Sumoylation as a therapeutic target in inflammatory bowel diseases and infections disorders

Abstract

The present disclosure provides methods of treating or preventing inflammatory bowel disease or sepsis in a subject by inhibiting SUMOylation in the subject.

Description

[0001] Client Ref. No. DU8225PCT Atty. Docket No. Duke-43397.601 SUMOYLATION AS A THERAPEUTIC TARGET IN INFLAMMATORY BOWEL DISEASES AND INFECTIONS DISORDERS PRIORITY STATEMENT This application claims priority to U.S. Provisional Patent Application No. 63 / 530,594, filed August 3, 2023, the entire contents of which are incorporated herein by reference. GOVERNMENT FUNDING This invention was made with government support under Federal Grant No. 1R21AT011387-01A1 awarded by the National Center for Complementary & Integrative Health (NCCIH). The government has certain rights in the invention. FIELD Provided herein are methods of treating or preventing disease, including IBD and sepsis, by inhibition of SUMOylation. BACKGROUND Inflammatory bowel diseases (IBD) are chronic, debilitating conditions for which there is currently no definitive cure. IBD shortens patients’ lifespans by nearly 10%, increasing the risk of colon cancer and sepsis. The two most common IBD diseases are ulcerative colitis (UC) and Crohn's disease (CD). The CDC estimates IBD affects nearly 4 million Americans with epidemiological studies reporting an over 50% increase in IBD prevalence in the last 15 years. IBD patients, and especially those with ulcerative colitis, have an over 10% risk of colorectal cancer, acute kidney injury, and sepsis. Current treatments intend to control inflammation using immunosuppressive drugs (e.g., anti-TNFα therapy) and prevent the lethal complications. However, these immunomodulatory treatments are very expensive, contributing to the daunting cost of IBD. More importantly, a significant percentage of IBD patients lose response to these treatments and experience detrimental side effects. Therefore, there is an unmet urgent need for novel treatments for IBD. The innate immune system orchestrates inflammatory responses to fight multiple disorders such as infections and cancer, but unregulated inflammation can become more dangerous than the original onset compromising vital organs. A typical example is severe sepsis, the most common cause of death in hospitalized patients killing over 250,000 Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 Americans each year. Sepsis is characterized by systemic inflammatory response syndrome (SIRS) and multi-organ dysfunction syndrome (MODS). In both syndromes, early TNFα production spreads in the bloodstream triggering systemic inflammation and septic shock. Metabolic acidoses are categorized into high- or non-anion gap metabolic acidosis, to determine the etiology and hence appropriate treatment. High anion gap metabolic acidosis (HAGMA) is the dominant blood gas anomaly in sepsis, and it is mostly due to ketoacidosis, lactic acidosis, or acute kidney injury (AKI). AKI is a life-threatening complication associated with higher mortality in sepsis, particularly in critically ill and elderly patients Sepsis and septic shock account for over 50% of AKI in ICUs. In addition to creatinine, which is a pivotal indicator for AKI prognosis, a study with 1505 patients reported that AKI negatively interacts with HAGMA, but not with lactate, as cofounder to worse 28- day mortality. Despite promising preclinical results, TNFα inhibition failed to improve survival in sepsis clinical trials, in part, because early TNFα production triggers multifactorial onslaughts leading to HAGMA and AKI. Thus, there remains a critical unmet need for novel treatments for sepsis. SUMMARY In some aspects, provided herein are methods of treating or preventing inflammatory bowel disease (IBD) in a subject comprising inhibiting SUMOylation in the subject. In some embodiments, the subject is a human. In some embodiments, inhibiting SUMOylation in the subject comprises performing vagal nerve stimulation on the subject and / or providing to the subject an inhibitor of SUMOylation. In some embodiments, inhibiting SUMOylation in the subject comprises performing vagal nerve stimulation on the subject and / or providing to the subject multiple inhibitors of SUMOylation. In some embodiments, inhibiting SUMOylation in the subject comprises providing to the subject an inhibitor of SUMOylation. In some embodiments, the inhibitor of SUMOylation inhibits SUMO maturation, SUMO activation, SUMO conjugation, SUMO ligation, and / or stimulates SUMO hydrolysis. In some embodiments, the inhibitor of SUMOylation is a SUMO E1 inhibitor, a SUMO E2 inhibitor, a SUMO E3 inhibitor, or a SENP inhibitor. In some embodiments, the inhibitor of SUMOylation is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, tannic acid, spectomycin B, viomellein, chaetochromin A, D-208, compound 22, momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, atromentin, compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 compound 12, compound 13 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor), ML-792, TAK-981, compound 23 (GSK145A), compound 24, compound 25, compound 26, compound 27, compound 34 (JCP-666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), compound 38 (VAE-561), compound 39, compound 40 (GN6767), compound 41 (GN6958), compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI-02), compound 46, compound 47, compound 48, compound 49, compound 50, compound 51, compound 52, compound 53, compound 54, a camptothecin derivative, topotecan, an ortho-quinone, nocardione A, 33-DINOR-dunnione, 33-DINOR-dehydrodunnione, β-lapachone, an iminoquinone, macrophilone A (compound 60) compound 61, a diterpene, triptolide, or compound 63. In some embodiments, inhibiting SUMOylation in the subject comprises providing to the subject multiple (e.g. two or more) inhibitors of SUMOylation. In some embodiments, the inhibitor of SUMOylation is TAK-981. In some embodiments, the method comprises providing the TAK-981 intravenously at a dose of 10 µg / kg to 5 mg / kg. In some aspects, provided herein are methods of treating or preventing sepsis in a subject, the method comprising performing vagal nerve stimulation on the subject and / or providing to the subject an inhibitor of SUMOylation. In some embodiments, the subject is a human. In some embodiments, the method comprises preventing sepsis in a subject, wherein the subject has an infection and is at risk of experiencing sepsis. In some embodiments, the method comprises treating sepsis in a subject. In some embodiments, inhibiting SUMOylation in the subject comprises performing vagal nerve stimulation on the subject and / or providing to the subject one or more inhibitors of SUMOylation. In some embodiments, inhibiting SUMOylation in the subject comprises performing vagal nerve stimulation on the subject and / or providing to the subject multiple inhibitors of SUMOylation. In some embodiments, inhibiting SUMOylation in the subject comprises providing to the subject an inhibitor of SUMOylation. In some embodiments, the inhibitor of SUMOylation inhibits SUMO maturation, SUMO activation, SUMO conjugation, SUMO ligation, and / or stimulates SUMO hydrolysis. In some embodiments, the inhibitor of SUMOylation is a SUMO E1 inhibitor, a SUMO E2 inhibitor, a SUMO E3 inhibitor, or a SENP inhibitor. In some embodiments, the inhibitor of SUMOylation is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, tannic acid, spectomycin B, viomellein, chaetochromin A, D-208, Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 compound 22, momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, atromentin, compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 12, compound 13 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor), ML-792, TAK-981, compound 23 (GSK145A), compound 24, compound 25, compound 26, compound 27, compound 34 (JCP-666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), compound 38 (VAE-561), compound 39, compound 40 (GN6767), compound 41 (GN6958), compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI-02), compound 46, compound 47, compound 48, compound 49, compound 50, compound 51, compound 52, compound 53, compound 54, a camptothecin derivative, topotecan, an ortho-quinone, nocardione A, 33-DINOR-dunnione, 33-DINOR-dehydrodunnione, β-lapachone, an iminoquinone, macrophilone A (compound 60) compound 61, a diterpene, triptolide, or compound 63. In some embodiments, inhibiting SUMOylation in the subject comprises providing to the subject multiple (e.g. two or more) inhibitors of SUMOylation. In some embodiments, the inhibitor of SUMOylation is TAK-981. In some embodiments, the method comprises providing to the subject the TAK-981 intravenously at a dose of 10 µg / kg to 5 mg / kg. In some aspects, provided herein is an inhibitor of SUMOylation for use in a method of treating or preventing IBD or sepsis in a subject. In some embodiments, the inhibitor is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, tannic acid, spectomycin B, viomellein, chaetochromin A, D-208, compound 22, momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, atromentin, compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 12, compound 13 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor), ML-792, TAK-981, compound 23 (GSK145A), compound 24, compound 25, compound 26, compound 27, compound 34 (JCP- 666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), compound 38 (VAE-561), compound 39, compound 40 (GN6767), compound 41 (GN6958), compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI-02), compound 46, compound 47, compound 48, compound 49, compound 50, compound 51, compound 52, compound 53, compound 54, a camptothecin derivative, topotecan, an ortho-quinone, nocardione A, 33- DINOR-dunnione, 33-DINOR-dehydrodunnione, β-lapachone, an iminoquinone, macrophilone A (compound 60) compound 61, a diterpene, triptolide, or compound 63. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 In some aspects, provided herein is an inhibitor of SUMOylation for use in the manufacture of a medicament for the treatment or prevention of IBD or sepsis in a subject. In some embodiments, the inhibitor is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, tannic acid, spectomycin B, viomellein, chaetochromin A, D-208, compound 22, momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, atromentin, compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 12, compound 13 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor), ML- 792, TAK-981, compound 23 (GSK145A), compound 24, compound 25, compound 26, compound 27, compound 34 (JCP-666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), compound 38 (VAE-561), compound 39, compound 40 (GN6767), compound 41 (GN6958), compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI-02), compound 46, compound 47, compound 48, compound 49, compound 50, compound 51, compound 52, compound 53, compound 54, a camptothecin derivative, topotecan, an ortho- quinone, nocardione A, 33-DINOR-dunnione, 33-DINOR-dehydrodunnione, β-lapachone, an iminoquinone, macrophilone A (compound 60) compound 61, a diterpene, triptolide, or compound 63. BRIEF DESCRIPTION OF THE DRAWINGS FIGS.1A-1F show inhibition of SUMOylation improved survival in polymicrobial peritonitis. (FIG.1A) Kaplan-Meier survival analyses in polymicrobial peritonitis induced by cecal ligation and puncture (CLP) in mice treated with vehicle or TAK981 (7.5 mg / kg; s.c.). *P<0.05 vs. Control, (n=25 / group, Survival Log-rank test. Experiment repeated twice at different days). (FIG.1B) blood urea nitrogen (BUN), (FIG.1C) IFNγ, and (FIG.1D) TNFα serum levels in control and septic mice treated with vehicle or TAK981. (FIG.1E) TNFα, and (FIG.1F) bacterial colony forming units in peritoneal lavage (PL) from control and CLP mice with or without splenectomy (spx) treated with vehicle or TAK981. *P<0.05 vs. Control (n=6 / group, Two-way ANOVA with Bonferroni’s post-hoc test). FIGS.2A-2F show inhibition of SUMOylation in fatal endotoxemia. (FIG.2A) Time- course of serum TNFα levels in endotoxemic (LPS, 7 mg / kg; i.p.) mice treated with vehicle (control) and TAK981 (7.5 mg / kg; s.c.) *P<0.05 vs. Control (n=6 / group, Two-way ANOVA with Bonferroni’s post-hoc test). Serum levels of (FIG.2B) potassium (K), (FIG.2C) anion Gap, (FIG.2D) blood urea nitrogen (BUN), and (FIG.2E) creatinine (Cre) at 48h post-LPS Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 in control and endotoxemic mice treated with vehicle or TAK981. *P<0.05 vs control (n=6 / group, unpaired two-tailed t-test). FIGS.3A-3D show splenectomy abrogated TAK981 control of systemic inflammation. (FIG.3A) Serum and organ TNFα levels at 1.5h in control and endotoxemic mice treated with vehicle or TAK981. *P<0.05 vs control;#P<0.05 vs LPS (n=6 / group, One- way ANOVA with Bonferroni’s post-hoc test). (FIG.3B) Serum TNFα levels in control or TAK981-treated after sham or splenectomy (SPX). (FIG.3C) Blood urea nitrogen (BUN) and (FIG.3D) creatinine (Cre) serum levels at 48h post-LPS in splenectomized (SPX) mice challenged with PBS (control) or LPS and treated with vehicle or TAK981. *P<0.05 vs sham (n=6 / group, unpaired two-tailed t-test). FIGS.4A-4F show endotoxemia induced a distinct temporal and substrate specificity for splenic SUMO isoforms. Western-blots (top) and densitometric analyses (bottom) of splenic SUMOylation at the indicated post-LPS time points. Bacterial endotoxin induced SUMOylation-1 of (FIG.4A) p150, (FIG.4B) p100, (FIG.4C) p250, and (FIG.4D) SUMOylation-2 / 3 of p70 and p260 (smear). Comparative densitometric analyses of (FIG.4E) SUMO1 and (FIG.4F) SUMO2 / 3 at the indicated post-LPS time-points. β-actin Western- blots (bottom panels) were used as an internal control for protein loading. were used as an internal control for protein loading. Western-blots represent experiments repeated at least twice on different days and graphs show mean±SEM of fold of induction (FI). *P<0.05 vs Control (n=3 / group, two-way ANOVA with Tukey’s post hoc test). FIGS.5A-5F show specific inhibition of SUMO1 unfettered serum TNFα levels in lethal endotoxemia. (FIG.5A) Serum and (FIG.5B) spleen levels of TNFα at 1.5h, and (FIG. 5C) potassium (K), (FIG.5D) anion gap (Gap), (FIG.5E) blood urea nitrogen (BUN) and (FIG.5F) creatinine (Cre) at 48h in wild-type (WT), SUMO1-KO (S1KO), and SUMO3-KO (S3KO) with vehicle or LPS. *P<0.05 vs control mice without LPS,#P<0.05 vs WT+LPS (n=6 / group, unpaired two-tailed t test). FIGS.6A-6E show adoptive transfer of SUMO1-null splenocytes enhanced inflammatory responses. Serum (FIG.6A) TNFα at 1.5h post-LPS and, (FIG.6B) potassium (K), (FIG.6C) anion Gap, (FIG.6E) blood urea nitrogen (BUN), and (FIG.6E) creatinine (Cre) at 48h post-LPS in control and endotoxemic mice after sham (control), splenectomy (SPX), and transfer of splenocytes from wild-type (SPLWT) or SUMO1-KO (SPLS1KO) mice. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 *P<0.05 vs control mice without LPS,#P<0.05 vs SPX+ SPLWT+LPS (n=6 / group, unpaired two-tailed t test). FIGS.7A-7D show blood chemistry after SUMO inhibition o in endotoxemic mice. (FIG.7A) Hematocrit, Serum levels of (FIG.7B) potassium (K), (FIG.7C) chloride (Cl), and (FIG.7D) total carbon dioxide (TCO2) at 48h post-LPS (LPS, 7 mg / kg; i.p.) in control and endotoxemic mice treated with vehicle or TAK981 (7.5 mg / kg; s.c.). *P<0.05 vs Control (n=3 / group, unpaired two-tailed t test). FIGS.8A-8F show blood chemistry of splenectomized mice after SUMO inhibition. (FIG.8A) Hematocrit, and serum levels of (FIG.8B) sodium (Na), (FIG.8C) potassium (K), (FG.8D) chloride (Cl), (FIG.8E) total carbon dioxide (TCO2), and (FIG.8F) anion Gap at 48h post-LPS (LPS, 7 mg / kg; i.p.) in splenectomized (SPX) mice challenged with PBS (control) or LPS (7 mg / kg; i.p.) and treated with vehicle or TAK981. FIGS.9A-9D show SUMOylation time-course during endotoxemia. Western-blots for SUMO1 (FIG.9A) and SUMO2 / 3 (FIG.9B) of spleen at the indicated post-LPS time points. Comparative densitometric analyses of (FIG.9C) SUMO1 and (FIG.9D) SUMO2 / 3 at the indicated post-LPS time-points. β-actin Western-blots (bottom panels) were used as an internal control for protein loading. were used as an internal control for protein loading. Western-blots represent experiments repeated at least twice on different days and graphs show mean±SEM of fold of induction (FI). *P<0.05 vs Control (n=3 / group, two-way ANOVA with Tukey’s post hoc test). FIGS.10A-10G show immune responses in endotoxemic SUMO1-KO and SUMO3- KO mice. Western-blots analyses of (FIG.10A) SUMO1 or (FIG.10B) SUMO2 / 3 of the spleen of control and endotoxemic (LPS, 7 mg / kg; i.p.) wild-type mice treated with vehicle (WT) or TAK981 (TAK, 7.5 mg / kg; s.c.), SUMO1-KO (S1KO), and SUMO3-KO (S3KO). β- actin Western-blots are internal control for protein loading. (FIG.10C) Serum, spleen, and liver TNFα levels at 1.5h post-LPS in endotoxemic (LPS, 7 mg / kg; i.p.) wild-type (WT), SUMO1-KO (S1KO), and SUMO3-KO (S3KO) mice. (FIG.10D) Hematocrit, (FIG.10E) Sodium, (FIG.10F) chloride, and (FIG.10G) total carbon dioxide at 48h post-LPS in wild- type (WT), SUMO1-KO (S1KO), and SUMO3-KO (S3KO).