administered twice daily at a dose of about 1200 mg to achieve a total daily dose of about 2400 mg. In some embodiments, the inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 2000 mg to about 2800 mg. In some embodiments, the inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 2000 mg to about 2800 mg. In some embodiments,WSGR Docket No. 60790-720.601the inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2040 mg. In some embodiments, the inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2720 mg.

[0184] In some embodiments, the therapeutically effective amount of an inhibitor of IL-23 and TNF-alpha is an increment of about 135 mg, about 270 mg, about 405 mg, about 540 mg, about 675 mg, about 810 mg, about 945 mg, about 1,080 mg, about 1, 215 mg, about 1,350 mg, about 1,485 mg, about 1,620 mg, about 1,755 mg, about 1,890 mg, about 2,025 mg, about 2,160 mg, about 2,295 mg, about 2,430 mg, about 2,565 mg, about 2,700 mg, about 2,835 mg, about 2,970 mg, about 3,105 mg, 3,240 mg, 3,375 mg, 3,510 mg, 3,645 mg, 3,780 mg, 3,915 mg, 4,050 mg, 4,185 mg, 4,320 mg, about 4,455 mg, about 4,725 mg, about 4,860 mg, about 4,995 mg, about 5,130 mg, about 5,265 mg, about 5,400 mg, about 5,535 mg, about 5,670 mg, about 5,805 mg, about 5,940 mg, about 6,075 mg, about 6,210 mg, about 6,345 mg, about 6,480 mg, about 6,615 mg, about 6,750 mg, about 6,885 mg, 7,020 mg, about 7,155 mg, about 7,290 mg, about 7,425 mg, about 7,560 mg, about 7,695 mg, about 7,830 mg, about 7,965 mg, about 8,100 mg, about 8,235 mg, about 8,370 mg, about 8,505 mg, about 8,640 mg, about 8,775 mg, about 8,910 mg, about 9,045 mg, about 9,180 mg, about 9,315 mg, about 9,450 mg, about 9,585 mg, about 9,720 mg, about 9,855 mg, about 9,990 mg, about 1,0125 mg, about 1,0260 mg, about 1,0395 mg, about 1,0530 mg, about 1,0665 mg, about 1,0800 mg, or about 1,0935 mg.Treatment schedule

[0185] In some embodiments, inhibitors of IL-23 and TNF-alpha described herein can be administered as a single dose. In some embodiments, inhibitors of IL-23 and TNF-alpha described herein can be administered in multiple doses. In some embodiments, inhibitors of IL-23 and TNF-alpha described herein can be administered once a day. In some embodiments, inhibitors of IL-23 and TNF-alpha described herein can be administered until a therapeutic effect is achieved. In some embodiments, inhibitors of IL-23 and TNF-alpha described herein can be administered once a day. In some embodiments, inhibitors of IL-23 and TNF-alpha described herein can be administered once a day until a therapeutic effect is achieved. In some embodiments, inhibitors of IL-23 and TNF-alpha described herein can be administered twice a day. In some embodiments, inhibitors of IL-23 and TNF-alpha described herein can be administered twice a day until a therapeutic effect is achieved.

[0186] In some embodiments an individual is administered an induction course of treatment followed by a maintenance course of treatment. In some embodiments, individuals are administered a maintenance course of treatment after a course of induction treatment, theWSGR Docket No. 60790-720.601induction treatment lasting about 8, 10, 12, 14, or 16 weeks. In some embodiments, individuals are administered a maintenance course of treatment after a course of induction treatment, the induction treatment lasting about 8 weeks. In some embodiments, individuals are administered a maintenance course of treatment after a course of induction treatment, the induction treatment lasting about 12 weeks. In some embodiments, individuals are administered a maintenance course of treatment after a course of induction treatment, the induction treatment lasting about 16 weeks. Such maintenance may proceed at a lower dose (about 10%, 20%, 30%, 40%, or 50%) lower than the induction dose or a lower frequency (e.g., QD vs BID). In some embodiments, maintenance is initiated after a clinical response is observed. In some embodiments, maintenance is initiated after clinical remission is observed. In some embodiments, patients who do not experience clinical improvement of remission are retreated (e.g., reinduced) after about 8, 10, 12, 14, or 16 weeks. The maintenance dose may be about 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 775 mg, 800 mg, 825 mg, 850 mg, 900 mg, 950 mg, 1,000 mg, 1,100 mg, 1,200 mg, 1,300 mg, 1,400 mg, 1,500 mg, or 1,600 mg. The maintenance dose may be about 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 775 mg, 800 mg, 825 mg, 850 mg, 900 mg, 950 mg, 1,000 mg, 1,100 mg, 1,200 mg, 1,300 mg, 1,400 mg, 1,500 mg, or 1,600 mg or more. The maintenance dose may be about 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 775 mg, 800 mg, 825 mg, 850 mg, 900 mg, 950 mg, 1,000 mg, 1,100 mg, 1,200 mg, 1,300 mg, 1,400 mg, 1,500 mg, or 1,600 mg or less.Clinical outcomes

[0187] The treatments prescribed herein can result in improvement in outcomes for patients afflicted with autoimmune and / or inflammatory conditions. In some embodiments, the treatments prescribed herein can improve symptoms of patients afflicted with autoimmune and / or inflammatory conditions. In some embodiments, the treatments prescribed herein can provide short-term relief of patients afflicted with autoimmune and / or inflammatory conditions. In some embodiments, the treatments prescribed herein can provide long-term relief of patients afflicted with autoimmune and / or inflammatory conditions. In certain embodiments the autoimmune and / or inflammatory conditions comprises colitis or ulcerative colitis.

[0188] Evaluation of adverse events following therapeutic administration can be important in determining an optimal therapeutic dosage. In some embodiments, administration of placebo controls can be helpful in determining an optimal therapeutic dosage. In some embodiments, treatment emergent adverse events can be monitored after single administration of inhibitors ofWSGR Docket No. 60790-720.601IL-23 and TNF-alpha. In some embodiments, treatment emergent adverse events can be monitored after single administration of placebo controls.

[0189] Clinical, endoscopic, histological, and composite instruments as well as physician assessments can be used to measure disease activity in patients with ulcerative colitis (UC). Nonlimiting examples of instruments for determining measures of disease activity include: Mayo Clinic score (MCS), modified MCS (mMS; excluding physician global assessment), partial MCS (pMS; MCS without endoscopic subscore), Robart's Histopathology Index (RHI), UC-100 score, and Inflammatory Bowel Disease Questionnaire (IBDQ). Changes in one or more measures of disease activity in patients with moderately to severely active UC can be used to assess severity of disease and / or effectiveness of a treatment in patients. Depending on the treatment setting, time point of evaluation, and feasibility of measurement, different scores may be used to demonstrate response.

[0190] The UC-100 scale ranges from 1-100 with 100 being the most severe. In certain embodiments, administering an inhibitor of IL-23 and TNF-alpha to an individual, as described herein can reduce a UC-100 score in an individual afflicted with an autoimmune disorder (e.g., colitis or ulcerative colitis). In certain embodiments, the UC-100 score is reduced by about 5 or more, about 10 or more, about 15 or more, about 20 or more, about 25 or more, about 30 or more, about 40 or more, about 45 or more, about 50 or more, about 55 or more, about 60 or more, about 65 or more, about 70 or more, about 75 or more, about 80 or more, about 85 or more, about 90 or more, about 95 or more, or about 99 or more, compared to a baseline. In certain embodiments, the baseline comprises the UC-100 of the individual prior to administering the inhibitor of IL-23 and TNF-alpha.

[0191] In certain embodiments, the UC-100 score is reduced to about 30 or less, about 29 or less, about 28 or less, about 27 or less, about 26 or less, about 25 or less, about 24 or less, about 23 or less, about 22 or less, about 21 or less, about 20 or less, about 19 or less, about 18 or less, about 17 or less, about 16 or less, about 15 or less, about 14 or less, about 13 or less, about 12 or less, about 11 or less, about 10 or less, about 9 or less, about 8 or less, about 7 or less, about 6 or less, or about 5 or less.

[0192] Robarts Histopathology Index (RHI) measures histological disease activity in ulcerative colitis. In certain embodiments, administering an inhibitor of IL-23 and TNF-alpha to an individual, as described herein can reduce an RHI in an individual afflicted with an autoimmune disorder (e.g., colitis or ulcerative colitis). In certain embodiments, the RHI is reduced by about 4 or more, about 5 or more, about 6 or more, about 7 or more, about 8 or more, about 9 or more, about 10 or more, about 11 or more, about 12 or more, about 13 or more, aboutWSGR Docket No. 60790-720.60114 or more, about 15 or more, about 16 or more, about 17 or more, about 18 or more, about 19 or more, about 20 or more, about 21 or more, about 22 or more, about 23 or more, about 24 or more, about 25 or more, about 26 or more, about 27 or more, about 28 or more, about 29 or more, about 30 or more, about 31 or more, or about 32 or more, compared to a baseline. In certain embodiments, the baseline comprises the RHI of the individual prior to administering the inhibitor of IL-23 and TNF-alpha. In certain embodiments, the RHI is reduced to about 10 or less, about 9 or less, about 8 or less, about 7 or less, about 6 or less, about 5 or less, about 4 or less, about 3 or less, about 2 or less, about 1 or less, or 0. In certain embodiments, the RHI is reduced to 0. In certain embodiments, the RHI is reduced to 1.

[0193] In certain embodiments, the treating achieves histological remission. A non-limiting example of a histological remission includes an absence of neutrophils from the mucosa (e.g., lamina propria and / or epithelium). In certain embodiments, a subscore of the RHI is 0. In certain embodiments, a subscore of 0 is achieved for lamina propria neutrophils. In certain embodiments, a subscore of 0 is achieved for neutrophils in epithelium. Another non-limiting example of a histological remission includes an absence of basal plasmacytosis. In certain embodiments, the absence of basal plasmacytosis comprises a presence of plasma cells between the base of the crypts and the muscularis mucosae in the colon.

[0194] The Mayo Clinic Score Outcome Assessment for ulcerative colitis disease activity provides an assessment of disease severity and can be used to monitor patients during a therapy. The Full Mayo Clinic Score (MCS) comprises 4 categories: rectal bleeding subscore (RBS), stool frequency (SF), physician’s global assessment (PGA), and endoscopy subscore (ES), each rated with a score from 0 to 3 with a total score ranging from 0 to 12, and higher scores indicating greater disease severity. In certain embodiments, a score of 3 to 5 points can indicate a mildly active disease, a score of 6 to 10 points can indicate a moderately active disease, and a score of 11 to 12 points can indicate a severely active disease.

[0195] Two abridged versions, the partial Mayo Clinic score (partial MCS) that excludes the endoscopy subscore is a nine-point score comprising the RBS, SF, and PGA subscores, and the modified Mayo Clinic score (modified MCS) is a six-point score comprising only the RBS and SF subscores, can also be used to provide an assessment of disease severity and can be used to monitor patients during a therapy. The Mayo score can correlate with patient assessment of change in UC activity.

[0196] In certain embodiments, a method disclosed herein achieves remission by a Full Mayo Score. In certain embodiments, a method disclosed herein achieves remission by a Partial Mayo score. In certain embodiments, a method disclosed herein achieves remission by a ModifiedWSGR Docket No. 60790-720.601Mayo score. In certain embodiments, clinical remission of UC can be determined from a Mayo score of about 2 points or less.

