Treatment of major CTLA-4 checkpoint-associated immunodeficiency disorders with 1H-indole-3-carboxyaldehyde or 1-methylrindole-3-carboxylic acid

Abstract

This disclosure relates to a method for treating CTLA-4 checkpoint-associated immunodeficiency, comprising administering a therapeutically effective amount of a pharmaceutical composition comprising 1H-indole-3-carboxyaldehyde (3-IAld) or 1-methylindole-3-carboxylic acid.

Description

[Technical Field]

[0001] Cross-reference of related applications This international application claims priority to U.S. Provisional Patent Application No. 63 / 509,223, filed on 20 June 2023, which is incorporated herein by reference in its entirety.

[0002] field This disclosure relates to a method for treating CTLA-4 checkpoint-related immunodeficiency disorder in patients. [Background technology]

[0003] Primary CTLA-4 checkpoint-associated immunodeficiency is characterized by various combinations of enteropathy, hypogammaglobulinemia, recurrent respiratory infections, granulomatous lymphocytic interstitial lung disease, lymphocyte infiltration of non-lymphoid organs (intestines, lungs, brain, bone marrow, and kidneys), autoimmune thrombocytopenia or neutropenia, autoimmune hemolytic anemia, and lymphadenopathy. Heterozygous CTLA4 mutations in humans are associated with severe immunodysregulation, known as CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI) disease. Another type of primary CTLA-4 checkpoint-associated immunodeficiency has been reported in the literature in patients carrying bi-allele mutations in the "lipopolysaccharide-responsive beige-like anchor (LRBA)" gene. LRBA deficiency also leads to secondary loss of CTLA-4, so this recessive disorder is called "LRBA deficiency with autoantibodies, T-reg cell deficiency, autoimmune infiltration, and enteropathy (LATAIE)."

[0004] Currently, hematopoietic stem cell transplantation (HSCT) is the only treatment option for patients with primary CTLA-4 checkpoint-associated immunodeficiency. All other proposed treatments remain largely unsatisfactory, with the exception of immunoglobulin replacement therapy to correct hypogammaglobulinemia and prevent infections.

[0005] Systemic corticosteroids provided temporary relief from gastrointestinal symptoms in some patients.

[0006] Treatment with abatacept or sirolimus is recommended to control lymphocyte activation in all patients with CTLA-4-associated CNS syndrome. High-dose steroids remain suggested as first-line treatment, as is recommended for other demyelinating CNS disorders. Adding high-dose IVIG appears worthwhile if a rapid clinical response is not observed. As a second-line strategy, rituximab or cyclophosphamide should be considered while continuing first-line therapy, as is recommended for autoimmune encephalitis.

[0007] In conclusion, systemic immunosuppressants and abatacept may provide partial control, but require continuous administration. Despite the significant risk of treatment-related mortality, allogeneic hematopoietic stem cell transplantation offers a possible treatment option for patients with CTLA-4 dysfunction who are eligible for this approach. Another important issue is the need for multiple changes in immunosuppressants due to side effects or steroid dependence. However, untreated or inadequately treated patients require treatment as they develop further CTLA-4-related symptoms, suggesting a progressive and natural course of the disease.

[0008] Furthermore, while cancer immunotherapy using immune checkpoint inhibitors has been highly successful, its therapeutic effects are limited by various resistance mechanisms (Schoenfeld et al, 2020) or associated toxic effects, including frequent gastrointestinal, endocrine, and cutaneous toxicity, as well as potentially fatal neurotoxicity and cardiotoxicity (Choi et al, 2020). Therefore, novel therapeutic strategies that offer manageable side effects to existing immunotherapies will enhance and expand their therapeutic efficacy and applications. [Overview of the project]

[0009] This disclosure provides, at least in part, a method for treating CTLA-4 checkpoint-associated immunodeficiency in a patient, comprising administering a therapeutically effective amount of a pharmaceutical composition containing 1H-indole-3-carboxyaldehyde (3-IAld) to a patient in need thereof. In some embodiments, CTLA-4 checkpoint-associated immunodeficiency is selected from the group consisting of: CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI), lipopolysaccharide-responsive beige-like anchor (LRBA) deficiency (LATAIE), regulatory T (Treg) cell deficiency, autoimmune infiltration, intestinal disease, enteritis, immune-mediated colitis, gastrointestinal disorders, and gastric atrophy. In another embodiment, CTLA-4 checkpoint-associated immunodeficiency is immune-mediated colitis.

[0010] In one embodiment, the pharmaceutical composition comprises a pharmaceutically acceptable polymer. In another embodiment, the pharmaceutically acceptable carrier is at least one polymer. In another embodiment, the pharmaceutically acceptable carrier is a group of polymers. In another embodiment, the group of polymers is Eudragit® polymers. In another embodiment, the pharmaceutical composition is formulated for intestinal delivery. In another embodiment, the pharmaceutical composition is administered orally. In another embodiment, the pharmaceutical composition is in the form of capsules, tablets, gel tablets, gel capsules, gels, liquids, or gums. In another embodiment, the pharmaceutical composition is in the form of tablets or capsules.