#P<0.05 vs wild-type endotoxemic mice (n=6 / group, Two-way ANOVA with Bonferroni’s post-hoc test). FIGS.11A-11D show adoptive transfer of SUMO1-null and SUMO3-null splenocytes into wild-type splenectomized mice. (FIG.11A) Hematocrit, (FIG.11B) Sodium, (FIG.11C) Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 chloride, and (FIG.11D) total carbon dioxide at 48h post-LPS in wild-type (WT) control mice, splenectomized (SPX), and transfer of splenocytes from wild-type (SPLWT), SUMO1- KO (SPLS1KO) mice. *P<0.05 vs SPX+SPLS1KOwithout LPS (n=6 / group, unpaired two-tailed t-test). FIGS.12A-12N show vagal stimulation improved preclinical outcomes in colitis. Time course of (FIG.12A) body weight and (FIG.12B) Disease Activity Index (DAI), (FIG. 12C) serum levels of corticosterone (HPA), norepinephrine, and heart rate variability (HRV measured as the Low / High-Frequency ratio) in 3%DSS mice with or without stress or (FIG. 12D) stress mice with sham or vagal stimulation (VS). (FIG.12E) Body weight and (FIG. 12F) DAI in stress DSS mice with sham or vagal stimulation. (FIG.12G) Intestinal bleeding, (FIG.12H) DAI, (FIG.12I) body weight, (FIG.12J) intestinal permeability, colon weights, (FIG.12K) representative macroscopic colons and group colon lengths, (FIG.12L) colonic H&E (20x, scale 200µM), (FIG.12M) histological score, and (FIG.12N) colon IL6, TNFα, and IL10 levels at day 7 in control and DSS mice with sham or vagal stimulation. Data are representative of experiments that were replicated on different dates. *P<0.05 vs Control (n=5, Two-way ANOVA with Bonferroni’s post-hoc test). *P<0.05 vs Control, #P<0.05 vs DSS (n=5, One-way ANOVA with Bonferroni’s post-hoc test). Graphs represent the mean ± SEM of experiments repeated twice on different dates. FIGS.13A-13P show vagal regulation of colitis is IL10-independent and mediated by the spleen. Time course of (FIG.13A) DAI, (FIG.13B) body weight, (FIG.13C) intestinal permeability, and (FIG.13D) colon IL6 and TNFα levels in wild type mice and IL10-KO (IL10KO) mice after 3% DSS. Time course of (FIG.13E) DAI, (FIG.13F) body weight, (FIG.13G) intestinal permeability, and (FIG.13H) colon IL6 and TNFα levels in IL10-KO (IL10KO) DSS mice with sham or vagal stimulation (VS). Time course of (FIG.13I) DAI, (FIG.13J) body weight, (FIG.13K) intestinal permeability, and (FIG.13L) colon IL6 and TNFα levels in DSS mice with sham surgery or splenectomy (SPX). Time course of (FIG. 13M) DAI, (FIG.13N) body weight, (FIG.13O) intestinal permeability, and (FIG.13P) colon IL6 and TNFα levels in splenectomized (SPX) DSS mice with sham or vagal stimulation. *P<0.05 vs Control, #P<0.05 vs DSS (n=5, One-way ANOVA with Bonferroni’s post-hoc test). Graphs represent the mean ± SEM of experiments repeated twice on different dates. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 FIGS.14A-14I show α7nAChR-independent vagal protection against colitis. (FIG. 14A.14G) DAI, (FIG.14B, 14H) body weight, (FIG.14C, 14I) representative colons, (FIG. 14D, 14J) group colon lengths and weight, (FIG.14E, 14K) intestinal permeability, and (FIG. 14F, 14L) colon immune cytokines in wild-type (WT) or α7nAChR-KO (α7KO), and in α7nAChR-KO (α7KO) DSS mice with sham or vagal stimulation (VS). *P<0.05 vs Control, #P<0.05 vs WT or DSS (n=4, One-way ANOVA with Bonferroni’s post-hoc test). Graphs represent the mean ± SEM of experiments repeated twice on different dates. FIGS.15A-15E show that nicotine improves colitis outcomes through α7nAChRs. (FIG.15A, 15F) DAI, (FIG.15B, 15G) intestinal bleeding, (FIG.15C, 15H) body weight, (FIG.15D, 15I) colon length, intestinal permeability, and (FIG.15E, 15J) colon immune cytokines in wild-type or α7nAChR-KO (α7KO) DSS mice treated with vehicle or nicotine. *P<0.05 vs Control, #P<0.05 vs DSS (n=5, One-way ANOVA with Bonferroni’s post-hoc test). Graphs represent the mean ± SEM of experiments repeated twice on different dates. FIGS.16A-16E show SUMOylation in human biopsy and vagal regulation of colitis. (FIG.16A) DEseq normalized counts for Ubc9 expression in human biopsy samples from 29 non-IBD patients (NIBD) and 91 CD patients (p=0.34). (FIG.16B) Western blots and (FIG. 16C) IHC for SUMO1 and SUMO2 / 3 of colon biopsy from NIBD, UC, and CD patients. Data shows representative results of 7 biopsies from 18 human samples analyzed. Brackets point to the typical high molecular weight smears induced by poly SUMOylation. (FIG.16D) Western blots, densitometric analyses, and (FIG.16E) IHC for SUMO1 and SUMO2 / 3 of colon samples from control or 3% DSS mice with sham or vagal stimulation. Western blots represent experiments repeated at least twice on different days and graphs show mean ± SEM of fold of induction (FI). *P<0.05 vs Control, #P<0.05 vs DSS (n=5 / group, two-way ANOVA with Tukey’s post hoc test). FIGS.17A-17P show bone marrow transfer from SUMO1-null mice confers protection to colitis. (FIG.17A) DAI, (FIG.17B) representative colons, (FIG.17C) group colon lengths and intestinal permeability, and (FIG.17D) immune cytokines in wild-type, SUMO1-KO, and SUMO3-KO control mice after DSS. (FIG.17E) DAI, (FIG.17F) representative colons, (FIG.17G) group colon lengths and intestinal permeability, and (FIG. 17H) immune cytokines in SUMO1-KO DSS mice with sham or VS. (FIG.17I) DAI, (FIG. 17J) representative colons, (FIG.17K) group colon lengths and intestinal permeability, and (FIG.17L) immune cytokines in SUMO3-KO DSS mice with sham or VS. (FIG.17M) DAI, Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 (FIG.17N) representative colons, (FIG.17O) group colon lengths and intestinal permeability, and (FIG.17P) immune cytokines in irradiated wild-type DSS (Ir.WT+DSS) mice transferred with wild-type, SUMO1-KO, or SUMO3-KO bone marrow. *P<0.05 vs Control or WT or Control, #P<0.05 vs DSS (n=4, One-way ANOVA with Bonferroni’s post-hoc test). Graphs represent the mean ± SEM of experiments repeated twice on different dates. FIG.18A-18D show inhibition of SUMOylation by TAK981. SUMO1 and SUMO2 / 3 western blot analyses of (FIG.18A, FIG.18B) colon from mice with 7 days 3%DSS treatment and (FIG.18C, FIG.18D) spleen from endotoxemic (LPS, 7.5mg / Kg for 1.5h) mice. The figure shows a representative experiment repeated 2 times for colon samples and 3 times for spleen samples. FIGS.19A-19E show inhibition of SUMOylation with TAK981 improved colitis outcomes. (FIG.19A) DAI, (FIG.19B) body weight, (FIG.19C) representative colons and group colon lengths, (FIG.19D) H&E of colonic tissues (10x, 20x, scale 200uM), (FIG.19E) histological score, intestinal permeability, and (FIG.19F) colon immune cytokines in control or colitis mice treated with vehicle or TAK981. *P<0.05 vs Control, #P<0.05 vs DSS (n=5, One-way ANOVA with Bonferroni’s post-hoc test). FIGS.20A-20M show TAK981 treatment rescued mice from established colitis. (FIG. 20A, 20D, 20G) DAI, (FIG.20B, 20E, 20H) intestinal bleeding, (FIG.20C, 20F, 20I) body weight, (FIG.20J) intestinal permeability and peritoneal lavage (PL) bacteria counts, (FIG. 20K) colon immune cytokines, and (FIG.20L) representative colons, and (FIG.20M) colonic H&E (scale 200uM) in 3% DSS colitis mice treated with vehicle or ‘rescue’ TAK981 (rTAK) when treatment was started 3 days after DSS challenge, TNBS or Oxazolone (OXA) mice treated with vehicle or TAK981. *P<0.05 vs Vehicle (n=4, Two-way ANOVA with Bonferroni’s post-hoc test). *P<0.05 vs Control, #P<0.05 vs OXA (n=4, One-way ANOVA with Bonferroni’s post-hoc test). FIG.21. Pharmacologic inhibition of SUMOylation as an alternative strategy for vagal regulation of colitis. Stress can induce parasympathetic autonomic dysfunction as determined by HRV that contributes to colitis. Conversely, vagal stimulation improves murine colitis via the spleen. VS regulates colon SUMOylation mainly inhibiting DSS- induced SUMO2 / 3. SUMO1-KO mice delay the onset of colitis and SUMO3-KO mice are more resistant to colitis. The transfer of SUMO1-KO, but not SUMO3-KO, bone marrow confers resistance to wild-type mice against colitis. Inhibition of SUMOylation with TAK981 Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 rescued mice from established DSS colitis and protected them against TNBS and oxazolone- induced colitis. DETAILED DESCRIPTION Section headings as used in this section and the entire disclosure herein are merely for organizational purposes and are not intended to be limiting. 1. Definitions Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. In case of conflict, the present document, including definitions, will control. Preferred methods and materials are described below, although methods and materials similar or equivalent to those described herein can be used in practice or testing of the present disclosure. All publications, patent applications, patents and other references mentioned herein are incorporated by reference in their entirety. The materials, methods, and examples disclosed herein are illustrative only and not intended to be limiting. The terms “comprise(s),” “include(s),” “having,” “has,” “can,” “contain(s),” and variants thereof, as used herein, are intended to be open-ended transitional phrases, terms, or words that do not preclude the possibility of additional acts or structures. The singular forms “a,” “and” and “the” include plural references unless the context clearly dictates otherwise. The present disclosure also contemplates other embodiments “comprising,” “consisting of” and “consisting essentially of,” the embodiments or elements presented herein, whether explicitly set forth or not. For the recitation of numeric ranges herein, each intervening number there between with the same degree of precision is explicitly contemplated. For example, for the range of 6- 9, the numbers 7 and 8 are contemplated in addition to 6 and 9, and for the range 6.0-7.0, the number 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, and 7.0 are explicitly contemplated. As used herein, the term “subject” broadly refers to any animal, including but not limited to, human and non-human animals (e.g., dogs, cats, cows, horses, sheep, poultry, fish, crustaceans, etc.). In some embodiments, the subject is a human. In some embodiments, the subject has, is suspected of having, or is at risk of having inflammatory bowel disease. In some embodiments, the subject has, is suspected of having, or is at risk of having sepsis. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 As used herein, the term “preventing” refers to prophylactic steps taken to reduce the likelihood of a subject (e.g., an at-risk subject) from contracting or suffering from a particular disease, disorder, or condition. The likelihood of the disease, disorder, or condition occurring in the subject need not be reduced to zero for the preventing to occur; rather, if the steps reduce the risk of a disease, disorder or condition across a population, then the steps prevent the disease, disorder, or condition for an individual subject within the scope and meaning herein. As used herein, the terms “treatment,” “treating,” and the like refer to obtaining a desired pharmacologic and / or physiologic effect against a particular disease, disorder, or condition. Preferably, the effect is therapeutic, i.e., the effect partially or completely cures the disease and / or adverse symptom attributable to the disease. “Treatment” may refer to reducing the amount or severity of a particular condition, disease state (e.g., inflammatory bowel disease, sepsis), or symptoms thereof, in a subject presently experiencing or afflicted with the condition or disease state. The terms do not necessarily indicate complete treatment (e.g., total elimination of the condition, disease, or symptoms thereof). "Treatment,” encompasses any administration or application of a therapeutic or technique for a disease (e.g., in a mammal, including a human), and includes inhibiting the disease, arresting its development, relieving the disease, causing regression, or restoring or repairing a lost, missing, or defective function; or stimulating an inefficient process. The term “therapeutically effective dose” refers to an amount of an agent, e.g., an inhibitor of SUMOylation, that results in the reduction of symptoms in a patient, treatment of prevention of a disease or condition, or results in a desired biological outcome. As used herein, the terms “administration” and “administering” refer to the act of providing an agent (e.g. an inhibitor of SUMOylation) to a subject. Suitable routes of administration include, without limitation: topical, parenteral (e.g. by injection, such as intramuscular, intravenous, intraperitoneal, subcutaneous, etc.), oral, rectal, and the like. As used herein, the terms “co-administration” and “co-administering” refer to the administration of at least two agent(s) or therapies to a subject. In some embodiments, the co- administration of two or more agents or therapies is concurrent. In other embodiments, a first agent / therapy is administered prior to a second agent / therapy. Those of skill in the art understand that the formulations and / or routes of administration of the various agents or therapies used may vary. The appropriate dosage for co-administration can be readily Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 determined by one skilled in the art. In some embodiments, when agents or therapies are co- administered, the respective agents or therapies are administered at lower dosages than appropriate for their administration alone. Thus, co-administration is especially desirable in embodiments where the co-administration of the agents or therapies lowers the requisite dosage of a potentially harmful (e.g., toxic) agent(s), and / or when co-administration of two or more agents results in sensitization of a subject to beneficial effects of one of the agents via co-administration of the other agent. 2. Methods Inflammatory bowel disease (IBD) refers to a spectrum of chronic disorders characterized by inflammation in the gastrointestinal tract. Two common types of IBD include ulcerative colitis ad Crohn’s disease. Ulcerative colitis is a condition involving inflammation and scores / ulcers along the lining of the large intestine and rectum. Crohn’s disease is a type of IBD characterized by inflammation of the lining of the gastrointestinal tract which can often involve deeper layers of tissue. Crohn’s disease most commonly affects the small intestine, but it can also affect the large intestine and / or the upper gastrointestinal tract. IBD is generally characterized by diarrhea, rectal bleeding, abdominal pain, fatigue, and weight loss. In some aspects, provided herein are methods of treating and / or preventing inflammatory bowel disease (IBD) in a subject involving inhibiting SUMOylation in the subject. In some embodiments, a method of treating IBD in a subject involves inhibiting SUMOylation in a subject presently experiencing one or more symptoms of IBD or already diagnosed with IBD. In some embodiments, a method of preventing IBD in a subject involves inhibiting SUMOylation in a subject at risk of having IBD, thereby reducing the likelihood of IBD developing in the subject. Sepsis refers to a dangerous and extreme immune response to infection that can lead to tissue damage, organ failure, or death. Sepsis is characterized by systemic inflammatory response syndrome (SIRS) and multi-organ dysfunction syndrome (MODS). In both syndromes, early TNFα production spreads in the bloodstream triggering systemic inflammation and septic shock. Common symptoms of sepsis include fever, elevated heart rate, weak pulse, low blood pressure, rapid breathing, difficulty breathing, clammy and / or Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 sweaty skin, confusion, convulsions, pale skin, lethargy, difficulty waking up, low urine output, and body pain. In some aspects, provided herein are methods of treating and / or preventing sepsis in a subject. In some embodiments, methods of treating sepsis in a subject comprising providing an inhibitor of SUMOylation to a subject currently experiencing one or more symptoms of sepsis, thereby treating sepsis in the subject. In some embodiments, methods of preventing sepsis in a subject comprise providing to a subject at risk of having sepsis an inhibitor of SUMOylation, thereby preventing sepsis in the subject. For example, a subject may be at risk of having sepsis due to an infection, such as an infection (e.g. a bacterial infection, a viral infection). In some embodiments, preventing sepsis in a subject comprises inhibiting SUMOylation in a subject having an infection, thereby preventing sepsis in the subject. SUMOylation refers to a post-translational modification that involves the covalent, reversible attachment of small ubiquitin-like modifier (SUMO) polypeptide to a target protein. In particular, SUMOylation involves the covalent attachment of SUMO polypeptide to a lysine residue of a target protein. SUMOylation is a dynamic and reversible process that is controlled by an enzymatic cascade similar to the ubiquitinoylation pathway. SUMOylation requires an ATP-dependent heterodimeric SUMO-activating enzyme (SAE) referred to as E1, a SUMO conjugating enzyme referred to as E2, and E3 ligases that assist in the recognition of substrates. SUMOylation is reversed by SUMO / sentrin specific peptidases (SENPs) that hydroylze SUMO from the substrate. There is only one known SUMO E2 enzyme, Ubc9. There are six known SENPs (SENP1-3, and SENP5-7) and various known SUMO E3 ligases, including those in the SP-RING domain family, TRIM superfamily, and noncanonical ligases (e.g. SIM-containing SUMO E3 ligases). The first step in the SUMOylation pathway is maturation of the SUMO protein. Two isoforms of SENPs, SENP1 and SENP2, are primarily responsible for cleaving the tetrapeptide C-terminal cap to reveal a di-glycine motif. The di-glycine motif eventually forms the isopeptide bond with the ε-amino group of a lysine residue on the protein substrate. After SUMO maturation has occurred, the E1 enzyme activates the protein for conjugation. Activation involves formation of an E1-SUMO complex, after which SUMO transferred to a cysteine residue on the SUMO-conjugating enzyme, SUMO E2. SUMO E2 catalyzes formation of the isopeptide bond between the side chain amine of lysine and the C-terminal glycine residue of SUMO, often with the assistance of an E3 ligase, which can increase the Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 rate of SUMOylation. E3 ligases are also involved in colocalization of the E2-SUMO complex and the target protein. However, it has been demonstrated that SUMOylation can occur without the use of an E3 ligase. In some embodiments, the methods provided herein involve providing to a subject one or more inhibitors of SUMOylation. The inhibitor of SUMOylation may inhibit SUMO maturation (e.g. by inhibiting SENP1 or SENP2), SUMO activation (e.g. by inhibiting E1), SUMO conjugation (e.g. by inhibiting the SUMO conjugating enzyme Ubc9), SUMO ligation (e.g. by inhibiting an E3 ligase), or may activate the reversal of SUMOylation (e.g. by increasing the activity of SENPs involved in hydrolysis). In some embodiments, the inhibitor of SUMOylation is a natural product inhibitor. In some embodiments, the natural product inhibitor of SUMOylation is a natural product E1 inhibitor. In some embodiments, the natural product inhibitor is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, or tannic acid. The structural of these natural product inhibitors is shown below, where 1 is ginkgolic acid, 2 is anacardic acid, 3 is kerriamycin B, 4 is the polyphenolic inhibitor davidiin, and 5 is the polyphenolic inhibitor tannic acid.