[0197] In certain embodiments, a reduction of about 3 points or more on the Mayo score constitutes a clinically response. In certain embodiments, administering an inhibitor of IL-23 and TNF-alpha to an individual, as described herein can reduce a Mayo score (MS) in an individual afflicted with an autoimmune disorder (e.g., colitis or ulcerative colitis). In certain embodiments, the MS is reduced by about 1 or more, by about 2 or more, by about 3 or more, by about 4 or more, about 5 or more, about 6 or more, about 7 or more, about 8 or more, about 9 or more, about 10 or more, about 11 or more, or about 12, compared to a baseline. In certain embodiments, the baseline comprises the MS of the individual prior to administering the inhibitor of IL-23 and TNF-alpha.

[0198] In certain embodiments, the MS is a Full Mayo score. In certain embodiments, the Mayo score is a Partial Mayo score. In certain embodiments, the Mayo score is a Modified Mayo score.

[0199] Inflammatory Bowel Disease Questionnaire (IBDQ) comprises a physician-administered questionnaire to assess health-related quality of life in patients afflicted with inflammatory bowel disease (IBD) (e.g., UC and Crohn disease). An IBDQ is a 32-item Likertbased questionnaire divided into four dimensions: bowel symptoms (10 items), systemic symptoms (5 items), emotional function (12 items), and social function (5 items). Responses to each question are graded from 1 to 7 (1 being the worst situation and 7 the best). Accordingly, the total IBDQ score ranges between 32 (very poor health-related quality of life) to 224 (perfect health-related quality of life). The score of patients in remission can be >170. An increase in IBDQ score of >16 points can represent a clinically meaningful improvement in health -related quality of life in patients afflicted with IBD.

[0200] In certain embodiments, administering an inhibitor of IL-23 and TNF-alpha to an individual, as described herein can improve an IBDQ in an individual afflicted with an autoimmune disorder (e.g., colitis or ulcerative colitis). In certain embodiments, the IBDQ is increased by about 15 or more, by about 2 or more, by about 3 or more, by about 4 or more, about 5 or more, about 6 or more, about 7 or more, about 8 or more, about 9 or more, about 10 or more, or about 11, compared to a baseline. In certain embodiments, the baseline comprises the MS of the individual prior to administering the inhibitor of IL-23 and TNF-alpha.

[0201] Mayo Endoscopic Score (MES) is a clinical tool to assess the severity of inflammation in patients with ulcerative colitis. Mayo endoscopic subscore (MES or ES) ranges from 0 to 3, with higher scores indicating greater disease severity. In certain embodiments,WSGR Docket No. 60790-720.601administering a local inhibitor of IL-23 and TNF-alpha to an individual, as described herein can reduce an MES in an individual afflicted with an autoimmune disorder (e.g., colitis or ulcerative colitis). In certain embodiments, the MES score is reduced by about 1 or more compared to a baseline. In certain embodiments, the MES score is reduced by about 2 or more compared to a baseline. In certain embodiments, the MES score is reduced by about 3 compared to a baseline. In certain embodiments, the baseline comprises the MES of the individual prior to administering the inhibitor of IL-23 and TNF-alpha.

[0202] In certain embodiments, administering a inhibitor of IL-23 and TNF-alpha to an individual, as described herein can achieve endoscopic normalization in an individual afflicted with an autoimmune disorder (e.g., colitis or ulcerative colitis). In certain embodiments, administering a local inhibitor of IL-23 and TNF-alpha to an individual, as described herein can induce an endoscopic response in an individual afflicted with an autoimmune disorder (e.g., colitis or ulcerative colitis).

[0203] In certain embodiment, administering a inhibitor of IL-23 and TNF-alpha to an individual, as described herein can improve a histological response in an individual afflicted with an autoimmune disorder (e.g., colitis or ulcerative colitis). In certain embodiments, the administering reduces neutrophils in the epithelium of the intestine of the individual. In certain embodiments, the administering reduces neutrophils in the epithelium of the intestine of the individual to about less than 5%. Non-limiting examples of histological improvement include: no crypt destruction, no erosions, no ulcerations, and no granulation tissue.

[0204] In certain embodiment, administering an inhibitor of IL-23 and TNF-alpha to an individual, as described herein can increased expression of mature epithelial cell associates genes.Non-limiting examples of patient selection criteria

[0205] Therapeutic efficacy may vary according to factors such as the disease state, age, sex, and weight of the individual. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may be at least 18 years of age. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may be at most 55, 60, 65, 70, or 75 years of age. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may be male. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may be female. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may not be gender based. In some embodiments, an individual being treatedWSGR Docket No. 60790-720.601with inhibitors of IL-23 and TNF-alpha described herein can be a healthy volunteer. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein can be of any ethnic origin. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may have a body mass index (BMI) of about 18.0 to 35.0 kg / m2(inclusive). In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein can be healthy as determined by a responsible physician. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein can have normal bowel habits as defined by usual passage of daily stool.

[0206] In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may not have a clinically relevant history of abnormal physical or mental health. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may not have clinically relevant abnormal laboratory results. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may not have history or clinical evidence of any disease and / or existence of any surgical or medical condition which might interfere with the absorption, distribution, metabolism or excretion of the study drug. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may not have any other concomitant disease or condition that could interfere with the conduct of the study. In some embodiments, an individual being treated with inhibitors of IL-23 and TNF-alpha described herein may not have veins unsuitable for venipuncture and / or cannulation.

[0207] While preferred embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention.

[0208] All publications, patent applications, issued patents, and other documents referred to in this specification are herein incorporated by reference as if each individual publication, patent application, issued patent, or other document was specifically and individually indicated to be incorporated by reference in its entirety. Definitions that are contained in text incorporated by reference are excluded to the extent that they contradict definitions in this disclosure.WSGR Docket No. 60790-720.601Table 1: Amino acid sequences or polynucleotide sequences disclosed are represented herein. SEQ ID NO: Sequence Description1 FKVMG IL-23: ID-L253T CDR1 2 TITTGGSPDYSDSVKG IL-23: ID-L253T CDR2 3 RLWAHITSSGHDV IL-23: ID-L253T CDR3 4 GSAMG IL-23: 10E2 CDR1 5 VI I WTGGYT YYADS VK IL-23: 10E2 CDR2 6 RTPNSSWFPTTLYEYD IL-23: 10E2 CDR3 7 NY IMG IL-23: 10G10 CDR1 8 I ITANGVATDYRDSVK IL-23: 10G10 CDR2 9 RTRSGSLFDPGTFDY IL-23: 10G10 CDR3 DVQLVESGGGQVQPGGSLSLSCESSVSIWSFKVMGWFRQ VPGKQRELVATITTGGSPDYSDSVKGRFTISRDYDKRTL10 YLQMNSLKPEDTAVYYCAGRLWAHITSSGHDVWGQGTQV IL-23: ID-L253T TVSS EVQLVESGGGLAQAGGSLRLSCAPSGRAFSGSAMGWFRQ VPGKERE FVGVI I WTGGYT YYADS VKGRFT I SRDSAKNT11 VYLQMDSLKPEDTAVYYCAARTPNSSWFPTTLYEYDYWG IL-23: 10E2QGTQVTVSS EVQLVESGGGLVQAGTSLRLSCAASGRTFSNYIMGWFRQ APGKEREFVAIITANGVATDYRDSVKGRFTISRDNARNT12 GYLEMNSLKPEDTAVYFCALRTRSGSLFDPGTFDYWGQG IL-23: 10G10TQVTVSS EVQLVESGGGQVQPGGSLSLSCESSVSIWSFKVMGWFRQ VPGKQRELVATITTGGSPDYSDSVKGRFTISRDYDKRTL13 YLQMNSLKPEDTAVYYCAGRLWAHITSSGHDVWGQGTQV IL-23: D1E ID-L253T TVSS101 SHWMY TNF-alpha: ID-38F CDR1 102 EINTNGLITHYGDSVK TNF-alpha: ID-38F CDR2 103 NQHGLN TNF-alpha: ID-38F CDR3 104 NQKGLN TNF-alpha: Q65F2 CDR3 105 NQMGLN TNF-alpha: Q65F1 CDR3 106 NERGLN TNF-alpha: Q65D3 CDR3 107 VHWMY TNF-alpha: Q65F1 CDR1 108 NHWMC TNF-alpha: Q65D1 CDR1 109 EINTNGLITHYGDSVH TNF-alpha: ID27F CDR2 110 EINTNGLITKYGDSVH TNF-alpha: ID28F CDR2 111 EINTNGLITSYVDSVK TNF-alpha: Q65F2 CDR2 112 EINTNGLITKYIDSVR TNF-alpha: Q65F3 CDR2 113 EINTNGLITNYVDSVK TNF-alpha: Q62F2 CDR2 114 E INTNGL IT KY I DS VG TNF-alpha: Q65F1 CDR2 115 EINTNGLITKYADFVK TNF-alpha: Q65D1 CDR2 116 EINTNGLITKYADSTK TNF-alpha: Q65D3 CDR2117 EINTNGLITKYGDSVK TNF-alpha: Q65B1 CDR2WSGR Docket No. 60790-720.601DVQLVESGGGLVQPGGSLKLSCAASGFDFSSHWMYWVRQ118 APGKELEWLSEINTNGLITHYGDSVKGRFTVSRNNAANK TNF-alpha: ID-38F MYLELTRLEPEDTALYYCARNQHGLNKGQGTQVTVSS119 IHWMY TNF-alpha: Q62E10 CDR1 120 EINTNALITKYADSVK TNF-alpha: Q62E10 CDR2 121 TQNGAA TNF-alpha: Q62E10 CDR3 122 SRNGAA TNF-alpha: Q65F6 CDR3 123 ARNGAA TNF-alpha: Q65F11 CDR3 124 TQNGKT TNF-alpha: Q62F10 CDR3QVQLVESGGGLVQPGGSLRLSCTTSGLDFGIHWMYWFRQ125 APGKELEWVAEINTNALITKYADSVKGRFTISRDNAKNT TNF-alpha: Q62E10 LFLQMNDLKSEDTAVYYCSNTQNGAAKGQGVQVTVSSDVQLVESGGGLVQPGGSLKLSCAASGFDFSSHWMYWVRQ APGKELEWLSEINTNGLITHYGDSVKGRFTVSRNNAANK MYLELTRLEPEDTALYYCARNQHGLNKGQGTQVTVSSGG TNF-alpha / IL-23 dual 201 GGSGGGGSKGGGGSGGGGSEVQLVESGGGQVQPGGSLSL SCESSVSIWSFKVMGWFRQVPGKQRELVATITTGGSPDY inhibitor: FA1K SDSVKGRFTISRDYDKRTLYLQMNSLKPEDTAVYYCAGR LWAHITSSGHDVWGQGTQVTVSSDVQLVESGGGQVQPGGSLSLSCESSVSIWSFKVMGWFRQ VPGKQRELVATITTGGSPDYSDSVKGRFTISRDYDKRTL YLQMNSLKPEDTAVYYCAGRLWAHITSSGHDVWGQGTQV IL-23 / TNF-alpha dual 202 TVSSGGGGSGGGGSKGGGGSGGGGSEVQLVESGGGLVQP GGSLKLSCAASGFDFSSHWMYWVRQAPGKELEWLSEINT inhibitor: FA1K-based NGLITHYGDSVKGRFTVSRNNAANKMYLELTRLEPEDTA LYYCARNQHGLNKGQGTQVTVSSDVQLVESGGGLVQPGGSLKLSCAASGFDFSSHWMYWVRQ APGKELEWLSEINTNGLITHYGDSVKGRFTVSRNNAANK TNF-alpha fragment of 203 MYLELTRLEPEDTALYYCARNQHGLNKGQGTQVTVSSGG cleaved TNF-alpha / IL-23GGSGGGGSK dual inhibitorGGGGSGGGGSEVQLVESGGGQVQPGGSLSLSCESSVSIW SFKVMGWFRQVPGKQRELVATITTGGSPDYSDSVKGRFT IL-23 fragment of cleaved 204 ISRDYDKRTLYLQMNSLKPEDTAVYYCAGRLWAHITSSG TNF-alpha / IL-23 dual HDVWGQGTQVTVSS inhibitorDVQLVESGGGQVQPGGSLSLSCESSVSIWSFKVMGWFRQ VPGKQRELVATITTGGSPDYSDSVKGRFTISRDYDKRTL IL-23 fragment of cleaved 205 YLQMNSLKPEDTAVYYCAGRLWAHITSSGHDVWGQGTQV IL-23 / TNF-alpha dual TVSSGGGGSGGGGSK inhibitorWSGR Docket No. 60790-720.601GGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFDF SSHWMYWVRQAPGKELEWLSEINTNGLITHYGDSVKGRF TNF-alpha fragment of 206 TVSRNNAANKMYLELTRLEPEDTALYYCARNQHGLNKGQ cleaved IL-23 / TNF-alpha GTQVTVSS dual inhibitor301 GGGGSKGGGGS Linker 302 GGGGSGGGGSKGGGGSGGGGS LinkerWSGR Docket No. 60790-720.601EXAMPLES