[0011] In one embodiment, the pharmaceutical composition is administered at intervals of every other day (qod). In another embodiment, the pharmaceutical composition is administered in 3-IAld doses of at least about 3 mg / kg, at least about 4 mg / kg, at least about 5 mg / kg, at least about 6 mg / kg, at least about 7 mg / kg, at least about 8 mg / kg, at least about 9 mg / kg, at least about 10 mg / kg, at least about 11 mg / kg, at least about 12 mg / kg, at least about 13 mg / kg, at least about 14 mg / kg, at least about 15 mg / kg, at least about 16 mg / kg, at least about 17 mg / kg, or at least about 18 mg / kg. In another embodiment, the 3-IAld dose is about 18 mg / kg.

[0012] This disclosure provides, at least in part, a method for treating CTLA-4 checkpoint-associated immunodeficiency in a patient, comprising administering a therapeutically effective amount of a pharmaceutical composition containing 1-methylindole-3-carboxylic acid to a patient in need thereof. In some embodiments, CTLA-4 checkpoint-associated immunodeficiency is selected from the group consisting of: CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI), lipopolysaccharide-responsive beige-like anchor (LRBA) deficiency (LATAIE), regulatory T (Treg) cell deficiency, autoimmune infiltration, intestinal disease, enteritis, immune-mediated colitis, gastrointestinal disorders, and gastric atrophy. In another embodiment, CTLA-4 checkpoint-associated immunodeficiency is immune-mediated colitis.

[0013] In one embodiment, the pharmaceutical composition comprises a pharmaceutically acceptable carrier. In another embodiment, the pharmaceutically acceptable carrier is at least one polymer. In another embodiment, the pharmaceutically acceptable carrier is a group of polymers. In another embodiment, the group of polymers is Eudragit® polymers. In another embodiment, the pharmaceutical composition is formulated for intestinal delivery. In another embodiment, the pharmaceutical composition is administered orally. In another embodiment, the pharmaceutical composition is in the form of capsules, tablets, gel tablets, gel capsules, gels, liquids, or gums. In another embodiment, the pharmaceutical composition is in the form of tablets or capsules. [Brief explanation of the drawing]