[0002] Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 In some embodiments, the natural product inhibitor of SUMOylation is a natural product inhibitor or SUMO-E2. In some embodiments, the natural product inhibitor is spectomycin B, viomellein, chaetochromin A, D-208, or compound 22. The structure of these exemplary natural product inhibitors is shown below, where spectomycin B is 18, viomellein is 19, chatochromin A is 20, D-208 is 21, and compound 22 is 22. In some embodiments, the inhibitor of SUMOylation is a natural product SENP inhibitor. In some embodiments, the natural product inhibitor is momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, or atromentin. The structure of such natural product inhibitors is shown below, where momordin Ic is 28, streptonigrin is 29, NSC76919 is 30, NSC45384 is 31, vialinin A is 32, and atromentin is 33.

[0003] Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 In some embodiments, the inhibitor of SUMOylation is a semisynthetic product. Exemplary semisynthetic products include, for example, compound 6 and compound 7. The structure of compounds 6 and 7 are show below. In some embodiments, the inhibitor of SUMOylation is a synthetic product. Exemplary synthetic inhibitors include compound 8, compound 9, compound 10, compound11, compound 12, compound 13 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor). The structure of compounds 8-15 are shown below. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 In some embodiments, the inhibitor of SUMOylation is ML-792 or TAK-981. The structure of ML-792 is shown below as 16, and the structure of TAK-981 is shown below as 17.

[0004] Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 In some embodiments, the inhibitor of SUMOylation is a Ubc9 inhibitor. Exemplary Ubc9 inhibitors include compound 23 (GSK145A), compound 24, compound 25, compound 26, and compound 27. The structures of compounds 23-27 are shown below. In some embodiments, the inhibitor of SUMOylation is a SENP inhibitor. In some embodiments, the inhibitor of SUMOylation is the SENP inhibitor compound 34 (JCP-666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), or compound 38 (VAE- 561). The structures of compounds 34-38 are provided below.

[0005] Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 inhibitor of SENP. In some embodiments, the inhibitor of SUMOylation is compound 39, compound 40 (GN6767), or compound 41 (GN6958). In some embodiments, the inhibitor of SUMOylation is compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI-02), compound 46, compound 47, compound 48, compound 49, compound 50, compound 51, compound 52, compound 53, or compound 54. The structures of compounds 43-54 are shown below.