[0209] The following illustrative examples are representative of embodiments of compositions and methods described herein and are not meant to be limiting in any way.Example 1: Human clinical study of orally delivered bispecific anti-TNF-alpha / anti-IL-23 domain antibody

[0210] Healthy adult subjects and patients with mild to severe ulcerative colitis (UC) were enrolled in 3-part first-in-human Phase 1 clinical studies to evaluate an orally delivered bispecific anti-TNF-alpha / anti-IL-23 domain antibody (SEQ ID NO: 201); hereinafter in the examples, “TNF-alpha / IL-23 dual inhibitor”) (Table 2). TNF-alpha / IL-23 dual inhibitor contained two humanized single domain heavy chain VHH antibodies of camelid origin targeting TNF-alpha (SEQ ID NO: 118) and IL-23pl9 (SEQ ID NO: 13) connected by a trypsin-labile linker (SEQ ID NO: 302), which can enable monomer separation within the small and / or large intestine. TNF-alpha / IL-23 dual inhibitor can inhibit TNF-alpha and IL-23 activity as an intact molecule and after trypsin cleavage into its monomeric arms yielding cleavage products comprising a liberated anti-TNF-alpha monomer arm (SEQ ID NO: 203) and a liberated anti-IL-23 monomer arm (SEQ ID NO: 204). TNF-alpha / IL-23 dual inhibitor single domain antibodies were engineered for high stability among intestinal and inflammatory proteases to enable oral dosing.

[0211] Part 1 (single ascending dose) and Part 2 (multiple dose) in healthy volunteers showed no safety or tolerability concerns at TNF-alpha / IL-23 dual inhibitor doses above Phase lb dose levels.

[0212] Part 3, a Phase lb study, was a randomized, placebo-controlled, double-blind, doubledummy, multiple dose exploratory study. Twenty-two UC patients were enrolled in the study: (1) a 810mg oral dose of TNF-alpha / IL-23 dual inhibitor administered once a day (QD) (n = 7 patients), (2) a 810mg oral dose of TNF-alpha / IL-23 dual inhibitor administered twice a day (BID) for a total oral dose of 1620mg of TNF-alpha / IL-23 dual inhibitor per day (n = 9 patients), or (3) an oral placebo administered BID (n = 6 patients). 17 of the 22 patients completed the treatment. Of the 17 completers, three parallel groups of patients were evenly randomized to receive daily for 42 consecutive days: (1) a 810mg oral dose of TNF-alpha / IL-23 dual inhibitor administered once a day (QD) (n = 6 patients), (2) a 810mg oral dose of TNF-alpha / IL-23 dual inhibitor administered twice a day (BID) for a total oral dose of 1620mg of TNF-alpha / IL-23 dual inhibitor per day (n = 5 patients), or (3) an oral placebo administered BID (n = 6 patients) (Tables 2-9). Patients enrolled to receive doses of TNF-alpha / IL-23 dual inhibitor QD received TNF-alpha / IL-23 dual inhibitorWSGR Docket No. 60790-720.601for one of their doses and matching placebo capsule for their second daily dose (Table 3).Table 2: Pharmacological Properties of TNF-alpha / IL-23 dual inhibitor.Property Assay [description THF-WhaM-23 duat mhimerBinding aftemy § contifmed by ELISA Sitety Assay* 7HFs Potency (H.iSAj nhssNsn si' human sTNFu btedteg Io.21 nM INFs potency fce&based assay! tehibftten st sT UFs NA induced ce&iar touoty 6.-23 Potency (EUSA) AhibSten st H.-23; Binding te IL-23R^cass Small intestteat stability % Potency alter 2-hsur ista iCsfec NA 4-hcur ijsafetesH NA Huntes % Patency alter Faecal Ststntiiy 4-hear 3-ate«ta St (TNFcj. 67 (lt-23)16-hour incahata NA24-tottr incs batten NATable 3: Exemplary Phase lb Dosing and Dose Regimen of TNF-alpha / IL-23 dual inhibitor.Treatment Arm Treatment Assignment Dosing RegimenArm 1 TNF-alpha / IL-23 dual inhibitor TNF-alpha / IL-23 dual inhibitor 6 capsules 810 mg once a day (QD) AM+Matching Placebo 6 capsules PM Arm 2 TNF-alpha / IL-23 dual inhibitor TNF-alpha / IL-23 dual inhibitor 6 capsules 810 mg twice a day (BID) AM+TNF-alpha / IL-23 dual inhibitor 6 capsules PMArm 3 Placebo twice a day (BID) Matching Placebo 6 capsules AM+Matching Placebo 6 capsules PM

[0213] The study population met Mayo Score criterion for moderate-severe UC. Key inclusion criteria comprised a Mayo Score of 4 to 12, including Mayo ES of > 2 as determined by central reader and rectal bleeding subscore (RBS) of > 1 and stool frequency subscore (SFS) of > 1 (Tables 4-6). Key exclusion criteria included prior primary efficacy failure or secondary loss of response to >1 biologic or new small molecule therapy or prior primary non-response, secondary loss ofWSGR Docket No. 60790-720.601response or contraindication to anti-TNFa, anti-IL-12 / 23 or anti-IL-23.

[0214] Patients exhibited clear evidence of active disease, and the population was mostly biologic naive population. The intention-to-treat (ITT) population of 22 patients was randomized with 17 patients completing the Phase lb study and 5 patients discontinued from the study, including 1 patient discontinued after 5 days of dosing QD, 1 discontinued patient had evidence of improvement BID, and 3 treatment failures BID with 2 patients having significant disease burden at baseline (Table 4). For the ITT population, missing data was addressed with Last Observation Carried Forward (LOCF) methodology.Table 4: Patient Population Summary of Phase lb for TNF-alpha / IL-23 dual inhibitor.TRF -alpha? iL-23 dual aihibitor lEfflrmtor W Ktalg® aww Bielists n s&s©WSGR Docket No. 60790-720.601Table 5: Patient Population Characteristics of Phase lb for TNF-alpha / IL-23 dual inhibitor.TNF-Btphd? S.-23 Placebo dual tatabitw dual inftSwtor Total I QO(N = 71 i 81Omn BID (N=9 } {N=22} I Age, years 53 (17) 49 (18) 49 (16) 50 (16) Sex, female 3 (50) 2 (29} 7 (78) 12 (55) Extermve Disaase (Psnccljt'ss) 2 {33} 2 (29) 1 (11} 5 {23) Full Mayo score 8 (6, 9) 9 (6, 10) 7 (6, 11 > 8 {6, 11} Modified Mayo score 6 (4f7) 7 (5, 8) 5 (4, 9) 6 (4, 9) Number (®f«) patients with 1 (17) 6 (85} 5 (56) 12 (55) FCAL, median 453.5 (69, 0001) 1559.0 (322, 3893) 1302.0 (12, 5403) 1226.0 (12, 6001) CRP, median 2.90 (2.9, 157.8) 7.30 (5.3, 56.5} 5.80 (2.9, 38.6} 5 60 (2.9, 157.8) Prior sise of biologies -3 (0) 2 (29)10 (0) 2 (9) Concoirnstant UC rneds at baseline None 0 (O G) 1 (14) _ 1 ( 11} _ 2 (9) _ CBi"ticosteroids 1 (17) 1 { 14} 2 {22) 4 (18) AminoMiitcyiaites 5 {83) 6 (86) 8 (89? 19 (86)Table 6: Patient Population Medication History of Phase lb for TNF-alpha / IL-23 dual inhibitor.THf-a-Epha? i TNF-atphaf Rscefeo inhibitor | inhibitor Tefesf

[0215] Sample Collection and Analysis

[0216] Two sigmoidscopies were performed over the course of Part 3 of the study: one at the beginning of study to determine patient eligibility (Baseline: Visit 2) and one at the end of treatment (for completer patients; Week 6: Visit 8). At each endoscopy, 10 biopsies were collected from the most inflamed lesions / sites observed during the sigmoidoscopy.