[0014] [Figure 1] This represents the chemical structure of 1H-indole-3-carboxyaldehyde, also known as indole-3-aldehyde or 3-formylindole (3-IAld, MF:C9H7NO, IUPAC name:1H-indole-3-carbaldehyde). [Figure 2-1]Figures 2A–2H show graphs and photographs demonstrating that 3-IAld protects against inflammatory lesions in DSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and 3-IAld as shown in the experimental schedule (Figure 2A). Mice were evaluated for survival% (Figure 2B), weight change% (Figure 2C), disease activity index (Figure 2D), rectal bleeding (Figure 2E), colon histology by PAS staining (Figure 2F), histological score (Figure 2G), and epithelial barrier function (ZO-1 and Ki-67 staining) (Figure 2H). Photographs were taken with a high-resolution microscope at 20x magnification (scale bar, 200 μm). White arrows indicate rectal bleeding. Yellow arrows indicate inflammatory cell recruitment. Each in vivo experiment included 4–6 mice per group (16–24 mice in each experiment). Data are expressed as mean ± SEM. H2O, untreated mice. ***P<0.001, ****P<0.0001. [Figure 2-2] Figures 2A–2H show graphs and photographs demonstrating that 3-IAld protects against inflammatory lesions in DSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and 3-IAld as shown in the experimental schedule (Figure 2A). Mice were evaluated for survival% (Figure 2B), weight change% (Figure 2C), disease activity index (Figure 2D), rectal bleeding (Figure 2E), colon histology by PAS staining (Figure 2F), histological score (Figure 2G), and epithelial barrier function (ZO-1 and Ki-67 staining) (Figure 2H). Photographs were taken with a high-resolution microscope at 20x magnification (scale bar, 200 μm). White arrows indicate rectal bleeding. Yellow arrows indicate inflammatory cell recruitment. Each in vivo experiment included 4–6 mice per group (16–24 mice in each experiment). Data are expressed as mean ± SEM. H2O, untreated mice. ***P<0.001, ****P<0.0001. [Figure 3]Figures 3A–3H are graphical representations showing that 3-IAld provides epithelial barrier integrity and promotes an anti-inflammatory state in mouse colitis. As shown in Figure 2, C57BL / 6 mice subjected to DSS plus anti-CTLA-4 induced colitis and administered 3-IAld were evaluated for epithelial function markers (Figure 3A); dextran-FITC (Figure 3B) and serum sCD14 levels (Figure 3C); cytokine (Figure 3D) and calprochitin levels in colon homogenate (Figure 3E); and AhR-dependent gene expression (Figures 3F, 3G, 3H). Data are expressed as mean ± SEM. H2O, untreated mice. *P<0.05, **P<0.01, ***P<0.001. ns, not significant. [Figure 4] Figures 4A–4F show graphs and photographs illustrating that 3-IAld protects against immune-mediated colitis. NSG mice were injected with hPBMC and treated with anti-CTLA-4 mAb and 3-IAld. Mice were sacrificed after 21 days and evaluated for survival% (Figure 4A), weight change% (Figure 4B), colon histology (Figure 4C) (periodate-Schiff staining) and histological score (Figure 4D), ZO-1 protein expression (Figure 4E), and inflammatory cytokine expression (Figure 4F). Photographs were taken using a high-resolution microscope (Olympus BX51) at 20x magnification (scale bar, 200 μm). Each in vivo experiment included 4–6 mice per group (16–24 mice per experiment). Data are expressed as mean ± SEM. *P<0.05, **P<0.01, ****P<0.0001. Two-way ANOVA, Bonferroni post-hoc test. H2O, untreated mice. [Figure 5]Figures 5A-5D are graphical representations and photographic images of how 3-IAld restricts the progression of immune-mediated colitis in Rag1-deficient mice. Rag1- / - mice injected with CD4+ T cells were treated with αCTLA-4 mAb and 3-IAld. Mice were sacrificed on day 21, and percent weight change (Figure 5A), gross histology (Figure 5B), histological score (Figure 5C), and colonic histology (periodic acid-Schiff staining) (Figure 5D) were evaluated. Photographs were taken with a high-resolution microscope at 20× magnification (scale bar, 200 μm). For histology, the data represent two independent experiments. Each in vivo experiment included 4 mice per group (16 mice per experiment). Data are presented as mean ± SEM. **P<0.01, ****P<0.0001. Two-way ANOVA, Bonferroni's post-test. H2O, untreated mice. [Figure 6] Figures 6A-6C are graphical representations and photographic images showing how 3-IAld protects from DSS+ anti-CTLA-4-induced colitis in IL-10- / - mice. Mice were treated with DSS in drinking water for 1 week, followed by a 1-week recovery period, and then 100 μg of anti-CTLA-4 mAb and 3-IAld were administered as shown in Figure 2A. Mice were evaluated for percent weight change (Figure 6A), colonic histology (PAS staining) (Figure 6B), and histological score (Figure 6C), and photographs were taken with a high-resolution microscope at 20× magnification (scale bar, 200 μm). Data are presented as mean ± SEM. H2O, untreated mice. *P<0.05, **P<0.01. [Figure 7] Figures 7A-7B are graphical representations and photographic images showing how 3-IAld protects IL-10- / - mice from pathology after short-term treatment with anti-CTLA-4 monoclonal antibody. Mice were administered 100 μg of anti-CTLA-4 mAb and 3-IAld as described above and evaluated for percent weight change (Figure 7A) and colonic histology (PAS staining) (Figure 7B). Photographs were taken with a high-resolution microscope at 20× magnification (scale bar, 200 μm). [Figure 8-1]Figures 8A - 8H are graphical representations and photographic images showing how 3 - IAld protects IL - 10 - / - mice from the pathology after long - term treatment with anti - CTLA - 4 monoclonal antibody. Control C56BL / 6 and