[0006] Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 In some embodiments, the inhibitor of SUMOylation is a camptothecin derivative. In some embodiments, the inhibitor of SUMOylation is topotecan (compound 55). In some embodiments, the inhibitor of SUMOylation is an ortho-quinone. In some embodiments, the inhibitor of SUMOylation is nocardione A (compound 56), 33-DINOR-dunnione (compound 57), 33-DINOR-dehydrodunnione (compound 58), or β-lapachone (compound 59). In some embodiments, the inhibitor of SUMOylation is an iminoquinone. In some embodiments, the Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 inhibitor of SUMOylation is macrophilone A (compound 60) or compound 61. In some embodiments, the inhibitor of SUMOylation is a diterpene. In some embodiments, the inhibitor of SUMOylation is triptolide (compound 62). In some embodiments, the inhibitor of SUMOylation is compound 63. The structures of compounds 55-63 are shown below. The inhibitor of SUMOylation may be provided to the subject in a pharmaceutical formulation comprising one or more pharmaceutically acceptable excipients (e.g. carriers, vehicles, excipients, preservatives, stabilizers, or other auxiliary agents that do not induce an undesirable biological effect in the subject). In some embodiments, the inhibitor of SUMOylation is provided to the subject orally. In some embodiments, the inhibitor of SUMOylation is provided to the subject parenterally (e.g. by injection). For example, in some embodiments the inhibitor of SUMOylation is provided to the subject by a suitable injection route, including intramuscular, intravenous, subcutaneous, intraperitoneal, and the like. In some embodiments, the inhibitor of SUMOylation is provided to the subject rectally. The dose of the inhibitor of SUMOylation may depend on the age of the subject, the weight of the subject, the precise inhibitor used, the disease condition afflicting the subject, and / or the severity of symptoms in the subject. The inhibitor of SUMOylation may be provided to the subject at any suitable dose (e.g. a therapeutically effective dose) at any suitable frequency and for any suitable duration to achieve the desired effect (e.g. the desired treatment and / or prevention of IBD, the desired treatment and / or prevention of sepsis). Suitable doses of the inhibitor of SUMOylation may be from 1 µg of the inhibitor / kg body Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 weight to the subject to 10 mg / kg. For example, a suitable dose may be 1µg / kg to 10 mg / kg, 10 µg / kg to 9.5 mg / kg, 20 µg / kg to 9 mg / kg, 30 µg / kg to 8.5 mg / kg, 40 µg / kg to 8 mg / kg, 50 µg / kg to 7.5 mg / kg, 100 µg / kg to 7 mg / kg, 200 µg / kg to 6.5 mg / kg, 300 µg / kg to 6 mg / kg, 400 µg / kg to 5.5 mg / kg, 500 µg / kg to 5 mg / kg, 600 µg / kg to 4.5 mg / kg, 700 µg / kg to 4 mg / kg, 800 µg / kg to 3.5 mg / kg, 900 µg / kg to 3 mg / kg, or 1 mg / kg to 2 mg / kg. In some embodiments, the inhibitor of SUMOylation is TAK-981 (also referred to herein as TAK981). In some embodiments, the methods herein comprise providing to the subject TAK-981. In some embodiments, TAK-981 is provided to the subject parenterally (e.g. by injection). In some embodiments, TAK-981 is provided to the subject intravenously at a dose of 10 µg / kg to 5 mg / kg. In some embodiments, the inhibitor of SUMOylation is provided to the subject once per day. In some embodiments, the inhibitor of SUMOylation is provided to the subject multiple times per day. In some embodiments, the inhibitor of SUMOylation is provided to the subject every other day, every 3 days, every 4 days, every 5 days, every 6 days, every 7 days, every 8 days, every 9 days, every 10 days, every 2 weeks, every 3 weeks, monthly, every 2 months, every 3 months, every 4 months, every 5 months, every 6 months, etc. In some embodiments, inhibiting SUMOylation in the subject involves performing vagal nerve stimulation on the subject. In some embodiments, the methods herein involve performing vagal nerve stimulation on the subject and providing to the subject one or more inhibitors of SUMOylation. It will be readily apparent to those skilled in the art that other suitable modifications and adaptations of the methods of the present disclosure described herein are readily applicable and appreciable, and may be made using suitable equivalents without departing from the scope of the present disclosure or the aspects and embodiments disclosed herein. Having now described the present disclosure in detail, the same will be more clearly understood by reference to the following examples, which are merely intended only to illustrate some aspects and embodiments of the disclosure, and should not be viewed as limiting to the scope of the disclosure. The disclosures of all journal references, U.S. patents, and publications referred to herein are hereby incorporated by reference in their entireties. The present disclosure has multiple aspects, illustrated by the following non-limiting examples. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 EXAMPLES Example 1 TAK981(Subasumstat) is the first-in-class selective inhibitor of SUMOylation. TAK981 is in clinical trials to potentiate immunotherapy in metastatic tumors and hematologic malignancies. Cancer patients have more than ten times higher risk of infections, but the effects of TAK981 in sepsis are unknown and previous studies on SUMO in infections are conflicting. Herein, it is demonstrated that inhibition of SUMOylation with TAK981 improved survival in mild polymicrobial peritonitis by enhancing innate immune responses and peritoneal bacterial clearance. The effects of TAK981 on the immune responses to bacterial endotoxin were evaluated, showing that TAK981 enhanced early TNFα production but did not affect the resolution of inflammation. Splenectomy decreased serum TNFα levels by nearly 60% and TAK981-induced TNFα responses. In the spleen, endotoxemia induced a distinct temporal and substrate specificity for SUMO1 and SUMO2 / 3, and both were inhibited by TAK981. Global genetic depletion of SUMO1, but not SUMO3, enhanced TNFα production and metabolic acidosis. The transfer of SUMO1-null, but not wild-type, splenocytes into splenectomized wild-type mice exacerbated TNFα production and metabolic acidosis in endotoxemia. These results indicate that specific regulation of splenic SUMO1 can modulate immune and metabolic responses to bacterial infection. MATERIALS AND METHODS Chemicals and reagents. LPS (Escherichia coli 0111:B4) was purchased from Sigma-Aldrich (Saint Louis, MO) and dissolved in sterile pyrogen-free PBS (Gibco, Life Technologies, Grand Island, NY). SUMO inhibitor TAK981 (Subasumstat) was purchased from ChemieTek (Indianapolis, IN). Animal experiments. Animal procedures were approved by the Institutional Animal Care & Use Committee of Duke University. All animal experiments were performed in 6-8- week-old (∼25 ± 5 g) male mice. Wild-type C57BL / 6 male mice were obtained from The Jackson Laboratory (Bar Harbor, ME). SUMO1-, SUMO3-knockout and littermates wild- type mice were produced. Knockout mice were genotyped by PCR using tail genomic DNA and the Extract-N-Amp Tissue PCR kit (Sigma Chemical, Saint Louis, MO). Mice were maintained on 12h light-dark cycle, with free access to food and water (ad libitum). Mice Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 were randomized into experimental groups and outcome assessors were blinded to group treatments. Experimental sepsis. Cecal ligation and puncture (CLP) were performed. Mice were anesthetized with 5% isoflurane, shaved, and the lower abdomen was disinfected. A one cm vertical incision was made to expose the cecum. Cecal ligation was done mid-way between the cecal tip and the ileocecal valve using a 6 / 0 proline. The cecal puncture was done once between the ligature site and cecal tip using a 25-gauge needle and the stool was extruded one mm. Then, the cecum was returned to the abdomen, and the abdominal wall was sutured in layers using a 6 / 0 proline. Mice were pretreated with vehicle or TAK981 (7.5 mg / Kg; s.c.) 12h before the CLP and every 24h for 4d starting right after the surgery. Survival was monitored every 12 hours for two weeks. Endotoxemia was performed using a lethal dose of LPS (7 mg / Kg; i.p.). LPS (E. coli LPS 0111:B4; Sigma Chemical, Saint Louis, MO) was dissolved in sterile pyrogen-free PBS (Gibco: Life Technologies, Grand Island, NY) and sonicated for 10 min immediately before use. Splenectomy. Splenectomy was performed. Mice were anesthetized with isoflurane 5% and subjected to an abdominal incision on the epigastrium and mesogastrium. The spleen was exposed by gentle retraction of the stomach to the side. The three main branches of the splenic artery were stabilized with nylon thread, ligated, and cut. The spleen was removed, and the abdominal wall was sutured with catgut and the skin with nylon thread. Mice were splenectomized 5 days before the experimental procedure and checked daily. Cytokine and organ function analyses. Mice were anesthetized by 5% isoflurane and blood was collected by cardiac puncture at the indicated time points, allowed to clot for 1h at room temperature, and centrifuged at 2,000g for 20 min at 4°C. Organs were collected at the indicated time points, snap frozen in liquid nitrogen, and homogenized on ice in RIPA buffer supplemented with protease inhibitor (Pierce protease inhibitor mini-tablets, A32955, Thermo Scientific, Rockford, IL). TNFα and IFNγ were analyzed by ELISA (Invitrogen, Life Technologies, NY) as per manufacturer instructions. Blood chemistry was analyzed at 48 hours in blood collected from the vena cava into heparin microtiter tubes and analyzed with CHEM8+ cartridges with the iSTAT1 blood analyzer (Abbott Laboratories, Chicago, IL). Peritoneal lavage and bacterial load analysis. Peritoneal lavage was performed. Briefly, 24 hours after CLP, mice were anesthetized with 5% isoflurane, the abdominal wall was disinfected with 70% ethanol, and 5 mL of sterile pyrogen-free PBS was injected i.p. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 using a 25g needle. Then, the abdomen was massaged for 1 minute, and 1 mL of peritoneal fluid was aspirated using a sterile syringe. Peritoneal aspirates were centrifuged at 2,000g for 20 min at 4°C and supernatants were frozen at -20 for further cytokines analysis. Peritoneal lavage bacterial load was analyzed. Freshly collected peritoneal aspirates were cultured in blood agar using a sterile inoculation loop and agar plates (Hardy Diagnostics, Santa Maria, CA) and were incubated at 37°C for 24 hours. Bacterial colonies (CFU / mL of peritoneal aspirate) were counted using an ImageJ Fiji software cell analyzer. Western-blots. Western blot assays were performed. The spleens were homogenized (1:10 w / v) in lysis buffer (50mM β-glycerophosphoric acid, disodium salt, 5-hydrate, 1mM EGTA pH 8.0, 0.5 mM Na3VO4 and triton x-100) supplemented with protease inhibitor (Pierce protease inhibitor mini-tablets, A32955, Thermo Scientific, Rockford, IL), 20mM SUMO degradation inhibitor N-ethylmaleimide (Sigma, Saint Louis, MO), and 0.5mM DL- Dithiothreitol (VWR, Radnor, PA), then centrifuged at 16,000g for 15min at 4°C. Supernatant protein concentration was determined with Bio-Rad Protein Assay (Bio-Rad, Life Science Research, Hercules, CA). Samples were run on the Bolt Bis-Tris Mini Gels (Novex, Invitrogen, Life Technologies, Grand Island, NY) at 150 V for 50 min. The samples were transferred onto the PVDF membranes (Hybond-P, Amersham, GE Healthcare, Pittsburg, PA) with the Bio-Rad Gel Transfer (Bio-Rad, Life Science Research, Hercules, CA). The membranes were blocked with 5% skimmed milk-TBST for 1 h at 4°C and incubated with antibodies against SUMO1 (1:1,000, PA5-96065, Thermo Scientific, Rockford, IL) or SUMO2 / 3 (1:1,000, PA5-19418, Thermo Scientific, Rockford, IL) overnight at 4°C. Then, washed in TBST and incubated with HRP-linked antibody to rabbit IgG (1:5,000, G21234, Thermo Scientific, Rockford, IL) for 1h at room temperature. The membranes were washed and processed with chemiluminescence (Amersham, Buckinghamshire, UK) and analyzed with Amersham gel imager 680 (Amersham, Buckinghamshire, UK). Splenocytes transfer. Splenocytes were isolated by harvesting spleens and mechanically disrupting using 70-μm nylon mesh, then filtering using a 70μm cell strainer in ice-cold PBS. The suspension was centrifuged 800g for 6 minutes at 4°C, and pellets were resuspended in RBC lysis buffer (Biolegend, Sand Diego, CA) for 5 minutes on ice. Cells were washed twice in ice-cold PBS before resuspending them in Trypan Blue 1:1 for cell counting (Countess™, Invitrogen, Life Technologies, Grand Island, NY). Splenocytes Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 (10x106live cells / 20g mouse) were injected i.p. in splenectomized recipient mice 24 hours before the LPS challenge. Statistical analyses. Statistical analyses were performed with GraphPad Prism