[0217] Clinical activity after TNF-alpha / IL-23 dual inhibitor treatment across multiple UC clinical endpoints were evaluated (Table 7). Evaluation of clinical endpoints with sigmoid colon biopsies were determined using: (1) Histology for grading of disease severity. Further pharmacodynamic endpoints determined using biopsy tissue included; (2) Profiling of protein phosphorylation levels; (3) Measurement of cytokine protein levels; and (4) RNA sequencing (RNAseq) for transcriptome profiling. Baseline vs Week 6 comparisons were evaluated for all data sets. Biopsies were also assayed for the presence of TNF-alpha / IL-23 dual inhibitor, liberated-WSGR Docket No. 60790-720.601anti-TNF and liberated-anti-IL-23 monomer by ELISA and mass spectrometry.Table 7: Exemplary Phase lb Exploratory Objectives and Endpoints of TNF-alpha / IL-23 dual inhibitor.Ohjecltve: Ta assess. change from baseiiBa / propartfon of: Ewrf polniLi cfaawga from bsseSms or projptQrtron &f f espoad&ra responders iTable 8: Summary of TEAEs of TNF-alpha / IL-23 dual inhibitor.TNF-alphai TNF-alpha?IL-23 dual I IL-23 dual Placebo inhibitor inhibitor Total (N=6) 810 mg QD 810mg BID (N=22)(N=9)Total Number of TEAEs 5 2 6 13 Number of patients with >1 TEAE 3 (50) 2 i29) 5 (56) 10 (46) Number of patients with TEAEs leading to study discontinuation 0 (0) 0 (0) 2 (22} 2 (9) Number of patients with >1 SAE 0 (0) 0 (0) 1 (1.1) 1 (5) Number of patients with SAEs leading to study discontinuation 0 (0) 0 (0) 1 (ll)11 (5) Number of patients with Si TEAE by maximum severityMlid 1 (17) 1 (14) 2 (22) 4 (18) Moderate 2 (33) 1 (14) 3 (22) 5 (23) Severe 0 (0) 0 (0) 1 (11) 1 (5)Number of patients with >1 TEAE by relationship to TreatmentPossibly related 0 (0) 0 (0) 1 (li)31 (5) Nat related 5 (S3) 2 (29) 5 (55) 12 (55) 1 Worsening of UC that led to hospitalization2. Abdominal pairs or mild severityTable 9: Summary of TEAEs of TNF-alpha / IL-23 dual inhibitor.WSGR Docket No. 60790-720.601TEAEs in Treatment Period by SOC and PT Safety Analysis Set Ga stroi rctestinaJ Disorders <7 a 8 (8.0)0O (0.0) 8 2 (22.2) 2 Abdotntas: pain upper rc f& Je 6 (0.(1; S 1 0:.15 1 Colitis ukerattve 0 (03} 0 1 (11.1} 1 Infections And Infestations 1 (1&.7) 1 0 (».©} O *3 (111) 1 Nasopharyngitis rs e I (1&.7) 1 Q (5 £) 0 1 (111) 1 MetabeHsm and Nutrition Disorders « (%)e 1 (16.7) 1 W43-1 0 {M) O Hyperglycemia 0 (LUO) 0 1 (14.3) 1 3 (0,0) 0 Hyppaibuminemfe 1 1 0 (O. G) 0 Q (60) 3 Blood And Lymphatic System Disorders rs j e 3 (16.7) 1 13 (iLO) a 0 {0.0} B Ar. SFfifo n (%}e 1 (167) 1 G $ G)0 G{00} 3 General Disorders and Administration Sit® Conditions n e <3 ifi.0} it 1 fU.3- 1 IS (».«) O Peripheral swelling r< (%)e 0 (03; 0 1 (14.3) 1 Investigations n {%) *2 1 7} 1 O (0.0} Q B (0.0) 0 Prosratlc specific antigen Increased n {%) e 1:7} 3 3 (G £■} 0 Q (6.3) 3 Respiratory, Thoracic ar«d Mediastinal Disorders si 1 (16.?) 1 a (a. S3) 8 o (©.a) a Cough I (16.7) 1 G (i; 4-)0 6 {00) 5 Skim and Subcutaneous Tissue Disorders r< (%} s 0 > 0. o; 6 O {O.0) O 1 tltlp Rash 6 (0.3; S <: U 1p TEAEs-4:reatraeftt emergent adverse ever-te. SGC-vsystem organ class. PT--p?eferred term, of c drtlcipemis Ir- analysis set fi---numlier of psftkipants with^

[0218] Tissue Cytokine

[0219] A panel of 8 cytokine proteins were measured at Baseline and Week 6: IFNy, IL-17A, IL-1β, IL-6, TNFa, IL-10, IL-22, and IL-23. IL-1β, IL-6, and TNFa are general pro-inflammatory cytokines. IFNy and IL-17A are pro-inflammatory cytokines associated with Thl and Thl7 subsets of CD4 T cells. IL-10 is generally an anti-inflammatory cytokine, but can increase during inflammation to counteract increases in pro-inflammatory cytokines (i.e. levels can be context-dependent). IL-22 and IL-23 were not detected in the majority of samples and were therefore not analyzed further.

[0220] Cytokines were measured using the Meso Scale Discovery (MSD) multiplex platform (Placebo n=6, QD n=6, BID n=5) (FIGURES 34-45). Picograms (pg) of cytokine proteins per milligram (mg) of tissue (pg / mg).

[0221] Log2 fold-change of various cytokines from Baseline to Week 6 were assessed for patient groups receiving placebo, TNF-alpha / IL-23 dual inhibitor QD, and TNF-alpha / IL-23 dual inhibitor BID (FIGURES 34-39).

[0222] Cytokine expression (pg / mg values) was assessed for various cytokines and shown to vary across treatment groups (FIGURES 40-45). Tissue levels of these cytokines vary across both healthy control and both healthy control and UC patient groups (Podolski M et al., Eur J Clin Invest.2023; doi:10.1111 / eci.14070).

[0223] All cytokines increased from Baseline to Week 6 in the Placebo group (both medianWSGR Docket No. 60790-720.601and mean values). Median / mean cytokine levels stayed the same or decreased from Baseline to Week 6 in both TNF-alpha / IL-23 dual inhibitor treatment groups (QD, BID). Median (and mean) values of log2fold-change were higher in Placebo than QD or BID groups for all cytokines.

[0224] Overall, a decrease in inflammatory cytokines in biopsies in patients treated with TNF-alpha / IL-23 dual inhibitor supports clinical observations that TNF-alpha / IL-23 dual inhibitor decreased disease across multiple clinical endpoints (full Mayo score, modified Mayo score, Endoscopy score, Stool frequency and rectal bleeding scores, etc.).

[0225] Serum Cytokine

[0226] Serum samples were also collected at Baseline and Weeks 1, 2, 3, 4, and 6. A panel of cytokine proteins were measured: IFNy, IL-17A, IL-6, TNFa, IL-10, and IL-22 (FIGURES 54-60). There were minimal differences between TNF-alpha / IL-23 dual inhibitor treated groups and placebo across time for these cytokines in serum samples. This data supports a proposed tissue-localized activity of TNF-alpha / IL-23 dual inhibitor with lack of systemic immune-modulation.

[0227] RNA sequencing gene co-expression network (GCN)

[0228] RNAseq pre-processing

[0229] Following RNA extraction of tissue RNA and quality control (RIN), a cDNA library was prepared from total extracted RNA. Then, high-throughput sequencing (e.g., Illumina) of the cDNA library was performed and quality control (Q30). Transcripts were mapped to the human transcriptome and quantification was performed of count reads mapping to transcripts. For gene counts matrix development, read counts for all genes / transcripts were compiled across all samples into a counts matrix. Preliminary exploration of principal component analysis and heatmaps were used to assess data distribution. During RNAseq preliminary exploration, hierarchical clustering of samples based on the top- 100 most variable genes and PCA revealed no distinct segregation or grouping patterns, indicating minimal observable heterogeneity in the sample transcriptomic profiles from the dataset.

[0230] A GCN analysis was conducted separating genes into biological ‘modules’ (e.g., according to co-expression of and connectivity between genes). The raw values for Visit 2 (Baseline) and Visit 8, along with the change from Visit 2 (Baseline) to Visit 8, for paired samples from each patient were evaluated in GCN modules. Statistical analysis of paired samples were analyzed using non-parametric paired T test and ANOVA. Parametric statistical analysis results were overlayed assuming normal distribution.

[0231] The change from Visit 2 (Baseline) to Visit 8 for paired samples from each patient were evaluated in GCN modules, identified based on a reference dataset.

[0232] The GCN analysis workflow was run independently for RNAseq stratified by cellWSGR Docket No. 60790-720.601lineage, and RNAseq stratified by cell class. RNAseq stratified by lineage yielded a total of 6 modules (e.g., glial, B cell, cell cycle, membrane transport, hormone secretion, and acute inflammatory response). GCN analysis of RNAseq of TNF-alpha / IL-23 dual inhibitor treatment groups stratified by cell class yielded a total of 44 modules (Table 10).Table 10: Exemplary modules identified by GCN analysis of RNAseq of TNF-alpha / IL-23 dual inhibitor treatment groups stratified by cell class.tasma KM jfctoktesS pressed tsssptmeINF« •«P w SKI a csii E R re spew:<£Ifni i igiyN.? s" as ssl'jc»>onGitalS>?!>tarns,'i r > 1 s. to ’■'M? ansi; FM >Ctftcy feiS p'-ii-s\ C<; V Cy<. a';

[0233] With 56 modules originally identified with gene frequencies varying from ~50 to -750 genes, 12 modules were filtered out with no defined identity and association with TNF-alpha / IL- 23 dual inhibitor treatment groups, due to a combination of (1) absence of statistical difference between Visit 2 (Baseline) and Visit 8 in TNF-alpha / IL-23 dual inhibitor treated groups, (2) no visible association with cell lineage; or (3) no significantly enriched process.

[0234] Module-trait correlations (MTC) were performed to identify GCN module cell lineage identities. Weak to modest correlation values (r2 < 0.4) were observed. Module cellular expression (MCE) was analyzed by scoring (e.g., Ucell scoring method) single cells based on GCN module gene content. MCE outperformed MTC in identifying cell lineages and was used to infer module identity. MCE was utilized to illustrate the expression of module gene content at a single cell level for every lineage.

[0235] GCN modules were scored with Hallmark gene sets. Modules (n=28) were found to be significantly associated with the Hallmark gene sets. Additional analysis was performed by scoring GCN modules with Gene Ontology (GO) Biological Processes (BP) gene sets, yielding a total of 38 modules with enriched gene sets.

[0236] Gene Module Changes

[0237] Changes in expression in gene modules per treatment were assessed between Baseline (Visit 2) and end of TNF-alpha / IL-23 dual inhibitor treatment Week 6 (Visit 8). Hub score analysisWSGR Docket No. 60790-720.601was also determined as a measure of how important or influential a particular gene (or node) was for the stability of the module.

[0238] Further additional analysis with enrichment with IL-23 gene set was performed for a single set of 37 genes, including IL12RB1, IL17A, and IL23A.

[0239] Consensus Gene Set Changes

[0240] Three publicly available consensus gene sets representing TNF-alpha (2 gene sets: “Hallmark_TNFA_ Signaling Via NFKB”, “REACTOME_TNF_ SIGNALING”) or IL-23 (1 gene set: “PID_IL23_Pathway”) pathway activity signatures were identified from the molecular signatures database (https: / / www.gsea-msigdb.org / gsea / msigdb). Changes in expression of each of these sets were assessed between Baseline (Visit 2) and end of TNF-alpha / IL-23 dual inhibitor treatment Week 6 (Visit 8).Example 2: Human clinical study of orally delivered bispecific anti-TNF-alpha / anti-IL-23 domain antibody

[0241] Healthy adult subjects and patients with mild to severe ulcerative colitis (UC) were enrolled in randomized, double-blinded, placebo-controlled Phase 1 clinical studies to evaluate an orally delivered TNF-alpha / IL-23 dual inhibitor, as described in Example 1.

[0242] Various treatment effects were observed showing clinical improvement in patient groups administered at a total daily dose of 1620 mg TNF-alpha / IL-23 dual inhibitor (SEQ ID NO: 201) BID (FIGURES 23-33 and TABLE 9)

[0243] Improvement in UC symptoms

[0244] Completer population showed rapid and robust improvement in UC symptoms (FIGURES 23-25 and 32-33). An improvement in UC-100 score was observed (FIGURE 24) with a reduction in mean combined stool frequency subscore (SFS) and rectal bleeding subscore (RBS), between Visit 2 (Baseline) and Visit 8 of patient groups receiving of TNF-alpha / IL-23 dual inhibitor BID. (FIGURE 23). Additionally, an improvement in symptomatic remission (e.g., SFS=0 or 1 without worsening and RBS=0) was observed with high dose TNF-alpha / IL-23 dual inhibitor BID (FIGURES 32-33 and 52-53).