IL - 10 - deficient mice were administered 100 μg of anti - CTLA - 4 mAb and 3 - IAld as shown in the experimental schedule (Figure 8A). The mice were evaluated for body weight change (Figure 8B), disease activity index (Figure 8C), clinical examination (Figure 8D), colon (Figure 8E) and histology (PAS staining) (in the inset, ZO - 1 and BrdU staining) (Figure 8F), calprotectin levels in intestinal homogenates (Figure 8G), and cytokine and defensin gene expression in the intestine (Figure 8H). Photographs were taken at high - resolution microscopy, 20 - fold magnification (scale bar, 200 μm). Data are presented as mean ± SEM (pooled from two experiments). H2O, untreated mice. *P < 0.05, **P < 0.01. [Figure 8-2] Figures 8A - 8H are graphical representations and photographic images showing how 3 - IAld protects IL - 10 - / - mice from the pathology after long - term treatment with anti - CTLA - 4 monoclonal antibody. Control C56BL / 6 and IL - 10 - deficient mice were administered 100 μg of anti - CTLA - 4 mAb and 3 - IAld as shown in the experimental schedule (Figure 8A). The mice were evaluated for body weight change (Figure 8B), disease activity index (Figure 8C), clinical examination (Figure 8D), colon (Figure 8E) and histology (PAS staining) (in the inset, ZO - 1 and BrdU staining) (Figure 8F), calprotectin levels in intestinal homogenates (Figure 8G), and cytokine and defensin gene expression in the intestine (Figure 8H). Photographs were taken at high - resolution microscopy, 20 - fold magnification (scale bar, 200 μm). Data are presented as mean ± SEM (pooled from two experiments). H2O, untreated mice. *P < 0.05, **P < 0.01. [Figure 8-3]Figures 8A–8H are graphs and photographs showing how 3-IAld protects IL-10- / - mice from pathology after long-term treatment with anti-CTLA-4 monoclonal antibody. Control C56BL / 6 and IL-10 deficient mice were administered 100 μg of anti-CTLA-4 mAb and 3-IAld as shown in the experimental schedule (Figure 8A). Mice were evaluated for body weight change (Figure 8B), disease activity index (Figure 8C), clinical laboratory tests (Figure 8D), colon (Figure 8E) and histology (PAS staining) (ZO-1 and BrdU staining in the inset) (Figure 8F), calprotein levels in intestinal homogenate (Figure 8G), and cytokine and protective gene expression in the gut (Figure 8H). Photographs were taken using a high-resolution microscope at 20x magnification (scale bar, 200 μm). Data are expressed as mean ± SEM (combined from two experiments). H2O, untreated mice. *P<0.05, **P<0.01. [Figure 9] Figures 9A-9B show graphs and photographs illustrating how 3-IAld reduces lymphocyte infiltration in IL-10-deficient mice after long-term treatment with anti-CTLA-4 monoclonal antibody. Mice were treated as shown in the legend in Figure 8A, and histological changes and lymphocyte infiltration were evaluated (quantified using ImageJ software) by anti-CD3 staining in different organs. DAPI (4',6-diamidino-2-phenylindole) was used for DNA staining. [Figure 10] Figures 10A–10D are graphs and photographs showing how 3-IAld slows disease progression in 16-week-old IL-10-deficient mice. Mice were treated with 3-IAld and 1% DSS, as shown in (Figure 10A), and evaluated for body weight % (Figure 10B), histological score (Figure 10C), and inflammatory bowel tissue lesions and epithelial and regenerative conditions (by PAS and Ki-67 staining, respectively) (Figure 10D). [Figure 11-1]Figures 11A–11F are graphs and photographs illustrating how the 3-IAld modified microbiome provides protection in DSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were subjected to DSS-colitis with or without anti-CTLA-4 treatment (Figure 11E) or without (Figures 11A–11D), and fresh fecal pellets (FMT) from control or 3-IAld treated mice were transplanted two days before and two days after colitis induction. Mice were sacrificed 7 days (Figures 11A-11D) or 14 days (Figure 11E) after colitis induction, and weight change % (Figures 11A, 11E), macroscopic pathology (Figure 11B), histological score (Figure 11C), colon histopathology (periodic acid-Schiff staining) (Figure 11D), and methylation / demethylation status of the Foxp3 promoter in mesenteric lymph nodes (Figure 11F) were evaluated. Photographs were taken using a high-resolution microscope at 10x and 20x magnification (scale bars, 500 μm and 200 μm). Data are representative of three independent experiments. Each in vivo experiment included 3 mice per group (6-12 mice in each experiment). Data are expressed as mean ± SD. H2O, untreated mice. *P<0.05, **P<0.01, ***P<0.001. [Figure 11-2]Figures 11A–11F are graphs and photographs illustrating how the 3-IAld modified microbiome provides protection in DSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were subjected to DSS-colitis with or without anti-CTLA-4 treatment (Figure 11E) or without (Figures 11A–11D), and fresh fecal pellets (FMT) from control or 3-IAld treated mice were transplanted two days before and two days after colitis induction. Mice were sacrificed 7 days (Figures 11A-11D) or 14 days (Figure 11E) after colitis induction, and weight change % (Figures 11A, 11E), macroscopic pathology (Figure 11B), histological score (Figure 11C), colon histopathology (periodic acid-Schiff staining) (Figure 11D), and methylation / demethylation status of the Foxp3 promoter in mesenteric lymph nodes (Figure 11F) were evaluated. Photographs were taken using a high-resolution microscope at 10x and 20x magnification (scale bars, 500 μm and 200 μm). Data are representative of three independent experiments. Each in vivo experiment included 3 mice per group (6-12 mice in each experiment). Data are expressed as mean ± SD. H2O, untreated mice. *P<0.05, **P<0.01, ***P<0.001. [Figure 12-1]Figures 12A–12J are graphs and photographs illustrating how 3-IAld does not interfere with the development of antitumor immunity. In Figures 12A–12E, C57BL / 6 mice were subcutaneously injected with B16 tumor cells and intraperitoneally administered 100 μg of anti-CTLA-4 mAb or isotype control at 3-day intervals for 4 doses (up to 16 days). 