Software (GraphPad Software, San Diego, CA). The sample size was calculated based on our preliminary results using G*Power3.1 and ANOVA (Fixed effects, omnibus, one-way), assuming effect size= 0.25, α=0.05, power (1-beta err prob)=0.8. Data are expressed as mean±sem and figures represent experiments repeated twice on different days. The Student's t-test (Mann-Whitney U test) was used to compare two experimental groups. Three or more groups were analyzed with parametric one-way ANOVA with multiple pair-wise comparisons. Normality and homogeneity of variance were confirmed with Kolmogorov- Smirnov analyses. Pair comparisons in nonparametric ANOVA tests were adjusted post-hoc with the Tukey test (in equal sample sizes) or Bonferroni’s for multiple hypothesis testing. The time courses and pair-wise comparisons were analyzed with the two-way ANOVA for repeated measures. Survival was analyzed with the Log-rank (Mantel-Cox) test. Statistical significance was established at p≤.05 with a 2-sided alpha according to Cohen (Cohen, J. Statistical power analysis for the behavioral sciences, (Routledge, 2013). RESULTS Inhibition of SUMOylation improved survival in polymicrobial peritonitis. Previous studies of SUMOylation were performed in endotoxemia, an experimental model that focuses on the immune responses to bacterial endotoxin (LPS) without an active pathogen. The effects of TAK981 on polymicrobial peritonitis were evaluated. Unlike endotoxemia, cecal ligation and puncture is the most clinically relevant experimental model of sepsis with polymicrobial peritonitis produced by the cecal puncture and necrotic tissue produced by cecal ligation. TAK981 treatment improved survival in mild polymicrobial peritonitis induced by cecal ligation and puncture (Fig.1a). Survival was recorded for over two weeks and no late deaths were observed. In line with previous studies, mild polymicrobial peritonitis induces a mild increase in blood urea nitrogen (Fig.1b). Next, the effects of TAK981 on the immune responses to bacterial infection were analyzed. TAK981 treatment did not affect blood urea nitrogen but increased the innate immune responses to bacterial infection including serum IFNγ and TNFα levels (Fig.1c,d). TAK981 treatment also increased TNFα levels in peritoneal lavage (Fig.1e), and significantly decreased bacteria Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 count in peritoneal lavage of septic mice (Fig.1f). Since the spleen is the major source of inflammatory cytokines in experimental sepsis, the experiments were repeated in splenectomized animals to confirm that splenectomy inhibited all the effects induced by TAK981. These results demonstrate that inhibition of SUMOylation with TAK981 enhances innate immune responses to fight infections. Inflammatory responses can become more detrimental than the original infection. Given the complexity of polymicrobial peritonitis, the immune and metabolic responses to bacterial endotoxin were next investigated using experimental models of endotoxemia, which is the most common and reliable model to study the innate immune system. TAK981 treatment given at 12h and right before the endotoxemic challenge (LPS, 7 mg / kg; i.p.) did not induce TNFα in control mice but increased TNFα production in lethal endotoxemia (Fig.2a). TAK981 increased acute TNFα production but did not affect immune resilience and resolution of serum TNFα levels. Serum TNFα levels peak at 1.5h and quickly resolve by nearly 80% at 3h. Likewise, TAK981 did not affect blood chemistry or anion gap in control mice, but increased hematocrit, hyperkalemia, and high anion gap metabolic acidosis (HAGMA) in lethal endotoxemia (FIG.7a; Fig.2b,c). These effects were specific because TAK981 did not affect natremia, chloremia, or total carbon dioxide (Fig.7b-d). These results indicate kidney dysfunction, and TAK981 increased blood urea nitrogen and serum creatinine by 25% and 50%, respectively in lethal endotoxemia without affecting control mice (Fig. 2d,e). These results show that SUMOylation is an integral mechanism that can be exploited to modulate systemic immune responses to bacterial infection. Inhibition of SUMOylation with TAK981 can enhance TNFα responses to fight bacterial infection in mild polymicrobial peritonitis. Splenectomy abrogated TAK981 control of systemic inflammation. Next, it was analyzed how TAK981 modulates systemic immune responses to bacterial endotoxin by analyzing the TNFα levels in different organs. Organ analyses showed the highest TNFα levels in the spleen, and the main effects of TAK981 were increasing TNFα levels specifically in the spleen and liver but not in other organs like the kidney or pancreas (Fig. 3a). Then, the role of the spleen and liver was analyzed by performing surgical splenectomy. Splenectomy prevented most of TAK981-induced immune and metabolic effects, including TNFα production, hyperkalemia, HAGMA, and kidney dysfunction markers blood urea nitrogen and creatinine (Fig.3b-d; Fig.8a-f). These results show that the immune and metabolic effects induced by TAK981 are mainly mediated by the spleen. Thus, Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 SUMOylation in the spleen appears to reprogram the immune and metabolic responses to bacterial infections. Endotoxemia induced a distinct temporal and substrate specificity for splenic SUMO isoforms. The specific SUMO isoforms in the spleen were analyzed by Western blots for SUMO1 and SUMO2 / 3. SUMO2 and SUMO3 are referred to together as SUMO2 / 3 because they share ∼95% sequence identity and are undistinguishable by current antibodies. Full SUMO1 blots appear to show that the levels of conjugated follow that of non-conjugated SUMO1 suggesting a global increase that could result from increased transcription. However, late conjugated SUMO2 / 3 levels outweigh non-conjugated levels showing a specific induction of SUMOylation (Fig 9a,b). Western blots were then analyzed by densitometry for quantification and statistical analyses. Endotoxin increased 2-fold 150KDa SUMO1 signal at 1.5h to return to a normal background at 3h with a slight late secondary increase around 24h (Fig.4a, Fig.9a). Endotoxemia also induced 100KDa SUMO1 signal at 1.5h with a progressive slight increase for up to 24h (Fig.4b). These early kinetics of SUMO1 at 100 and 150KDa mimic serum TNFα levels during endotoxemia. However, 150KDa, but not 100KDa, SUMO1 signal was also observed in control mice (without TNFα). Endotoxemia also induced a late 30-fold induction of 250KDa SUMO1 signal at 24 h (Fig.4c). SUMO2 / 3 also has a temporal substrate specificity. Endotoxemia induced an early 70KDa SUMO2 / 3 signal at 1.5h only (Fig.4d, Fig 9b).70KDa SUMO2 / 3 signal was not detected in control mice or late endotoxemia. The most significant effect of endotoxemia was the progressive late smear of 150-260KDa SUMO2 / 3 signal by 24h (Fig.4d). Quantitative comparative analyses show that the most significant effects of endotoxemia were early SUMO1 signals at 100 and 150KDa, and late p250 and early SUMO2 / 3 signal at 70KDa, and late 260KDa smear (Fig.4e,f, Fig.9c,d). Next, effects of TAK981 on these signals were investigated. TAK981 treatment inhibited all SUMOylation as confirmed by Western-blot analyses (Fig. 10a,b). These results show a distinct temporal and substrate specificity for splenic SUMO isoforms in response to bacterial endotoxin. Specific inhibition of SUMO1 unfettered TNFα production in endotoxemia. The specific SUMO isoform regulating immune responses were next investigated. SUMO1-null and SUMO3-null genetic mutant mice were bred. SUMO1-null and SUMO3-null mice are viable and survive through embryogenesis but SUMO2-null embryos die early during embryonic development and are not viable. Wild-type, SUMO1-null, and SUMO3-null mice had nearly undetectable serum TNFα levels in normal conditions, and wild-type and Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 SUMO3-null mice had similar responses to endotoxin (Fig.5a). By contrast, SUMO1-null mice exhibit unfettered TNFα production reminiscent of that found with global inhibition of SUMOylation with TAK981. SUMO1-null mice had almost 5-fold higher serum and splenic TNFα levels than wild-type endotoxemic mice (Fig.5a,b). These inflammatory responses were specific in the spleen because wild-type, SUMO1-null, and SUMO3-null mice all have statistically similar hepatic TNFα levels in control and endotoxemic mice (Fig.11a). Thus, specific SUMO1 inhibition unfetters splenic TNFα production and systemic inflammation during lethal endotoxemia. The specific role of SUMO1 and SUMO2 / 3 in HAGMA and kidney function markers were assessed. Endotoxemia induced similar hyperkalemia and creatinine levels in wild-type and SUMO3-null mice, but the latter had higher HAGMA and blood urea nitrogen (Fig.5c- f). Following TNFα responses, SUMO1-null mice also had higher hyperkalemia, HAGMA, blood urea nitrogen, and creatinine levels. Of note, hyperkalemia and creatinine levels mimicked TNFα responses, but not in SUMO3-null mice as they had similar TNFα responses than wild-type mice, but higher HAGMA and blood urea nitrogen. These results are specific to these markers and no differences were observed in hematocrit, natremia, or chloremia between wild-type, SUMO1-null, and SUMO3-null mice (Fig.10d-e). These results reveal a different role of SUMO1 and SUMO3 in regulating the immune and metabolic responses to lethal endotoxemia and show that specific SUMO1 inhibition increased serum TNFα levels and HAGMA. Adoptive transfer of SUMO1-null splenocytes unfettered TNFα production. The role of splenic SUMOylation was further analyzed with the adoptive transfer of splenocytes from SUMO1-null and SUMO3-null mice into splenectomized wild-type mice. Splenectomy attenuated serum TNFα levels and metabolic acidosis in lethal endotoxemia (Fig.6a-e). Then, a dose-response curve was generated to determine whether the transfer of wild-type splenocytes into splenectomized mice restores TNFα responses to endotoxin. The adoptive transfer of 1x107splenocytes intraperitoneally restored typical TNFα responses, HAGMA (hyperkalemia and anion gap), and kidney dysfunction (blood urea nitrogen and creatinine) in splenectomized endotoxemic mice (Fig.6a-e). These results were specific because the transferred splenocytes neither induced an effect by themselves nor affected the other markers, including hematocrit, natremia, or chloremia in control or endotoxic splenectomized mice (Fig.11a-d). The transfer of a higher number of splenocytes increased these responses in a concentration-dependent manner and started to induce unspecific responses. Next, the Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 role of SUMO1-null splenocytes was assessed by transferring splenocytes from SUMO1-null mice into wild-type splenectomized mice. Adoptive transfer of the of 1x107SUMO1-null splenocytes dramatically increased TNFα responses to bacterial endotoxin in splenectomized wild-type mice by 3-fold higher than the transfer of wild-type splenocytes (Fig.6a). This higher TNFα production induced by SUMO1-null splenocytes also triggered the responses observed in SUMO1-null mice, including higher hyperkalemia and anion gap levels. The transfer of SUMO1-null splenocytes also increased both blood urea nitrogen and creatinine in endotoxemic mice (Fig.6d,e). These results show that the specific transfer of SUMO1-null splenocytes is sufficient to recapitulate the immune and metabolic responses to lethal endotoxemia observed in SUMO1-KO mice. DISCUSSION. TAK981 is in clinical trials to potentiate immunotherapies in metastatic solid tumors and refractory hematologic malignancies such as large B cell lymphoma after CAR T-cells therapy. However, the effects of TAK981 in infectious disorders and sepsis are unknown, and the previous results of SUMOylation in infection are conflicting. As such, the effects of TAK981 in polymicrobial peritonitis induced by cecal ligation and puncture, the most clinically relevant experimental model of sepsis with a polymicrobial infection, were investigated herein. TAK981 treatment improved survival in mild polymicrobial peritonitis without antibiotics. These results also show that TAK981 treatment enhanced immune responses increasing both IFNγ and TNFα serum levels. TAK981 also increased TNFα levels and decreased bacterial counts in peritoneal lavage. Together, these results indicate that inhibition of SUMOylation with TAK981 potentiates immune responses that protect mice against mild bacterial infections. Splenic SUMOylation investigation revealed that TAK981 inhibits both SUMO1 and SUMO2 / 3. The results presented herein show that genetic depletion of SUMO1, but not SUMO3, mimics TAK981 treatment unleashing serum TNFα levels and HAGMA. These results show that specific SUMO1 inhibition is sufficient to unleash early complex inflammatory and metabolic responses to bacterial endotoxin. The transfer of SUMO1-null, but not wild-type, splenocytes into splenectomized