[0245] Endoscopic Improvement

[0246] Improvement in endoscopic response was observed in both TNF-alpha / IL-23 dual inhibitor treatment groups with a 40% response rate in the completer population and no worsening of endoscopic response was observed with high dose TNF-alpha / IL-23 dual inhibitor (FIGURE 26-28)

[0247] Mayo Score Clinical ResponseWSGR Docket No. 60790-720.601

[0248] Improvement in Mayo score and Modified Mayo Score were observed with high dose TNF-alpha / IL-23 dual inhibitor BID (FIGURE 30-33).

[0249] Histological improvement was observed with high dose TNF-alpha / IL-23 dual inhibitor BID following Robarts Histopathology Index (RHI) analysis (FIGURE 29 and 51).Example 3: RNAseq gene co-expression network analysis for human clinical study of orally delivered anti-TNF-alpha / anti-IL-23 dual inhibitor - Immune-associated modules

[0250] Healthy adult subj ects and patients with mild to severe UC were enrolled in randomized, double-blinded, placebo-controlled Phase 1 clinical studies to evaluate an orally delivered TNF-alpha / IL-23 dual inhibitor, as described in Example 1. The change from Visit 2 (Baseline) to Visit 8 for paired samples from each patient were evaluated in GCN modules, identified based on a single-cell RNA reference dataset, as described in Example 1.

[0251] Various immune-associated modules (e.g., aP T cells activation, B cells) were assessed as shown as described in FIGURES 1-6 and 11-14.

[0252] Immune-associated modules: of T cells activation

[0253] Module analysis associated with aP T cells activation for TNF-alpha / IL-23 dual inhibitor treatment groups were performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with aP T cells activation (e.g., G0:0046635 - positive regulation of alpha-beta T cell activation genes labelled).

[0254] FIGURE 1 shows differences observed for Log2FC of module expression levels (GVSA) of modules associated with aP T cells activation in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo or TNF-alpha / IL-23 dual inhibitor treatment groups.

[0255] Paired samples from patient groups receiving TNF-alpha / IL-23 dual inhibitor BID showed a significant decrease in expression (GVSA) of modules associated with aP T cells activation from Visit 2 (Baseline) to Visit 8 (FIGURE 2). Since aP T cells are pro-inflammatory, the decrease in expression of modules associated with aP T cells activation may be indicative of a less inflammatory environment.

[0256] Immune-associated modules: myeloid inflammatory TNF signaling

[0257] Modules associated with myeloid and myeloid inflamed lineages were assessed for expression by a particular cell lineage. Additional analysis was performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with inflammation, scoring GCN modules with enriched Hallmark gene sets associated with TNF signaling, and scoring GCN modules with enriched IL-23 signature (e.g., genes overlapping with TNFWSGR Docket No. 60790-720.601signaling).

[0258] Significant differences were observed for Log2FC of module expression levels (GVSA) of modules associated with myeloid inflammatory TNF signaling in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving TNF-alpha / IL-23 dual inhibitor QD compared to placebo (FIGURE 3). A significant increase was observed for expression (GVSA) of modules associated with myeloid inflammatory TNF signaling in paired samples from Visit 2 (Baseline) to Visit 8 in paired samples of patient groups receiving placebo (FIGURE 4) FIGURE 4 also shows differences observed for expression (GVSA) of modules associated with myeloid inflammatory TNF signaling from Visit 2 (Baseline) to Visit 8 in paired samples from patient groups receiving TNF-alpha / IL-23 dual inhibitor.

[0259] Immune-associated modules: inflammatory response

[0260] Modules associated with inflammatory response were assessed for expression by a particular cell lineage. A modest increased expression was observed for mature and immature epithelial cells. Additional analysis was performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with inflammatory processes.

[0261] Significant differences were observed for Log2FC of module expression levels (GVSA) of modules associated with inflammatory response in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving TNF-alpha / IL-23 dual inhibitor QD or BID compared to placebo (FIGURE 5). FIGURE 6 shows differences observed for expression (GVSA) of modules associated with inflammatory response from Visit 2 (Baseline) to Visit 8 in in paired samples from patient groups receiving placebo or TNF-alpha / IL-23 dual inhibitor.

[0262] Immune-associated modules: B cell

[0263] There is an increase in (mainly naive) B-cells in UC (and IBD in general) intestinal tissue that appears to drive inflammation and hinder mucosal healing. Anti-Microbial IgG (as well as IgA) is also increased in IBD conditions, as are auto-antibodies against human proteins.

[0264] The transcriptomes of healthy and adult UC colonic tissue revealed that the percentage of naive B cells was increased, and % memory B cells was decreased in inflamed UC tissue. (Penrose et al., 2021 Sci Rep 11, 9010). B cells accumulate in damaged areas of the colon during mucosal healing (MH). However these cells hinder the interaction between epithelium and stroma, limiting MH. (Frede et al. 2022. Immunity. 55. 2336-).

[0265] Modules associated with inflammatory response were assessed for expression by a particular cell lineage. An increased expression was observed for B cells. Additional analysis was performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with B cell activation and proliferation (e.g., GO: 0042113 - B cell activation genesWSGR Docket No. 60790-720.601labelled).

[0266] FIGURE 11 shows differences observed for Log2FC of module expression levels (GVSA) of modules associated with B cells in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo or TNF-alpha / IL-23 dual inhibitor. FIGURE 12 shows a significant decrease in expression (GVSA) of modules associated with B cells from Visit 2 (Baseline) to Visit 8 in paired samples of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID.

[0267] Immune-associated modules: Plasma cell misfolded protein response

[0268] There are conflicting reports about the involvement of the misfolded protein response, or unfolded protein response (UPR), in IBD and UC. UPR may be important for intestinal homeostasis and increased markers of UPR are present in mature / maturing epithelial cell populations that repopulate the intact intestinal barrier (Heijmans et al. 2013 Cell Rep.3(4).1128-39,. Garcia et al 2023 Biomedicines. 11(7):2066., Coleman & Haller (2019). Front Immunol.10:2825). UPR is also increased in inflamed UC tissue and may denote stress.

[0269] Modules associated with unfolded / misfolded protein response were assessed for expression by a particular cell lineage. An increased expression was observed for plasma cells. Additional analysis was performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with ER stress and unfolded protein response (UPR) signaling (e.g., GO:0006986 - response to unfolded protein genes labelled). UPR is a key player in IBD pathophysiology.

[0270] FIGURE 13 shows differences observed for Log2FC of module expression levels (GVSA) of modules associated with plasma cell misfolded protein response in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo or TNF-alpha / IL-23 dual inhibitor.

[0271] FIGURE 14 shows a significant increase in expression (GVSA) of modules associated with plasma cell misfolded protein response from Visit 2 (Baseline) to Visit 8 in paired samples of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID.

[0272] Clinical improvement in TNF-alpha / IL-23 dual inhibitor-treated patients, and increases in the stem cell maintenance and endocrine cell differentiation suggest that the UPR is contributing to the restoration of tissue homeostasis.Example 4: RNAseq gene co-expression network analysis for human clinical study of orally delivered anti-TNF-alpha / anti-IL-23 dual inhibitor - Epithelial-associated modules

[0273] Healthy adult subjects and patients with mild to severe ulcerative colitis (UC) wereWSGR Docket No. 60790-720.601enrolled in randomized, double-blinded, placebo-controlled Phase 1 clinical studies to evaluate an orally delivered TNF-alpha / IL-23 dual inhibitor, as described in Example 1. The change from Visit 2 (Baseline) to Visit 8 for paired samples from each patient were evaluated in gene co-expression network (GCN) modules, identified based on a single-cell RNA reference dataset, as described in Example 1.

[0274] Various epithelial-associated modules (e.g., stem cell maintenance and Myc signaling) were assessed as shown as described in FIGURES 7-10.

[0275] Epithelial-associated modules: stem cell maintenance

[0276] Stems cells are an important cell type responsible for differentiating into the other epithelial cell populations of the gut barrier (Luo 2022 Int J Biol Sci). An increase in pathways involved in stem cell maintenance could signify a stabilization of the stem cell population and a return of gut barrier homeostasis.

[0277] Module analysis associated with intestinal stem cell maintenance for TNF-alpha / IL-23 dual inhibitor treatment groups was performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with stem cell population and cell number (e.g., GO:0019827 - stem cell population maintenance genes labelled).

[0278] FIGURE 7 shows differences observed for Log2FC of module expression levels (GVSA) of modules associated with intestinal stem cell maintenance in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo or TNF-alpha / IL-23 dual inhibitor.

[0279] FIGURE 8 shows differences observed for expression (GVSA) of modules associated with intestinal stem cell maintenance from Visit 2 (Baseline) and Visit 8 in paired samples of patient groups receiving placebo or TNF-alpha / IL-23 dual inhibitor QD. Paired samples of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID from Visit 2 (Baseline) to Visit 8 showed a significant increase in expression (GVSA) of modules associated with intestinal stem cell maintenance (FIGURE 8).

[0280] Epithelial-associated modules: Myc signaling

[0281] Myc signaling is important to restore the population of intestinal stem cells (ISCs) upon damage, by mechanisms including signaling through the canonical Wnt pathway. Myc induces the expansion of Intestinal stem cells / Intestinal progenitor cells during intestinal regeneration (Kim et al. 2018. Dev Cell. 44(5):582). c-Myc is required for the formation of intestinal crypts (Bettess et al 2005. Mol Cell Biol.25(17):7868).

[0282] Increased expression in epithelial stem cells was observed in TNF-alpha / IL-23 dual inhibitor treatment groups, an effect also observed in inflammatory fibroblast and post-capillaryWSGR Docket No. 60790-720.601venules. Additional analysis was performed by scoring GCN modules with enriched Hallmark gene set associated with Myc signaling.

[0283] FIGURE 9 shows differences observed for Log2FC of module expression levels (GVSA) of modules associated with Myc signaling in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo or TNF-alpha / IL-23 dual inhibitor.

[0284] Paired samples of patient groups receiving placebo from Visit 2 (Baseline) to Visit 8 showed a significant decrease in expression (GVSA) of modules associated with Myc signaling (FIGURE 10). Paired samples of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID from Visit 2 (Baseline) to Visit 8 showed a significant increase in expression (GVSA) of modules associated with Myc signaling (FIGURE 10).

[0285] Taken together, the treatment with TNF-alpha / IL-23 dual inhibitor BID was followed by an increase in the expression (GVSA) of modules associated with intestinal stem cell maintenance (FIGURE 8 and 10). For some modules, the expression range of the placebo group is different from the treatment groups.Example 5: RNAseq gene co-expression network analysis for human clinical study of orally delivered anti-TNF-alpha / anti-IL-23 dual inhibitor - stratified by class

[0286] Healthy adult subjects and patients with mild to severe ulcerative colitis (UC) were enrolled in randomized, double-blinded, placebo-controlled Phase 1 clinical studies to evaluate an orally delivered TNF-alpha / IL-23 dual inhibitor, as described in Example 1. The gene coexpression network (GCN) analysis workflow was run independently for RNAseq stratified cell class, which yielded a total of 44 modules, as described in Example 1. The change from Visit 2 (Baseline) to Visit 8 for paired samples from each patient were evaluated in gene co-expression network (GCN) modules, identified based on a single-cell RNA reference dataset, as described in Example 1.