3-IAld was administered intragastricly every other day. Mice were evaluated for tumor growth (Figure 12A), histology (periodate-Schiff staining) (Figure 12B), immunohistochemistry of CD8+ and CD4+ tumor-infiltrating lymphocytes (TILs) (Figure 12C), number of positive TILs per high-power field (HPF) (Figure 12D), and expression of Cxcl9 and perforin (Figure 12E). (Figures 12F-12J) LLC cells were orthotopically injected into C57BL / 6 mice, and 200 μg of anti-PD-1 mAb or isotype control was administered intraperitoneally five times at 3-day intervals (maximum 18 days). 3-IAld was administered intragastricly every other day. Mice were evaluated for (Figure 12F) survival%, (Figure 12G) lung weight, (Figure 12H) macroscopic lung pathology, and (Figures 12I, 12J) CD4+CD25+Foxp3+ cells. Photographs were taken using a high-resolution microscope at 40x magnification (scale bar, 100 μm). Each in vivo experiment included 3 mice per group (9 mice in each experiment). Data are expressed as mean ± SEM. One-way ANOVA, Bonferroni post-hoc test. *P<0.05, **P<0.01, ***P<0.001. [Figure 12-2]Figures 12A–12J are graphs and photographs illustrating how 3-IAld does not interfere with the development of antitumor immunity. In Figures 12A–12E, C57BL / 6 mice were subcutaneously injected with B16 tumor cells and intraperitoneally administered 100 μg of anti-CTLA-4 mAb or isotype control at 3-day intervals for 4 doses (up to 16 days). 3-IAld was administered intragastricly every other day. Mice were evaluated for tumor growth (Figure 12A), histology (periodate-Schiff staining) (Figure 12B), immunohistochemistry of CD8+ and CD4+ tumor-infiltrating lymphocytes (TILs) (Figure 12C), number of positive TILs per high-power field (HPF) (Figure 12D), and expression of Cxcl9 and perforin (Figure 12E). (Figures 12F-12J) LLC cells were orthotopically injected into C57BL / 6 mice, and 200 μg of anti-PD-1 mAb or isotype control was administered intraperitoneally five times at 3-day intervals (maximum 18 days). 3-IAld was administered intragastricly every other day. Mice were evaluated for (Figure 12F) survival%, (Figure 12G) lung weight, (Figure 12H) macroscopic lung pathology, and (Figures 12I, 12J) CD4+CD25+Foxp3+ cells. Photographs were taken using a high-resolution microscope at 40x magnification (scale bar, 100 μm). Each in vivo experiment included 3 mice per group (9 mice in each experiment). Data are expressed as mean ± SEM. One-way ANOVA, Bonferroni post-hoc test. *P<0.05, **P<0.01, ***P<0.001. [Figure 13] Figures 13A to 13E graphically represent pharmacodynamic studies, including the analysis of unlabeled 3-IAld (Figure 13A), labeled 3-IAld (Figure 13B), potential metabolites of unlabeled 3-IAld (Figure 13C), and analysis of methylation forms of indole-3-formic acid, namely 1-methylindole-2-carboxylic acid and methyl indole-3-carboxylate levels (Figure 13D), where small fluctuations were detected relative to the basic endogenous level of 3-IAld at different time points. The presence of indole-3-formic acid was also confirmed in labeled 3-IAld (Figure 13E). [Figure 14]The graph shows the multiplicative increase in Cyp1a1 mRNA upon addition of 3-IAld or Ox-3-IAld. When human cell lines, including the liver cancer cell line HepG2, were exposed to different concentrations of the ligand indole-3-formic acid or Ox-3-IAld, dose-dependent induction of the AhR activation marker Cyp1A1 was observed between 1 and 100 μM (Figure 14). [Figure 15] The figure shows the histopathology of different organs examined in a blind manner after hematoxylin and eosin (H&E) staining. [Figure 16] This represents the chemical structure of 1-methylindole-3-carboxylic acid, also known as 1-methyl-3-indolecarboxylic acid, 1-methyl-1H-indole-3-carboxylic acid, 1-methyl-1H-indole-3-carboxylic acid, or 1ME3CA (MF:C10H9NO2, IUPAC name: 1-methyl-1H-indole-3-carboxylic acid). [Figure 17-1] Figures 17A to 17F are graphs and photographs showing that 1-methylindole-3-carboxylic acid protects against inflammatory lesions in DSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and 1-methylindole-3-carboxylic acid as shown in the experimental schedule, where 1-methylindole-3-carboxylic acid was administered every other day at a dose of either 0.09, 0.18, or 0.36 mg / mouse (Figure 17A). Mice were evaluated for body weight change (Figure 17B), disease activity index (Figure 17C), clinical morbidity and rectal bleeding (Figure 17D), histology of the colon and ileum by PAS staining (Figures 17E-17F), epithelial barrier function in the colon and ileum (ZO-1 staining), and lymphocyte infiltration (CD3 staining). Photographs were taken using a high-resolution microscope at 20x magnification (scale bar, 200 μm). Each in vivo experiment included 4-6 mice per group (16-24 mice per experiment). Data are expressed as mean ± SEM. H2O, untreated mice. One-way ANOVA, Bonferroni post-hoc test. ***P<0.001, ****P<0.0001. [Figure 17-2] Figures 17A to 17F are graphs and photographs showing that 1-methylindole-3-carboxylic acid protects against inflammatory lesions in DSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and 1-methylindole-3-carboxylic