wild-type mice potentiated the inflammatory responses and mimic the results observed in global SUMO1-null mice. Taken together, the results herein demonstrate that SUMOylation is an integral mechanism for Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 reprograming immune and metabolic responses to infections and a potential target for controlling infectious disorders. Example 2 The etiology of inflammatory bowel diseases (IBD), chronic debilitating conditions without cure that were originally regarded as psychosomatic disorders and shorten the lifespan of seven million patients worldwide by nearly 10%, is unknown. Herein it is shown that stress induces autonomic parasympathetic dysfunction that increases IBD susceptibility and mortality. Conversely, vagal stimulation restores neuromodulation and resolves colitis by inhibiting SUMOylation through a novel mechanism independent of the canonical IL10 / α7- cholinergic vagal pathway. Human colonic biopsies and experimental models show a significant increase in SUMO2 / 3 during colitis. Genetic inhibition of SUMO3 protects against colitis and delays the pathogenesis, whereas genetic depletion of SUMO1 halts the progression of colitis associated with immune cell influx. Bone marrow transplant from SUMO1-KO, but not SUMO3-KO, mice confer protection against colitis. Vagal stimulation inhibits both SUMOylation, delays the onset of colitis in SUMO1-KO mice, and protects SUMO3-KO mice. Treatment with TAK981, the first-in-class inhibitor of SUMOylation, rescues mice from established experimental UC, CD, and blocks the progression of acute colitis to sepsis. These results reveal a novel pathway of vagal neuromodulation to reprogram endogenous stress-adaptive responses through inhibiting SUMOylation. Taken together, these results reveal SUMO1 and SUMO3 as molecular markers and therapeutic targets for colitis stages and demonstrate that TAK981 can be used for treatment of IBD. MATERIAL AND METHODS Chemicals and reagents. Dextran sulfate sodium salt (DSS, MW 35-60KDa; MP biomedicals, Santa Ana, CA) was dissolved in autoclaved deionized water. 2,4,6- trinitrobenzene sulfonic acid (TNBS, Sigma; Saint Louis, MO) was used at a single dose of 150mg / kg (150µL, pr).4-Ethoxymethylene-2-phenyl-2-oxazolin-5-one (Oxazolone, Sigma; Saint Louis, MO) was used at 1% w / v in 50% ethanol (150µL, pr) 7 days after epicutaneous sensitization with 3% w / v Oxazolone in ethanol. Nicotine was dissolved in autoclaved drinking water at 20 µg / mL. Mice were treated with nicotine 3 days before and daily for 5 days after DSS treatment. TAK981 (ChemieTek, Indianapolis, IN) was dissolved in DMSO Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 (stock concentration 50 mg / mL, stored at -20°C) and diluted in pharmaceutical-grade corn oil (Sigma, St Louis, MO) at 7.5 mg / kg, and administered s.c.12 h before the start of colitis induction, then daily for 4 days. Animal experiments. Animal procedures were approved by the Institutional Animal Care & Use Committee of Duke University.6-8-week-old (∼25 ± 5 g) male C57BL / 6 male, IL10-KO mice, and α7nAChR-KO mice were obtained from The Jackson Laboratory (Bar Harbor, ME). SUMO1-, SUMO3-KO, and wild-type littermates’ mice were produced. Genotyping was performed by PCR using tail genomic DNA and the Extract-N-Amp Tissue PCR kit (Sigma Chemical, Saint Louis, MO). Mice were maintained on 12h light-dark cycle, with free access to food and water (ad libitum), randomized into experimental groups, and outcome assessors were blinded to group treatments. Restraint stress. Stress was induced by restraining the mice for 6 hours daily for 3 weeks in well-ventilated polypropylene tubes without physical compression or pain. Heart rate variability (HRV). Three stainless steel electrodes were implanted in the right and left pectoral muscles, and the abdominal wall using cardiac pacing wires (A & E Medical, Farmingdale, NJ),and tunneled under the skin. The electrodes were exteriorized from the mouse nape, and connected to MP150 research system (Biopac, Goleta, CA) for lead II electrocardiogram recording. Mice were allowed to recover for 5 days before recording lead II EKG. AcqKnowledge software was used for HRV frequency domain analyses based on the consecutive RR intervals of the 15 min EKG recordings. Low and high-frequency domains were calculated, and the ratio was used as an indicator of the sympathetic-vagal balance. Experimental IBD. Dextran Sulfate Sodium (DSS) colitis: In the initial experiments (Sup.Fig.S1a-h), 3 and 5% DSS in autoclaved drinking water were used for 5 days. Thereafter, all experiments were performed with 3% DSS. TNBS (2,4,6-trinitrobenzene sulfonic acid) colitis: Mice were fasted overnight and then given 150µL of TNBS (150mg / Kg) in 50% ethanol injected intra-rectally using polyurethane P20 tube (Becton Dickinson, Franklin Lakes, NJ) advanced gently 4 cm (from the anal verge) into the mouse rectum. The mice were held vertically for a minute after injection to make sure that TNBS was retained in the rectum. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 Oxazolone (4-Ethoxymethylene-2-phenyl-2-oxazolin-5-one) colitis was performed by shaving a 2x2 cm area of the abdomen, and pre-sensitizing mice with the percutaneous application of 150 µL of 3% oxazolone w / v in 100% ethanol. Seven days later, mice received 150 µL of 1% oxazolone w / v in 50% ethanol intrarectally, then held vertically for a minute to ensure the retainment of oxazolone in the rectum. In the three models of colitis, body weight, stool consistency, blood in stools by Hemoccult test (Hemoccult Sensa, Beckman Coulter, CA), and water consumption levels were recorded daily during the experiment to calculate Disease Activity Index (DAI). Intestinal permeability was analyzed with oral gavage of FITC dextran (0.6mg / g BW in PBS) 3 hours before terminal blood collection. Mice were euthanized on the indicated day, blood was collected by cardiac puncture and intestine was collected for further analysis. Disease Activity Index (DAI). DAI was recorded daily assessing preclinical parameters. Weight loss was scored (compared to baseline weight): 0, no loss or weight gain; 1, 5-10%; 2, 10-15%; 3, 15-20%; and 4, >20% loss from baseline weight. Stool consistency was scored: 0, well-formed stools; 1, sticky stools; 2, soft stools; 3, semi-solid loose stools; and 4, watery diarrhea. Rectal bleeding was scored: 0, no bleeding; 2, occult blood (by Hemoccult sensa, Beckman Coulter, CA); 3, bloody stools; and 4, gross rectal bleeding. Mice were immediately euthanized if they lost more than 20% of their baseline body weight. Vagal stimulation. Mice were anesthetized by 3-5% isoflurane, intubated, and connected to a ventilator to allow 45-55 mL / min airflow with 30% oxygen and 1-2% maintenance isoflurane. EKG leads were attached to the right and left forelimbs and hindlimbs to monitor heart rate. The front of the neck was shaved, and sterilized (70% ethanol and povidone-iodine) and a 2 cm longitudinal midline incision was made. Submandibular salivary glands were dissected and retracted laterally to expose the carotid artery and right cervical vagus nerve. The vagus nerve was carefully dissected from the carotid artery and connected to platinum-iridium microelectrodes. Biopac MP150 system was used to deliver continuous electrical stimulation (5V, 50 Hz, 100ms) for 15 minutes. Heart rate was monitored during the procedure to ensure effective vagal stimulation. After the stimulation, tissues were returned to anatomical positions, and the neck was closed with a 6 / 0 suture. Mice were monitored for 30 minutes in an incubation chamber to ensure full recovery before returning them to the cage. Sham mice were intubated, a longitudinal neck incision was performed, and the glands were retracted without touching the vagus nerve to avoid Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 mechanical stimulation. Vagal nerve stimulation was performed immediately before DSS treatment. Splenectomy. Mice were anesthetized with isoflurane 5% and subjected to an abdominal incision on the epigastrium and mesogastrium. The spleen was exposed by gentle retraction of the stomach to the side. The three main branches of the splenic artery were stabilized with nylon thread, ligated, and cut. The spleen was removed, and the abdominal wall was sutured with catgut and the skin with nylon thread. Mice were splenectomized 5 days before the experimental procedure. Peritoneal lavage and bacterial load analysis. Peritoneal lavage was performed. Six hours before euthanasia, mice were anesthetized with 5% isoflurane, the abdominal wall was disinfected with 70% ethanol, and 5 mL of sterile pyrogen-free PBS was injected i.p. using a 25G needle. Then, the abdomen was massaged for one minute, and 1 mL of peritoneal fluid was aspirated using a sterile syringe. Peritoneal aspirates were centrifuged at 2,000g for 20 min at 4°C and supernatants were frozen at -20°C for further cytokines analysis. Peritoneal lavage bacterial load was analyzed. Freshly collected peritoneal aspirates were cultured in blood agar using a sterile inoculation loop and agar plates were incubated at 37°C for 24 hours. Bacterial colonies (CFU / mL of peritoneal aspirate) were counted using an ImageJ Fiji software cell analyzer. Statistical analyses. Statistical analyses were performed with GraphPad Prism Software (GraphPad Software, San Diego, CA). The sample size was calculated based on our preliminary results using G*Power3.1 and ANOVA (Fixed effects, omnibus, one-way), assuming effect size= 0.25, α=0.05, power (1-β prob Type II error) =0.8. Data are expressed as mean±SEM and the figures represent experiments repeated twice on different days. The student’s t-test or Mann-Whitney U test was used to compare two experimental groups. Three or more groups were analyzed with parametric one-way ANOVA with multiple pair-wise comparisons. Normality and homogeneity of variance were confirmed with Kolmogorov- Smirnov analyses. Pair comparisons in nonparametric ANOVA tests were adjusted post-hoc with the Tukey test (in equal sample sizes) or Bonferroni’s for multiple hypothesis testing. The time courses and pair-wise comparisons were analyzed with the two-way ANOVA for repeated measures. Statistical significance was established at p≤.05 with a 2-sided alpha. Figures show representative data of experiments that were replicated on different dates. Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 RESULTS Vagal stimulation rescued mice from stress-induced autonomic dysfunction and IBD mortality. Restraint stress increased mice susceptibility to Dextran Sulfate Sodium (DSS) colitis and turned colitis lethal with all mice dead by day 5 after the challenge (Fig. 12a). There were no major changes in body weight between the groups, but the main differences were observed in intestinal bleeding and stool consistency worsening the overall multivariable colitis Disease Associate Index (DAI) in the stress mice (Fig.12b). Given that death prevented statistical group comparison, the figures show the time of death, and control DAI was monitored to confirm normal time course responses. Then, the three major networks of neuromodulation were analyzed in stress and control mice and found no significant changes in plasma corticosterone levels (examining the hypothalamic-pituitary- adrenal (HPA) axis) or norepinephrine (examining the sympathetic system) but there was a 3- fold increase in the low / high frequency (LF / HF) ratio, an established clinical approach to assess heart rate variability (HRV) and the parasympathetic tone (Fig.12c). Since high LF / HF reflects a parasympathetic deficiency, it was analyzed whether electrical stimulation of the parasympathetic vagus nerve could compensate for these effects and improve colitis outcome. Vagal stimulation decreased the ratio LF / HF (without affecting corticosterone or epinephrine) and significantly improved DAI in the colitis mice (Fig.12d-f). These results are consistent with the literature showing that stool consistency and intestinal bleeding / permeability are the first symptoms as reflected in early DAI scores before body weight changes are noticeable normally after day 5 of DSS treatment. Observers recording DAI and bleeding were blind to the treatments and alternated on different days. Vagal stimulation also rescued normal mice (without stress) and prevented colitis-induced intestinal bleeding, DAI, body weight loss, and intestinal permeability (Fig.12g-k). Vagal stimulation preserved intestinal integrity at macro and microscopic levels (Fig.12k-m) and damped inflammatory cytokines IL6 and TNFα but increased anti-inflammatory cytokine IL10 levels (Fig.12n). Since these results suggested that vagal stimulation improves colitis by inducing IL10, IL10-KO mice were assessed. IL10-KO mice were highly susceptible to DSS colitis with conspicuous intestinal bleeding within the first day (Fig.13a). Wild-type and IL10-KO mice had