[0287] Various modules stratified by class were assessed as shown as described in FIGURES 15-20. Significant differences between treatment groups were observed for exemplary modules strati (e.g., enteroendocrine cell differentiation, ribosome and rRNA processing).

[0288] Endocrine cell differentiation

[0289] No visible increase was observed by a particular cell lineage. Additional analysis was performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with enteroendocrine cell differentiation (e.g., GO: 0035883 - enteroendocrine cell differentiation genes labelled).

[0290] Significant differences were observed for Log2FC of module expression levelsWSGR Docket No. 60790-720.601(GVSA) of modules associated with endocrine cell differentiation in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID compared to placebo (FIGURE 15). FIGURE 16 shows differences observed for expression (GVSA) of modules associated with endocrine cell differentiation from Visit 2 (Baseline) and Visit 8 in paired samples from patient groups receiving TNF-alpha / IL-23 dual inhibitor BID.

[0291] Ribosome and rRNA processing

[0292] No visible increase was observed by a particular cell lineage. Additional analysis was performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with ribosome and rRNA processing (e.g., GO:0032790 - ribosome disassembly genes labelled).

[0293] FIGURE 17 shows differences observed for Log2FC of module expression levels (GVSA) of modules associated with ribosome and rRNA processing in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving placebo or TNF-alpha / IL-23 dual inhibitor. Paired samples from patient groups receiving TNF-alpha / IL-23 dual inhibitor BID from Visit 2 (Baseline) to Visit 8 showed a significant increase in expression (GVSA) for modules associated with ribosome and rRNA processing (FIGURE 18).

[0294] Epithelial mature immature

[0295] Analysis was performed by scoring GCN modules with enriched GO: BP and Hallmark gene sets with biological processes associated with epithelial mature immature, however, a trend was observed for enrichment with GO: BP associated with cell migration (e.g., GO:0030335 - positive regulation of cell migration genes labelled).

[0296] A significant decrease in Log2FC of module expression levels (GVSA) of modules associated with epithelial mature immature was observed in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID compared to placebo (FIGURE 19). Paired samples from patient groups receiving placebo from Visit 2 (Baseline) to Visit 8 showed a significant increase in expression (GVSA) of modules associated with epithelial mature immature (FIGURE 20). Paired samples from patient groups receiving TNF-alpha / IL-23 dual inhibitor BID from Visit 2 (Baseline) to Visit 8 showed a significant decrease in expression (GVSA) for modules associated with epithelial mature immature (FIGURE 20)Example 6: RNAseq gene co-expression network analysis for human clinical study of orally delivered anti-TNF-alpha / anti-IL-23 dual inhibitor - stratified by lineage

[0297] Healthy adult subjects and patients with mild to severe ulcerative colitis (UC) wereWSGR Docket No. 60790-720.601enrolled in randomized, double-blinded, placebo-controlled Phase 1 clinical studies to evaluate an orally delivered TNF-alpha / IL-23 dual inhibitor, as described in Example 1. The GCN analysis workflow was run independently for RNAseq stratified by cell lineage, which yielded a total of 6 modules (e.g., glial, B cell, cell cycle, membrane transport, hormone secretion, and acute inflammatory response), as described in Example 1. The change from Visit 2 (Baseline) to Visit 8 for paired samples from each patient was evaluated in gene co-expression network (GCN) modules, identified based on a single-cell RNA reference dataset, as described in Example 1.

[0298] Various modules stratified by lineage were assessed as shown as described in FIGURE 21-22. Significant differences between treatment groups were observed for exemplary modules stratified by lineage (e.g., hormone secretion).

[0299] Hormone secretion

[0300] Module cellular expression did not indicate association with a particular cell lineage. Additional analysis was performed by scoring GCN modules with enriched GO: BP gene sets with biological processes associated with hormone secretion.

[0301] A significant increase was observed for Log2FC of module expression levels (GVSA) of modules associated with hormone secretion in paired samples between Visit 2 (Baseline) and Visit 8 of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID compared to placebo (FIGURE 21). FIGURE 22 shows differences observed for expression (GVSA) of modules associated with hormone secretion from Visit 2 (Baseline) and Visit 8 in paired samples of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID.

[0302] Taken together, the examples show that TNF-alpha / IL-23 dual inhibitor can be an orally administered for the treatment of UC.

[0303] While preferred embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention.Example 7: Human clinical study of orally delivered TNF-alpha / IL-23 dual inhibitor to UC patients show efficient cleavage into monomeric arms, sustained monomeric activity in stool, and monomer delivery to UC colonic tissue with minimal systemic exposure

[0304] Healthy adult subjects and patients with mild to severe ulcerative colitis (UC) were enrolled in randomized, double-blinded, placebo-controlled Phase 1 clinical studies to evaluate anWSGR Docket No. 60790-720.601orally delivered TNF-alpha / IL-23 dual inhibitor, as described in Example 1. The following example shows biodistribution and tissue pharmacodynamic (PK) data collected as described in Example 1 from Part 3, a Phase lb study, which was a randomized, placebo-controlled, doubleblind, double-dummy, multiple dose exploratory study.

[0305] In Part 3, patients with Mayo endoscopy score >2, rectal bleeding score >1 and stool frequency score >1 were randomized 1:1:1 to receive daily for 42 consecutive days: (1) a 810mg oral dose of TNF-alpha / IL-23 dual inhibitor administered once a day (QD) (n = 7 patients), (2) a 810mg oral dose of TNF-alpha / IL-23 dual inhibitor administered twice a day (BID) for a total oral dose of 1620mg of TNF-alpha / IL-23 dual inhibitor per day (n = 9 patients), or (3) an oral placebo administered BID (n = 6 patients) (Tables 2-9). 22 patients were randomized, and 17 patients completed the study as follows: (1) a 810mg oral dose of TNF-alpha / IL-23 dual inhibitor administered once a day (QD) (n = 6 patients), (2) a 810mg oral dose of TNF-alpha / IL-23 dual inhibitor administered twice a day (BID) for a total oral dose of 1620mg of TNF-alpha / IL-23 dual inhibitor per day (n = 5 patients), or (3) an oral placebo administered BID (n = 6 patients). Patients enrolled to receive doses of TNF-alpha / IL-23 dual inhibitor QD received TNF-alpha / IL-23 dual inhibitor for one of their doses and matching placebo capsule for their second daily dose (Table 3). The mean age of the patients was 50 years. 45% of the patients were male. The median Mayo Score (MS) was 8 (range 6-11). Most patients were advanced therapy -naive.

[0306] In the TNF-alpha / IL-23 dual inhibitor BID arm, there was strong correlation between clinical activity, tissue drug concentrations, and PD outcomes, which included reduced colonic tissue IL-1β, IL-6, TNFa, IFNy, IL-17A, and IL-10 protein levels and changes in gene expression corresponding to reduced myeloid inflammation / TNF signaling and T & B cell responses, with increased activity of intestinal regeneration (FIGURES 61-68). No notable changes were observed in measured serum cytokine protein levels. In the TNF-alpha / IL-23 dual inhibitor QD arm, there was a tendency towards favorable tissue PD responses, suggesting a higher dose or longer duration of treatment are needed for higher response (FIGURES 61-68).

[0307] Clinical Remission Observed with High Dose TNF-alpha / IL-23 dual inhibitor

[0308] Clinical remission was observed with high dose TNF-alpha / IL-23 dual inhibitor BID (FIGURE 61) as determined by MCS (full MCS Mayo score < 2 with no individual subscore >1) and modified MCS (RBS=0, SF subscore=0 or 1, with no increase in SF from baseline, endoscopy subscore=0 or 1 based on central review, and symptomatic remission, score ranges from 0 to 9, with a higher score indicating more severe disease). Higher rates of MS and mMS clinical response, and SR, and greater decreases in MS, mMS, and UC-100 score were observed in the TNF-alpha / IL-23 dual inhibitor BID arm compared to placebo.WSGR Docket No. 60790-720.601

[0309] Pharmacokinetics in fecal samples show high active fecal monomer concentrations

[0310] High fecal concentrations of active liberated-anti-TNF and liberated-anti-IL-23 monomers and low fecal concentrations of intact TNF-alpha / IL-23 dual inhibitor confirm that TNF-alpha / IL-23 dual inhibitor is efficiently cleaved and monomer activity is maintained after oral dosing in UC patients (FIGURES 62-63). TNF-alpha / IL-23 dual inhibitor QD and TNF-alpha / IL-23 dual inhibitor BID patient groups show high and similar mean and median levels of active liberated-anti-TNF and liberated-anti-IL-23 monomers throughout treatment period. The data show that there can be less variability between samples with TNF-alpha / IL-23 dual inhibitor with BID dosing. Among the per protocol (completer) population, only one patient had detectable liberated-anti-IL-23 monomer in serum two weeks after cessation of treatment.

[0311] Fecal Recovery

[0312] Stool samples were collected prior to the first dose of TNF-alpha / IL-23 dual inhibitor and at Visit 4 (week 1 post initiation of dosing), visit 5 (week 2), visit 6 (week 3), visit 7 (week 4), visit 9 (week 6) and visit 10 (follow up) using at home stool sampling kits or by providing a stool sample at the time of the visit. Stool samples were frozen at -80 °C as soon as possible after collection. Stool samples were processed for downstream analysis as described in Crowe et al., 2019. Drug Dev and Industrial Pharmacy. 45, 387-394.

[0313] Activity Assays

[0314] Antigen-specific Enzyme-linked immunoassay (ELISA) or electrochemiluminescence immunoassay (ECLIA) protocols were used to detect liberated-anti-IL-23 monomer, liberated-anti-TNF monomer or TNF-alpha / IL-23 dual inhibitor in stool samples. These assays rely on intact molecules actively engaging target cytokine(s) to produce measurable assay signal. Therefore, only intact and active molecules are detected and the assays are considered “activity” assays as described in this example. Table 11 lists the lower limit of quantification (LLOQ) for each assay. The LLOQs reflect the minimum concentration quantifiable in the 5x diluted sample, taking into account any minimum in-assay dilution required before samples were loaded onto the assay plate. The minimum limit of detection (MLD) is the minimum concentration quantifiable in neat stool prior to the 5x dilution that occurs during sample processing. Only samples from TNF-alpha / IL-23 dual inhibitor -dosed patients were analyzed. Samples from placebo-dosed patients were not analyzed. No clear assay signal was detected for any analyte (liberated-anti-IL-23 monomer, liberated-anti-TNF monomer or TNF-alpha / IL-23 dual inhibitor) in any patient stool sample collected for QD or BID group patients prior to oral TNF-alpha / IL-23 dual inhibitor dosing.