acid as shown in the experimental schedule, where 1-methylindole-3-carboxylic acid was administered every other day at a dose of either 0.09, 0.18, or 0.36 mg / mouse (Figure 17A). Mice were evaluated for body weight change (Figure 17B), disease activity index (Figure 17C), clinical morbidity and rectal bleeding (Figure 17D), histology of the colon and ileum by PAS staining (Figures 17E-17F), epithelial barrier function in the colon and ileum (ZO-1 staining), and lymphocyte infiltration (CD3 staining). Photographs were taken using a high-resolution microscope at 20x magnification (scale bar, 200 μm). Each in vivo experiment included 4-6 mice per group (16-24 mice per experiment). Data are expressed as mean ± SEM. H2O, untreated mice. One-way ANOVA, Bonferroni post-hoc test. ***P<0.001, ****P<0.0001. [Figure 18] This photograph shows that 1-methylindole-3-carboxylic acid reduces lymphocyte infiltration in mice treated with anti-CTLA-4. C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and 1-methylindole-3-carboxylic acid as shown in the experimental schedule in Figure 17A. The mice were evaluated for histological changes and lymphocyte infiltration by anti-CD3 staining in the lungs. DAPI (4',6-diamidino-2-phenylindole) was used for DNA staining. [Figure 19]The graph shows the increase in mRNA levels for Il1b, Il10, Cyp1a1, Reg3g, and Il22, demonstrating that the addition of 1-methylindole-3-carboxylic acid promotes anti-inflammatory status and AhR activation in mouse colitis. C57BL / 6 mice were subjected to DSS + anti-CTLA-4 induced colitis, and 1-methylindole-3-carboxylic acid was administered as described in the experimental schedule in Figure 17A. Cytokine and AhR-dependent gene expression were evaluated. Data are expressed as mean ± SEM. H2O, untreated mice. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001. One-way ANOVA, Bonferroni post-hoc test. [Figure 20A] This shows the pharmacokinetics of 3-IAld in the intestines, serum, lungs, liver, brain, and kidneys over a 24-hour period. [Figure 20B] This paper describes the pharmacokinetics of 1-methylindole-3-carboxylic acid in the intestines, serum, lungs, liver, brain, and kidneys over a 24-hour period. [Figure 21] This study demonstrates the in vitro AhR agonist activity of 3-IAld and 1ME3CA in the reporter H1L6.1c3 cell line. [Figure 22-1] This figure shows the in vitro AhR activity of 3-IAld and 1ME3CA in A549 alveolar epithelial cells (Figure 22A), Calu-3 bronchial epithelial cells (Figure 22B), Caco-2 colon cancer cells (Figure 22C), and HepG2 hepatocellular carcinoma cells (Figure 22D). ITE and FICZ are reference AhR agonists. The assay was performed 4 hours after incubation. [Figure 22-2] This figure shows the in vitro AhR activity of 3-IAld and 1ME3CA in A549 alveolar epithelial cells (Figure 22A), Calu-3 bronchial epithelial cells (Figure 22B), Caco-2 colon cancer cells (Figure 22C), and HepG2 hepatocellular carcinoma cells (Figure 22D). ITE and FICZ are reference AhR agonists. The assay was performed 4 hours after incubation. [Figure 23-1]This study demonstrates that MECA provides dose-dependent protection from inflammatory lesions in DSSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and MECA as shown in the experimental schedule (Figure 23A). Mice were evaluated at ×40 magnification (scale bar, 100 μm) for body weight change (Figure 23B), disease activity index (Figure 23C), clinical morbidity and rectal bleeding (Figure 23D), and colon and ileum histology (PAS staining) (Figure 23E). Each in vivo experiment included 4-6 mice per group (20 mice in each experiment). [Figure 23-2] This study demonstrates that MECA provides dose-dependent protection from inflammatory lesions in DSSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and MECA as shown in the experimental schedule (Figure 23A). Mice were evaluated at ×40 magnification (scale bar, 100 μm) for body weight change (Figure 23B), disease activity index (Figure 23C), clinical morbidity and rectal bleeding (Figure 23D), and colon and ileum histology (PAS staining) (Figure 23E). Each in vivo experiment included 4-6 mice per group (20 mice in each experiment). [Figure 23-3] This study demonstrates that MECA provides dose-dependent protection from inflammatory lesions in DSSS + anti-CTLA-4 induced colitis. C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and MECA as shown in the experimental schedule (Figure 23A). Mice were evaluated at ×40 magnification (scale bar, 100 μm) for body weight change (Figure 23B), disease activity index (Figure 23C), clinical morbidity and rectal bleeding (Figure 23D), and colon and ileum histology (PAS staining) (Figure 23E). Each in vivo experiment included 4-6 mice per group (20 mice in each experiment). [Figure 24]MECA reduces lymphocyte infiltration in anti-CTLA-4 treated mice in a dose-dependent manner. Lung, spleen, liver, and kidney tissue samples showed a dose-dependent ability to reduce lymphocyte infiltration in anti-CTLA-4 treated mice administered with MECA. [Figure 25A] Figures 25A and 25B show the toxic effects of 3-IAld and 1ME3CA on PAS-stained tissue sections of naive C57BL / 6 mice. Photographs were taken using a high-resolution Olympus DP71 microscope with a 10x objective lens. Scale bar 400 μm. Naive, untreated mice. [Figure 25B] Figures 25A and 25B show the toxic effects of 3-IAld and 1ME3CA on PAS-stained tissue sections of naive C57BL / 6 mice. Photographs were taken using a high-resolution Olympus DP71 microscope with a 10x objective lens. Scale bar 400 μm. Naive, untreated mice. [Modes for carrying out the invention]