similar colon permeability and inflammatory markers at euthanasia on day 7 (Fig. 13c,d). Vagal stimulation completely protected IL10-KO mice from colitis during the first 4 days and prevented body weight loss and intestinal permeability without inhibiting IL6 or Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 TNFα by euthanasia at day 7 (Fig.13e-h). These results showed that vagal protection against colitis was independent of IL10. Splenectomy prevents vagal protection against colitis. The mechanism of vagal regulation of colitis is unknown as most of the colon lacks efferent vagal innervation. The spleen may be a link between the nervous and immune systems. Surgical splenectomy did not affect early stool consistency and intestinal bleeding but improved DAI scores after day 5 (Fig.13i). Splenectomy improved colon integrity, body weight, and intestinal permeability, and reduced colon IL6 and TNFα inflammatory responses by 50% without affecting IL10 levels (Fig.13j-l). Moreover, splenectomy prevented the protective effects of vagal stimulation in DAI, colon length, permeability, and immune cytokines (Fig.13m-p). Vagal stimulation controls colitis independently of α7-nicotinic acetylcholine receptors (α7nAChRs). It was next assessed whether α7nAChRs contribute to the vagal regulation of colitis. α7nAChR-KO mice had DAI scores like wild-type mice but lower body weight loss and intestinal permeability like splenectomized mice (Fig.14a-e). α7nAChR-KO mice had lower TNFα but similar IL6 and IL10 colon levels that wild-type mice (Fig.14f). Of note, vagal stimulation still improved DAI scores after day 4 in α7nAChR-KO mice (Fig. 14g). Although a significant effect on body weight loss and intestinal permeability was not detected, vagal stimulation reduced IL6 and TNFα but not IL10 colon levels in α7nAChR- KO mice on day 7 (Fig.14g-l). These results show that vagal stimulation induces two protective mechanisms, one dependent on α7nAChRs to induce IL10 and partially inhibition of TNFα and a second α7nAChR-independent mechanism to inhibit IL6 and TNFα. Since acetylcholine, the principal neurotransmitter of the vagus nerve, signals through nicotinic receptors, it was next analyzed whether nicotine mimics vagal neuromodulation of colitis. Treatment with nicotine was significantly less effective than vagal stimulation and did not improve DAI scores during the first 4 days, but slightly prevented intestinal bleeding and body weight loss on days 6 and 7 (Fig.15a-c). By euthanasia on day 7, nicotine prevented intestinal permeability and inhibited IL6 and TNFα by over 65% without affecting IL10 (Fig. 15d,e). Next, it was analyzed whether the effects of nicotine were mediated by α7nAChR using α7nAChR-KO mice. Nicotine did not improve DAI, intestinal bleeding, body weight, colon length, or intestinal permeability in α7nAChR-KO mice (Fig.15f-i). Surprisingly, nicotine inhibited colon levels of IL6 and TNFα in α7nAChR-KO mice (Fig.15j). These Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 results show that nicotine protects against inflammatory cytokines in colitis through a novel anti-inflammatory mechanism independent of α7nAChRs. SUMOylation in human biopsies and vagal regulation of colitis. The results in Example 1 demonstrate that SUMOylation is an important mechanism that orchestrates immune responses to bacterial infection. Thus, Ubc9 (the sole E2-conjugating enzyme of all SUMOylation) transcripts in the mRNA databank of human biopsy samples was assessed. There was no statistically significant difference between the Ubc9 transcripts of 91 CD patients and 29 non-IBD controls (p=0.34, Fig.16a). Next, different SUMO isoforms were investigated. There are three SUMO1-3 isoforms and SUMO2 and SUMO3 are referred to together as SUMO2 / 3 due to their homology.18 human samples from intestinal biopsies and surgical resection were collected to analyze SUMOylation by Western blot and immunohistochemistry. Human colonic biopsies did not show major changes in SUMO1 but a strong increase in SUMO2 / 3 levels in CD patients as shown by the high molecular “smear” representative of polySUMOylation of multiple proteins as compared to non-IBD patients (Fig.16b). Immunohistochemistry analyses of human biopsies provided similar results with a stronger SUMO2 / 3 staining in CD patients (Fig.16c). Western blots of colon samples from DSS mice showed similar pattern without major changes in SUMO1 but over 2-fold increase in SUMO2 / 3 smear ≈ 250KDa (Fig.16d). These results were consistent with the immunohistochemistry analyses showing a stronger and broader distribution of SUMO2 / 3 in the colon of colitis mice (Fig.16e). Vagal stimulation decreased SUMO1 levels by nearly 30% and strongly inhibited SUMO2 / 3 levels by 70% (Fig.16d,e). SUMO isoforms regulate different stages of colitis. The specific role of SUMO isoforms were analyzed in colitis and vagal neuromodulation by using SUMO1-KO and SUMO3-KO mice. SUMO2-KO mice were not analyzed because embryos die during early development. SUMO1-KO DAI scores plateaued on day 3 and mice were thereafter resistant to the progression of colitis (Fig.17a) By contrast, SUMO3-KO mice were very resistant to colitis during the first 5 days. Both SUMO1- and SUMO3-KO mice were more resistant to intestinal permeability and had lower IL6 and TNFα colon levels on day 7 (Fig.17b-d). Vagal stimulation protected SUMO1-KO mice to early colitis improving DAI until day 5 (Fig.17e). By contrast, vagal stimulation improved DAI scores in SUMO3-KO mice after day 4 (Fig.17i). Vagal stimulation prevented intestinal permeability in SUMO1-KO and SUMO3-KO mice by 65% and 35%, respectively (Fig.17g,k). By day 7, vagal stimulation also increased colon IL6 and IL10 levels in SUMO1-KO mice but only IL10 levels in Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 SUMO3-KO mice by nearly 4-fold (Fig.17h,l). The protection of SUMO1-KO mice after day 3 matches the timing of immune cell influx into the intestine, and suggests that protection induced by SUMO1 depletion may be due to immune cells. This hypothesis was tested by performing bone marrow transplants from wild-type, SUMO1-KO, and SUMO3-KO mice into irradiated wild-type mice. The transfer of wild-type and SUMO3-KO bone marrow restored normal DAI scores, body weight loss, intestinal permeability, and immune responses in irradiated wild-type mice (Fig.6m-p). However, the transfer of SUMO1-KO, but not SUMO3-KO, bone marrow significantly protected irradiated wild-type mice against DAI, body weight loss, permeability, and inflammatory responses while increasing IL10 production by nearly 3-fold. These results revealed a fundamental difference between SUMO1 and SUMO3 in the pathogenesis and progression of colitis. These results also show that vagal stimulation mimics early and late inhibition of SUMO3 and SUMO1, suggesting that it protects against colitis by inhibiting SUMOylation. Since vagal stimulation is an invasive surgical procedure, it was next tested whether pharmacological inhibition of SUMOylation mimics vagal stimulation and protects against colitis while avoiding nerve manipulation. Inhibition of SUMOylation with TAK981 rescued mice from established colitis. SUMOylation was inhibited using TAK981 (Subasumstat) which is the recently discovered first-in-class SUMO inhibitor that specifically inhibits the transfer of SUMO to the E2 conjugating enzyme Ubc9. First, it was confirmed that TAK981 inhibits both SUMO1 and SUMO2 / 3 levels in the colon of DSS mice (Fig.18). It was also confirmed that TAK981 inhibits SUMOylation in the spleen of endotoxemic mice. The specificity of the staining was confirmed in extracts from SUMO1-KO and SUMO3-KO mice. TAK981 protected against colitis, improved DAI through the time course, preserved the colon's macroscopic and histological integrity, and almost completely prevented intestinal permeability by nearly 90% (Fig.19a-e). TAK981 inhibited colon IL6 and TNFα by 80% and 65%, respectively without affecting IL10 levels (Fig.19f). Whether TAK981 treatment can rescue mice from established colitis was next investigated. TAK981 treatment, started three days after the DSS challenge, rescued mice from established colitis, and improved DAI by preventing intestinal bleeding and body weight loss (Fig.20a-c). TAK981 also improved body weight, DAI, and intestinal integrity in experimental CD induced by TNBS (2,4,6- trinitrobenzene sulfonic acid, Fig.20d-f). These results led to the question of whether TAK981 may preserve intestinal integrity and Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 inhibit colitis progression to sepsis, a lethal complication in IBD patients. Thus, the potential of TAK981 in oxazolone-induced colitis, an experimental model of severe ulcerative colitis associated with mucosal damage that progresses to sepsis, was investigated. TAK981 treatment improved DAI, prevented intestinal bleeding, body weight loss, and preserved intestinal integrity (Fig.20g-k). TAK981 treatment was especially effective preventing intestinal permeability and peritoneal bacterial leakage by nearly 90% (Fig.20l). These results correlated with the potential of TAK981 to inhibit inflammatory cytokines IL6 and TNFα by nearly 70% without affecting IL10 (Fig.20m). Together, these results demonstrate that inhibition of SUMOylation with TAK981 can mimic vagal regulation of SUMOylation during colitis and might provide clinical advantages for treating IBD (Fig.21). DISCUSSION The results presented herein show for the first time the different roles of SUMO isoforms in vivo, with SUMO1-KO mice being resistant to colitis progression, whereas SUMO3-KO mice are resistant to the early stages of colitis. This timing matches the time of immune cells influx into the intestine around day 3 and prompted bone marrow transplants from SUMO1-KO and SUMO3-KO mice into wild-type mice. The transplant of SUMO1- null, but not SUMO3-null, bone marrow conferred protection to irradiated wild-type mice against colitis. These results demonstrate a fundamental difference between SUMO1 and SUMO3 isoforms and their contribution to colitis. The typical high molecular weight smears observed in CD patients and murine models reflect polySUMOylation of multiple proteins. Indeed, SUMOylation controls multiple pathways at different levels by targeting several proteins. The results herein show for the first time that SUMOylation is regulated by neuromodulation and that vagal stimulation is a potential strategy to control SUMOylation in colitis. Vagal stimulation did not induce a major effect on SUMO1 but significantly decreased SUMO2 / 3. Vagal stimulation improved early colitis like those results observed in SUMO3-KO mice and improved late colitis reminiscent of SUMO1-KO mice. Since these results suggested that vagal stimulation protects against colitis by inhibiting SUMOylation, it was tested whether pharmacological inhibition of SUMOylation mimics vagal stimulation and protects against colitis. This strategy is clinically relevant because pharmacologic inhibition of SUMOylation can provide major clinical advantages to prevent surgical nerve manipulation and improve treatment efficacy as compared to surgical vagal stimulation. TAK981 (Subasumstat) is the recently discovered first-in-class SUMO inhibitor that specifically inhibits the transfer of SUMO to the E2 conjugating enzyme UB9. TAK981 is Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 currently in a dose-escalation Phase 1 / 2 clinical study to evaluate its safety, tolerability, and pharmacokinetics to potentiate immunotherapies in patients with advanced or metastatic solid tumors or relapsed / refractory hematologic malignancies such as diffuse large B cell lymphoma after CAR T-cells therapy (Clinicaltrials.gov, NCT03648372, Estimated completion date: October 2024). TAK981 treatment improved both DSS experimental colitis and TNBS experimental CD. From a clinical perspective, TAK981 treatment not only prevented but also rescued mice from established colitis even when treatment was started three days after the beginning of the DSS challenge when mice already have severe symptoms of colitis including intestinal bleeding. TAK981 treatment also prevented the most detrimental effects of IBD including intestinal permeability and bacterial leakage progressing to sepsis in oxazolone-induced colitis, an experimental model of severe ulcerative colitis associated with mucosal damage that progresses to sepsis. Taken together, the results herein demonstrate that SUMOylation is a promising therapeutic target and TAK981 can provide therapeutic advantages for IBD patients and decrease the risk of sepsis. TAK981 treatment may provide clinical for those patients with severe IBD unresponsive to vagal stimulation and to avoid costly surgical nerve manipulation. These results further support the design of comprehensive strategies to control SUMOylation and reprogram the immune system through nerve stimulation and complementary pharmacologic treatment.