[0315] Table 11: Assay LLOQs in Fecal Samples.WSGR Docket No. 60790-720.601Target Molecule Assay Assay MLD Assay MLD Graphing LLOQ (ng / ml) (pM) MLD (pM) (ng / ml)TNF-alpha / IL-23 dual 12.5 0.0004695 0.00045962.5inhibitor (27.2 kDa)liberated-anti-TNF 80 400 0.02985 0.02931 monomer (13.4 kDa)liberated-anti-IL-23 80 400 0.02878 0.02931 monomer (13.9 kDa)

[0316] Graphing Data for Samples with < MLD Values

[0317] Dotted lines representing the MLD values of each assay were included on graphs when appropriate. Because the MLD of the liberated-anti-IL-23 monomer and liberated-anti-TNF monomer assays are close (0.02985 and 0.02878, respectively), they were averaged and shown as a single line on the graphs for ease of visualization.

[0318] To visualize individual samples that were < MLD for each assay, these samples were assigned a value for graphing purposes only (Table 12). The assigned value for each molecule is unique so the symbols for each < MLD sample for each molecule do not overlap with < MLD symbols for the other molecules on the graph.

[0319] Table 12: Assigned Values for Samples < MLD.Molecule Assigned Value (pM)TNF-alpha / IL-23 0.0001dual inhibitorliberated-anti-TNF 0.00005monomerliberated-anti-IL- 0.000223 monomer

[0320] Monomer concentration in colonic tissue increases with oral dose by ELISA-based assay

[0321] Colonic biopsy samples were collected and processed to evaluate colonic tissue activity using three ELISA-based assays that were developed bespoke for measurement of drug in biopsy tissue. Specific ELISA detection for TNF-alpha / IL-23 dual inhibitor, liberated-anti-IL-23 monomer and liberated-anti-TNF monomer incorporated cytokine ligand (TNF-alpha or IL-23) in the detection format to account for the effect of endogenous cytokine present in biopsies. Higher levels of active liberated-anti-TNF and liberated-anti-IL-23 monomers were detected in colonicWSGR Docket No. 60790-720.601tissue from BID patients than QD patients (FIGURE 64). TNF-alpha / IL-23 dual inhibitor was detected in a single sample, at ~15x lower levels than liberated-anti-TNF.

[0322] Monomer concentration in colonic tissue increases with oral dose by Mass Spectrometry

[0323] Colonic biopsy samples were collected and processed to evaluate colonic tissue concentration using mass spectrometry. Higher levels of active liberated-anti-TNF and liberated-anti-IL-23 monomers were detected in colonic tissue from TNF-alpha / IL-23 dual inhibitor BID patients than TNF-alpha / IL-23 dual inhibitor QD patients (FIGURE 65). TNF-alpha / IL-23 dual inhibitor BID patients exhibited 20-35 fold higher median levels of liberated-anti-TNF and liberated-anti-IL-23 monomers in colonic tissue compared to TNF-alpha / IL-23 dual inhibitor QD patients.

[0324] Active monomer colonic tissue levels as detected by ELISA and Mass Spectrometry

[0325] The measured colonic tissue concentrations of liberated-anti-TNF monomer by ELISA and mass spectrometry were similar (FIGURE 66). Tissue concentrations in FIGURE 66 were plotted as liberated-anti-TNF monomer or liberated-anti-IL-23 monomer per mg of biopsy protein as measured by BCA protein assay. Liberated-anti-IL-23 monomer ELISA detection was not achievable in the 0.001-0.1 pmol / mg range due to assay sensitivity.

[0326] Lower inflammatory tissue cytokines and T-cell associated tissue cytokines detected after treatment as measured by gene expression

[0327] RNA was extracted from ulcerative colitis biopsies and gene expression was analyzed via RNA-seq as described in Example 1. Lower inflammatory tissue cytokine gene (IL1B, IL-6, and TNF) expression was observed after TNF-alpha / IL-23 dual inhibitor treatment (FIGURE 67). Lower T-cell associated tissue cytokine gene (IL12B, IL23 A, IL17A, IL22, IFNG, IL10) expression was observed after TNF-alpha / IL-23 dual inhibitor treatment (FIGURE 68). Log2FC were calculated based on comparisons between Baseline and Week 6.

[0328] Part 3 conclusions

[0329] High and sustained levels of active liberated-anti-TNF and liberated-anti-IL-23 monomers in feces confirmed that TNF-alpha / IL-23 dual inhibitor is efficiently cleaved, and activity is maintained after oral dosing in UC patients. Levels of liberated-anti-TNF and liberated-anti-IL-23 monomers in sigmoid colonic tissue increased with oral dose, with minimal systemic exposure. In the BID arm, TNF-alpha / IL-23 dual inhibitor demonstrated consistent activity across multiple clinical and tissue PD endpoints in patients with UC.Example 8: RNAseq consensus gene set analysis for human clinical study of orally deliveredWSGR Docket No. 60790-720.601anti-TNF-alpha / anti-IL-23 dual inhibitor

[0330] Expression levels of consensus gene sets representing TNF-alpha (2 gene sets:“Hallmark_TNFA_ Signaling Via NFKB”, “REACTOME_TNF_ SIGNALING”) or IL-23 (1 gene set: “PID_IL23_Pathway”) pathways were examined. Reductions in the expression of these canonical gene sets for both TNF and IL-23 are indicative of target engagement and neutralization of both TNF and IL-23 activity in intestinal tissues. Significant decreases were observed for Log2FC of consensus gene sets representing TNF-alpha (2 gene sets:“Hallmark_TNFA_ Signaling_Via_NFKB”, “REACTOME_TNF_ SIGNALING”) pathway activity expressions levels (GVSA) between Visit 2 (Baseline) and Visit 8 of patient groups receiving TNF-alpha / IL-23 dual inhibitor BID compared to placebo. A Significant decrease was observed for Log2FC of consensus gene sets representing IL-23 (“PID_IL23_Pathway”) pathway activity expressions levels (GVSA) between Visit 2 (Baseline) and Visit 8 of patient groups receiving TNF-alpha / IL-23 dual inhibitor QD compared to placebo (FIGURE 69).Example 9: Safety, tolerability, and efficacy of TNF-alpha / IL-23 dual inhibitor.

[0331] The safety, tolerability, and efficacy of a TNF-alpha / IL-23 dual inhibitor is assessed following 12 weeks of induction therapy compared to a placebo (FIGURE 70). Eligible subjects include advanced therapy (AV)-naive and AV-experienced patients. A screening endoscopy is performed 0-5 weeks prior to randomization of subjects. Subjects are stratified by concomitant corticosteroid at randomization, and AT failure and randomized 1:1:1 to receive TNF-alpha / IL-23 dual inhibitor Dose 1 (low dose, twice daily), TNF-alpha / IL-23 dual inhibitor Dose 2 (high dose, twice daily), or placebo. After 12 weeks, an endoscopy is performed, and the week 12 primary endpoint (clinical remission) is assessed before subjects proceed to the open label maintenance phase of the trial.

[0332] In the open label maintenance phase of the trial, long-term safety, tolerability, and efficacy of a TNF-alpha / IL-23 dual inhibitor is evaluated. Subjects deemed responsive to the TNF-alpha / IL-23 dual inhibitor at week 12 are randomized 1:1 and administered TNF-alpha / IL-23 dual inhibitor once daily or twice daily for 38 weeks. At study conclusion, an endoscopy is performed, and subjects are evaluated for clinical remission. Subjects deemed non-responsive to the initial 12-week TNF-alpha / IL-23 dual inhibitor are administered high-dose TNF-alpha / IL-23 dual inhibitor twice daily for 12 weeks. Subjects who are responsive to the additional 12-week treatment regimen are then randomized 1:1 and administered TNF-alpha / IL-23 dual inhibitor once daily or twice daily.

[0333] While preferred embodiments of the present invention have been shown andWSGR Docket No. 60790-720.601described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention.

[0334] All publications, patent applications, issued patents, and other documents referred to in this specification are herein incorporated by reference as if each individual publication, patent application, issued patent, or other document was specifically and individually indicated to be incorporated by reference in its entirety. Definitions that are contained in text incorporated by reference are excluded to the extent that they contradict definitions in this disclosure.

Claims

WSGR Docket No. 60790-720.601CLAIMS WHAT IS CLAIMED IS:

1. A method of treating an individual afflicted with moderate to severe ulcerative colitis, the method comprising: administering a local inhibitor of interleukin-23 (IL-23) and tumor necrosis factor alpha (TNF-alpha) to an intestine of the individual.

2. The method of claim 1, wherein the individual is afflicted with moderate ulcerative colitis.

3. The method of claim 1, wherein the individual is afflicted with severe ulcerative colitis.

4. The method of claim 1, wherein the treating achieves a clinical response.

5. The method of claim 1, wherein the treating achieves symptomatic remission.

6. The method of claim 1, wherein the treating achieves histologic remission.

7. The method of claim 1, wherein the treating achieves remission by a Modified Mayo score or a Full Mayo score.

8. A method of inducing intestinal stem cell maintenance and / or proliferation in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

9. The method of claim 8, wherein the method induces intestinal stem cell maintenance.

10. The method of claim 9, wherein the method induces intestinal stem cell proliferation.

11. A method of altering plasma cell and misfolded protein response in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

12. A method of altering alpha beta T cell activation in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

13. A method of altering myeloid inflammatory signaling in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.WSGR Docket No. 60790-720.60114. A method of altering TNF signaling in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

15. A method of altering inflammatory response in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

16. A method of altering ribosome and rRNA processing in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

17. A method of altering mature epithelial cells in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

18. A method of altering immature epithelial cells in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

19. A method of inducing a reduction in the number of B-cells in an intestinal tissue of an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL- 23 and TNF-alpha to an intestine of the individual.

20. A method of inducing a reduction in the number of naive B-cells in the intestinal tissues of an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL- 23 and TNF-alpha to an intestine of the individual.

21. A method of increasing Myc signaling in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

22. A method of increasing endocrine cell differentiation in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

23. The method of claim 22, wherein the method induces enteroendocrine cell differentiation.WSGR Docket No. 60790-720.60124. A method of increasing hormone secretion in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

25. A method of reducing expression of consensus gene sets representing TNF-alpha or IL-23 pathways in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

26. A method of reducing a Robarts Histopathology Index (RHI) in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

27. The method of claim 26, wherein the RHI is reduced by 6 or more compared to baseline.

28. The method of claim 26, wherein the RHI is reduced by 7 or more compared to baseline.

29. The method of claim 26, wherein the RHI is reduced by 8 or more compared to baseline.

30. The method of any one of claims 27 to 29, wherein baseline comprises the RHI of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha.

31. The method of claim 26, wherein the RHI is reduced to 7 or less.

32. The method of claim 26, wherein the RHI is reduced to 6 or less.

33. The method of claim 26, wherein the RHI comprises a subscore of 0.

34. The method of claim 33, wherein a subscore of 0 is for lamina propria neutrophils and / or neutrophils in epithelium.

35. The method of claim 26, wherein the intestine of the individual has an absence of neutrophils from the mucosa.

36. The method of claim 26, wherein the intestine of the individual has an absence of basal plasmacytosis.

37. The method of claim 36, wherein the intestine of the individual further comprises a presence of plasma cells between the base of the crypts and the muscularis mucosae in the colon.WSGR Docket No. 60790-720.60138. A method of reducing a Modified Mayo score in an individual afflicted with ulcerative colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

39. The method of claim 38, wherein the Modified Mayo score is reduced by 2 or more compared to baseline.

40. The method of claim 38, wherein the Modified Mayo score is reduced by 3 or more compared to baseline.

41. The method of claim 38, wherein the Modified Mayo score is reduced by 4 or more compared to baseline.

42. The method of claim 38, wherein the Modified Mayo score is reduced by 5 or more compared to baseline.

43. The method of any one of claims 39 to 42, wherein baseline comprises the Modified Mayo score of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha.