[0015] Some aspects of this disclosure relate to a method for treating CTLA-4 checkpoint-associated immunodeficiency in a patient, comprising administering a therapeutically effective amount of a pharmaceutical composition comprising 1H-indole-3-carboxyaldehyde (3-IAld) to a patient in need thereof.

[0016] Additional aspects of the present disclosure relate to a method for treating CTLA-4 checkpoint-associated immunodeficiency in a patient, comprising administering a therapeutically effective amount of a pharmaceutical composition comprising 1-methylindole-3-carboxylic acid to a patient in need thereof.

[0017] Before this disclosure is described in more detail, it should be understood that this disclosure is not limited to any particular composition or process step described and is therefore naturally subject to change. As will be apparent to those skilled in the art by reading this disclosure, each of the individual embodiments described and illustrated herein has other components and features that can be readily separated from or combined with the features of any of several other embodiments without departing from the scope or spirit of this disclosure. Any of the enumerated methods may be carried out in the order of the enumerated events or in any other logically possible order.

[0018] The headings provided herein are not intended to limit the various aspects of this disclosure, which can be defined by referring to this entire specification. Furthermore, the terms used herein are intended solely to describe specific aspects and not to limit them.

[0019] I. Terminology To make this disclosure easier to understand, certain terms are defined first. Where used in this application, each of the following terms shall have the meaning set forth below, unless otherwise expressly provided herein. Additional definitions are provided throughout this application.

[0020] As described herein, any concentration range, percentage range, ratio range, or integer range, unless otherwise indicated, should be understood to include any integer values ​​within the range described and, where appropriate, fractions thereof (e.g., one-tenth and one-hundredth of an integer).

[0021] Throughout this disclosure, the terms and entities “a” or “an” refer to one or more of those entities; for example, “chimeric polypeptide” is understood to represent one or more chimeric polypeptides. Accordingly, the terms “a” (or “an”), “one or more”, and “at least one” are interchangeable herein.

[0022] Furthermore, as used herein, “and / or” shall be considered a specific disclosure of each of two designated features or components, whether one is accompanied by the other or not. Accordingly, the term “and / or” as used herein in expressions such as “A and / or B” is intended to include “A and B,” “A or B,” “A” (alone), and “B” (alone). Similarly, the term “and / or” as used in expressions such as “A, B, and / or C” is intended to include each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone). In addition, “or” is used to mean a public list of components in a list. For example, “X includes A or B” means that X includes A, X includes B, X includes A and B, or X includes A or B and any other component.

[0023] The terms “approximately” or “essentially include” refer to a value or composition that falls within the acceptable margin of error for a particular value or composition as determined by those skilled in the art, and this depends in part on how that value or composition is measured or determined, i.e., on the limits of the measuring system. For example, “approximately” or “essentially from” may mean a standard deviation of 1 or less, or greater than 1, according to the convention of the art. Alternatively, “approximately” or “essentially from” may mean a range of up to 10%. Furthermore, particularly with respect to biological systems or processes, the term may mean up to one order of magnitude or up to five times the value. Where a particular value or composition is provided in the application and claims, unless otherwise stated, the meaning of “approximately” or “essentially from” should be considered to be within the acceptable margin of error for that particular value or composition.

[0024] As used herein, the term “approximately” means, when applied to one or more values ​​of interest, a value that is similar to the given reference value. In certain embodiments, unless otherwise stated or otherwise evident from the context (unless such a number would exceed 100% of the possible values), the term “approximately” means a range of values ​​that fall in any direction (above or below) 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% or less of the given reference value.