Claims

Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 CLAIMSWhat is claimed is:

1. A method of treating or preventing inflammatory bowel disease (IBD) in a subject, the method comprising inhibiting SUMOylation in the subject.

2. The method of claim 1, wherein inhibiting SUMOylation in the subject comprises performing vagal nerve stimulation on the subject and / or providing to the subject an inhibitor of SUMOylation.

3. The method of claim 2, comprising providing to the subject an inhibitor of SUMOylation.

4. The method of claim 3, wherein the inhibitor of SUMOylation inhibits SUMO maturation, SUMO activation, SUMO conjugation, SUMO ligation, and / or stimulates SUMO hydrolysis.

5. The method of claim 3, wherein the inhibitor of SUMOylation is a SUMO E1 inhibitor, a SUMO E2 inhibitor, a SUMO E3 inhibitor, or a SENP inhibitor.

6. The method of claim 3, wherein the inhibitor of SUMOylation is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, tannic acid, spectomycin B, viomellein, chaetochromin A, D-208, compound 22, momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, atromentin, compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 12, compound 13 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor), ML-792, TAK-981, compound 23 (GSK145A), compound 24, compound 25, compound 26, compound 27, compound 34 (JCP-666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), compound 38 (VAE-561), compound 39, compound 40 (GN6767), compound 41 (GN6958), compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI-02), compound 46, compound 47, compound 48, compound 49, compound 50, compound 51, compound 52, compound 53, compound 54, a camptothecin derivative, topotecan, an ortho-quinone, nocardioneClient Ref. No. DU7335US Atty. Docket No. Duke-42465.202 A, 33-DINOR-dunnione, 33-DINOR-dehydrodunnione, β-lapachone, an iminoquinone, macrophilone A (compound 60) compound 61, a diterpene, triptolide, or compound 63.

7. The method of claim 3, wherein the inhibitor of SUMOylation is TAK-981.

8. The method of claim 7, comprising providing to the subject the TAK-981 intravenously at a dose of 10 µg / kg to 5 mg / kg.

9. A method of treating or preventing sepsis in a subject, the method comprising performing vagal nerve stimulation on the subject and / or providing to the subject an inhibitor of SUMOylation.

10. The method of claim 9, comprising preventing sepsis in a subject, wherein the subject has an infection and is at risk of experiencing sepsis.

11. The method of claim 9, comprising treating sepsis in a subject.

12. The method of any one of claims 9-11, comprising providing to the subject an inhibitor of SUMOylation.

13. The method of claim 12, wherein the inhibitor of SUMOylation inhibits SUMO maturation, SUMO activation, SUMO conjugation, SUMO ligation, and / or stimulates SUMO hydrolysis.

14. The method of claim 12, wherein the inhibitor of SUMOylation is a SUMO E1 inhibitor, a SUMO E2 inhibitor, a SUMO E3 inhibitor, or a SENP inhibitor.

15. The method of claim 12, wherein the inhibitor of SUMOylation is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, tannic acid, spectomycin B, viomellein, chaetochromin A, D-208, compound 22, momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, atromentin, compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 12, compound 13Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor), ML-792, TAK-981, compound 23 (GSK145A), compound 24, compound 25, compound 26, compound 27, compound 34 (JCP-666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), compound 38 (VAE-561), compound 39, compound 40 (GN6767), compound 41 (GN6958), compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI-02), compound 46, compound 47, compound 48, compound 49, compound 50, compound 51, compound 52, compound 53, compound 54, a camptothecin derivative, topotecan, an ortho-quinone, nocardione A, 33-DINOR-dunnione, 33-DINOR-dehydrodunnione, β-lapachone, an iminoquinone, macrophilone A (compound 60) compound 61, a diterpene, triptolide, or compound 63.

16. The method of claim 12, wherein the inhibitor of SUMOylation is TAK-981.

17. The method of claim 16, comprising providing to the subject the TAK-981 intravenously at a dose of 10 µg / kg to 5 mg / kg.

18. The method of any one of the preceding claims, wherein the subject is a human.

19. An inhibitor of SUMOylation for use in a method of treating or preventing IBD or sepsis in a subject.

20. The inhibitor of claim 19, wherein the inhibitor is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, tannic acid, spectomycin B, viomellein, chaetochromin A, D-208, compound 22, momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, atromentin, compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 12, compound 13 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor), ML-792, TAK-981, compound 23 (GSK145A), compound 24, compound 25, compound 26, compound 27, compound 34 (JCP-666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), compound 38 (VAE-561), compound 39, compound 40 (GN6767), compound 41 (GN6958), compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI- 02), compound 46, compound 47, compound 48, compound 49, compound 50,Client Ref. No. DU7335US Atty. Docket No. Duke-42465.202 compound 51, compound 52, compound 53, compound 54, a camptothecin derivative, topotecan, an ortho-quinone, nocardione A, 33-DINOR-dunnione, 33-DINOR- dehydrodunnione, β-lapachone, an iminoquinone, macrophilone A (compound 60) compound 61, a diterpene, triptolide, or compound 63.

21. An inhibitor of SUMOylation for use in the manufacture of a medicament for the treatment or prevention of IBD or sepsis in a subject.

22. The inhibitor of claim 21, wherein the inhibitor is ginkgolic acid, anacardic acid, kerriamycin B, daviddin, tannic acid, spectomycin B, viomellein, chaetochromin A, D-208, compound 22, momordin Ic, streptonigrin, NSC76919, NSC45384, vialinin A, atromentin, compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 12, compound 13 (CID9549553), compound 14 (COH000), compound 15 (quinoxaline SAE inhibitor), ML-792, TAK-981, compound 23 (GSK145A), compound 24, compound 25, compound 26, compound 27, compound 34 (JCP-666), compound 35, compound 36 (VAE-499), compound 37 (VAE-500), compound 38 (VAE-561), compound 39, compound 40 (GN6767), compound 41 (GN6958), compound 42, compound 43, compound 44 (SPI-01), compound 45 (SPI- 02), compound 46, compound 47, compound 48, compound 49, compound 50, compound 51, compound 52, compound 53, compound 54, a camptothecin derivative, topotecan, an ortho-quinone, nocardione A, 33-DINOR-dunnione, 33-DINOR- dehydrodunnione, β-lapachone, an iminoquinone, macrophilone A (compound 60) compound 61, a diterpene, triptolide, or compound 63.

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