44. The method of claim 38, wherein the Modified Mayo score is reduced to 2 or less.

45. The method of claim 38, wherein the Modified Mayo score is reduced to 1 or less.

46. A method of reducing a Full Mayo score in an individual afflicted with ulcerative colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

47. The method of claim 46, wherein the Full Mayo score is reduced by 2 or more compared to baseline.

48. The method of claim 46, wherein the Full Mayo score is reduced by 3 or more compared to baseline.

49. The method of claim 46, wherein the Full Mayo score is reduced by 4 or more compared to baseline.

50. The method of any one of claims 47 to 49, wherein baseline comprises the Modified Mayo score of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha.WSGR Docket No. 60790-720.60151. The method of claim 46, wherein the Full Mayo score is reduced to 2 or less, and no individual sub score is greater than 1.

52. The method of claim 46, wherein the Full Mayo score is reduced to 1 or less.

53. A method of reducing a UC-100 score in an individual afflicted with ulcerative colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

54. The method of claim 53, wherein the UC-100 score is reduced by 10 or more compared to baseline.

55. The method of claim 53, wherein the UC-100 score is reduced by 20 or more compared to baseline.

56. The method of claim 53, wherein the UC-100 score is reduced by 30 or more compared to baseline.

57. The method of claim 53, wherein the UC-100 score is reduced by 40 or more compared to baseline.

58. The method of any one of claims 54 to 57, wherein baseline comprises the UC-100 score of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha.

59. The method of claim 53, wherein the UC-100 score is reduced to 25 or less.

60. The method of claim 53, wherein the UC-100 score is reduced to 10 or less.

61. A method of inducing histological improvement in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

62. The method of claim 61, wherein the administering reduces neutrophils in the epithelium of the intestine of the individual to about less than 5%.

63. The method of claim 61, wherein the histological improvement is selected from the group consisting of: no crypt destruction in the intestine of the individual, no erosions in the intestine of the individual, no ulcerations in the intestine of the individual, and no granulation tissue in the intestine of the individual.WSGR Docket No. 60790-720.60164. A method of inducing endoscopic response in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

65. The method of claim 64, further comprising reducing a Mayo endoscopic score in the individual.

66. The method of claim 65, wherein the Mayo endoscopic score is reduced by 1 or more compared to baseline.

67. The method of claim 66, wherein baseline comprises the Mayo endoscopic score of the individual prior to administering the local inhibitor of IL-23 and TNF-alpha.

68. The method of claim 65, wherein the Mayo endoscopic score is 0 or 1.

69. A method of inducing mucosal regeneration in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

70. A method of inducing symptomatic remission in an individual afflicted with colitis, the method comprising: administering a local inhibitor of IL-23 and TNF-alpha to an intestine of the individual.

71. The method of claim 70, further comprising reducing a symptom stool frequency subscore (SFS) in the individual.

72. The method of claim 70, wherein the SFS score is reduced by 1 or more compared to baseline.

73. The method of claim 70, wherein the SFS score is reduced by 2 or more compared to baseline.

74. The method of claim 70, wherein the SFS score is reduced by 3 or more compared to baseline.

75. The method of claim 70, further comprising reducing a rectal bleeding subscore (RBS) in the individual.WSGR Docket No. 60790-720.60176. The method of claim 75, wherein the RBS score is reduced by 1 or more compared to baseline.

77. The method of claim 75, wherein the RBS score is reduced by 2 or more compared to baseline.

78. The method of any one of claims 8 to 37 or 61 to 77, wherein the colitis is an inflammatory bowel disease.

79. The method of claim 78, wherein the inflammatory bowel disease is Crohn’s disease or ulcerative colitis.

80. The method of any one of claims 1 to 79, wherein the local inhibitor is an oral inhibitor.

81. The method of any one of claims 1 to 80, wherein the individual is refractory to a previous treatment for colitis or ulcerative colitis.

82. The method of any one of claims 1 to 81, wherein in a serum cytokine level of the individual is not altered by administering the local inhibitor of IL-23 and TNF-alpha to the individual.

83. The method of claim 82, wherein the cytokine is selected from the list consisting of: IFNy, IL- 17 A, IL-1β, IL-6, TNFa, IL- 10, IL-22, and combinations thereof.

84. The method of any one of claims 1 to 83, wherein in a tissue cytokine level of the individual is altered by administering the local inhibitor of IL-23 and TNF-alpha to the individual.

85. The method of claim 84, wherein the cytokine is selected from the list consisting of: IFNy, IL-17A, IL-1β, IL-6, TNFa, IL-10, IL-22, and combinations thereof.

86. The method of claim 84, wherein in a tissue is selected from the group consisting of intestinal tissue, colonic tissue, rectal tissue, and esophageal tissue.

87. The method of any one of claims 1 to 86, wherein the local inhibitor of IL-23 and TNF-alpha is dosed twice daily.

88. The method of any one of claims 1 to 87, wherein the local inhibitor of IL-23 and TNF-alpha is dosed twice daily for 1, 2, 4, 5, 6, 8, 10, or 12 weeks.WSGR Docket No. 60790-720.60189. The method of any one of claims 1 to 86, wherein the local inhibitor of IL-23 and TNF-alpha is dosed once daily.

90. The method of any one of claims 1 to 86 or 89, wherein the local inhibitor of IL-23 and TNF- alpha is dosed once daily for 1, 2, 4, 5, 6, 8, 10, or 12 weeks.

91. The method of any one of claims 1 to 90, wherein the local inhibitor of IL-23 and TNF-alpha comprises a multispecific polypeptide comprising a first antigen binding domain that binds to TNF-alpha and a second antigen binding domain that binds to IL-23.

92. The method of claim 91, wherein the first antigen binding domain that binds to TNF-alpha comprises a VHH.

93. The method of claim 91 or 92, wherein the second antigen binding domain that binds to IL- 23 comprises a VHH.

94. The method of any one of claims 91 to 93, wherein the first antigen binding domain that binds to TNF-alpha and the second antigen binding domain that binds to IL-23 are coupled by a linker.

95. The method of claim 94, wherein the linker is a protease cleavable linker.

96. The method of claim 94, wherein the linker is a trypsin cleavable linker.

97. The method of claim 94 to 96, wherein the linker is a polypeptide linker.

98. The method of any one of claims 94 to 97, wherein the linker is a polypeptide linker comprising an amino acid sequence as set forth in SEQ ID NO: 302.

99. The method of any one of claims 91 to 98, wherein the first antigen binding domain that binds to TNF-alpha comprises:a) a complementarity determining region 1 (CDR1) as set forth in SEQ ID NOs: 101, 107, 108, or 119;b) a complementarity determining region 2 (CDR2) as set forth in SEQ ID NOs: 102, 109 to 117, or 120; and / orWSGR Docket No. 60790-720.601c) a complementarity determining region 3 (CDR3) as set forth in SEQ ID NOs: 103 to 106 or 121 to 124.

100. The method of claim 99, wherein the first antigen binding domain that binds to TNF- alpha comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 118 or 125.

101. The method of claim 100, wherein the first antigen binding domain that binds to TNF- alpha comprises an amino acid sequence that is identical to any one of SEQ ID NOs: 118 or 125.

102. The method of any one of claims 91 to 101, wherein the second antigen binding domain that binds to IL-23 comprises:a) a complementarity determining region 1 (CDR1) as set forth in SEQ ID NOs: 1, 4, or 7;b) a complementarity determining region 2 (CDR2) as set forth in SEQ ID NOs: 2, 5, or 8; and / orc) a complementarity determining region 3 (CDR3) as set forth in SEQ ID NOs: 3, 6, or 9.

103. The method of claim 102, wherein the second antigen binding domain that binds to IL-23 comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to any one of SEQ ID NOs: 10 to 12.

104. The method of claim 103, wherein the second antigen binding domain that binds to IL-23 comprises an amino acid sequence that is identical to any one of SEQ ID NOs: 10 to 12.

105. The method of any one of claims 1 to 104, wherein the local inhibitor of IL-23 and TNF- alpha comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 201.

106. The method of claim 105, wherein the local inhibitor of IL-23 and TNF-alpha comprises an amino acid sequence that is identical to SEQ ID NO: 201.WSGR Docket No. 60790-720.601107. The method of any one of claims 1 to 105, wherein the local inhibitor of IL-23 and TNF- alpha comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, 98%, 99% or 100% identical to SEQ ID NO: 202.

108. The method of claim 107, wherein the local inhibitor of IL-23 and TNF-alpha comprises an amino acid sequence that is identical to SEQ ID NO: 202.

109. The method of any one of claims 1 to 108, wherein the local inhibitor of IL-23 and TNF- alpha is administered at a total daily dose of about 600 mg to about 3200 mg.

110. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 600 mg to about 1600 mg.

111. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1200 mg to about 3200 mg.

112. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1200 mg to about 1600 mg.

113. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1800 mg to about 2400 mg.

114. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2400 mg to about 3200 mg.

115. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 600 mg.

116. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 800 mg.

117. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1200 mg.

118. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1300 mg.

119. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1400 mg.WSGR Docket No. 60790-720.601120. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 1300 mg to about 1400 mg.

121. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1360 mg.

122. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1600 mg.

123. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 1800 mg.

124. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 2000 mg to about 2800 mg.

125. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose from about 2000 mg to about 2800 mg.

126. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2040 mg.

127. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2720 mg.

128. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 2400 mg.

129. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha is administered at a total daily dose of about 3200 mg.

130. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha comprises the amino acid sequence set forth in SEQ ID NO: 201.

131. The method of claim 109, wherein the local inhibitor of IL-23 and TNF-alpha consists of the amino acid sequence set forth in SEQ ID NO: 201.

132. The method of any one of claims 1 to 109, wherein the local inhibitor of IL-23 and TNF- alpha is administered twice daily at a dose of about 600 mg to achieve a total daily dose of about 1200 mg.WSGR Docket No. 60790-720.601133. The method of any one of claims 1 to 109, wherein the local inhibitor of IL-23 and TNF- alpha is administered twice daily at a dose of about 900 mg to achieve a total daily dose of about 1800 mg.

134. The method of any one of claims 1 to 109, wherein the local inhibitor of IL-23 and TNF- alpha is administered twice daily at a dose of about 1200 mg to achieve a total daily dose of about 2400 mg.

135. The method of any one of claims 1 to 134, wherein the local inhibitor of IL-23 and TNF- alpha is administered for 8, 12 or 16 weeks followed by a maintenance dose.

136. The method of any one of claims 1 to 134, wherein the local inhibitor of IL-23 and TNF- alpha is administered for 12 weeks followed by a maintenance dose.

137. The method of claims 135 or 136, wherein the maintenance dose is from about 1300 to about 1400 milligrams.

138. The method of claims 135 or 136, wherein the maintenance dose is about 1360 milligrams.

139. The method of any one of claims 1 to 138, wherein the local inhibitor of IL-23 and TNF- alpha is administered for another period of 12 weeks if the individual is a non-responder.

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