[0025] As used herein, a patient who is "homozygous" for a particular gene mutation has the same mutation in each allele.

[0026] As used herein, a patient who is "heterozygous" for a particular gene mutation has that mutation on one allele and different mutations on other alleles.

[0027] The term "antibody" may include, for example, monoclonal antibodies (mAbs), polyclonal antibodies, multispecific (e.g., bispecific) antibodies, recombinant antibodies, human antibodies, chimeric antibodies, and humanized antibodies. Furthermore, the term "antibody" may also encompass recombinantly expressed antigen-binding proteins and antigen-binding synthetic peptides.

[0028] An "antigen" refers to any molecule, such as a peptide, that can trigger an immune response or to which a TCR can bind. An immune response may include antibody production, activation of specific immune-qualified cells, or a combination thereof. Those skilled in the art will readily understand that virtually any macromolecule, including proteins or peptides, can function as an antigen. Antigens may be endogenously expressed, i.e., expressed by genomic DNA or recombinantly. Antigens and / or epitopes may be specific to certain tissues, such as cancer cells, or they may be expressed broadly. In addition, larger molecular fragments may also function as antigens. In one embodiment, an antigen is a tumor antigen.

[0029] As used herein, “antitumor effect” refers to a biological effect that may manifest as a reduction in tumor volume, a decrease in the number of tumor cells, a decrease in tumor cell proliferation, a decrease in the number of metastases, an extension of the patient’s overall survival or progression-free survival, an extension of the patient’s expected lifespan, or an improvement in various physiological symptoms of the patient associated with the tumor. The antitumor effect may also refer to the prevention of tumor development, such as by a vaccine.

[0030] Indole-3-aldehyde, or 3-IAld, is a metabolite derived from the microbial degradation of the amino acid tryptophan. The identification of 3-IAld is further described in U.S. Patent Application Publication 2016 / 0206595 (incorporated herein by reference). 3-IAld is an agonist of the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor involved in a wide range of physiological activities, including the maintenance of mucosal homeostasis in barrier organs (Stockinger B, et al., Ann. Rev. Immunol. 2014;32:403-32). By activating AhR in type 3 innate lymphoid cells, 3-IAld induces the production of IL-22, a key cytokine in host defense of mucosal surface and tissue repair, as well as in regulating microbial composition (Zelante T, et al., Immunity. 2013; 39(2): 372-85; Borghi M, et al., Immunol. 2019; 10: 2364). Therefore, the 3-IAld-AhR-IL-22 axis represents a functional signaling unit for enhancing barrier function, which has promising activity in pathological conditions characterized by epithelial damage, mucosal changes, and hyperinflammatory responses. The formula for indole-3-aldehyde (or indole-3-carbaldehyde, 3-IAld, MF: C9H7NO, IUPAC: IH-indole-3-carbaldehyde) is shown below and in Figure 1. Since 3-IAld has no potential side effects, good tolerability is expected in humans. [ka]

[0031] Endogenous tryptophan (Trp) metabolites play a crucial role in mammalian gut immune homeostasis. In the gastrointestinal tract, dietary AhR ligands promote local IL-22 production by innate lymphoid cells (ILCs) (Qiu et al, 2012) (referred to here as ILC3) (Spits et al, 2013) (Lee et al, 2011). Metabolomics analysis has revealed that gut bacteria influence host metabolism and immunity through various chemically distinct metabolites, including amino acid metabolites (Wikoff et al, 2009). In particular, the fact that dietary Trp deficiency impairs gut immunity and alters the gut microbial community in mice (Hashmoto et al, 2012) suggests that mucosal homeostasis is a multifactorial phenomenon in which Trp metabolism is a key regulatory component. However, the sources and properties of any such AhR ligand, the effects of microbial imbalances on mucosal reactivity driven by AhR and IL-22, and whether activation of AhR by microbiome-derived metabolites also occurs are all unknown. The enzyme Trp2,3-dioxygenase (Opitz et al, 2011), mainly expressed in the liver, regulates Trp concentration after nutrient Trp uptake under normal conditions. However, the high expression level of IDO1 in mucosal sites during immune activation (Dai et al., 2010) suggests that IDO1 is the dominant enzyme that regulates mucosal immune reactivity through local amino acid nutrient levels, gut microbiota size and metabolic activity, and the host's own immunomodulatory activity via L-kynurenine production. Therefore, these data qualify IDO1 as an important molecule in directing the host-microbiota symbiotic relationship and their integration into the adaptive immunity of vertebrate hosts.

[0032] The microbiome-AhR axis may represent a crucial strategy pursued through co-evolutionary symbiosis to fine-tune the incidental host mucosal reactivity to Trp catabolism. For this purpose, the drug 3-IAld may offer significant advantages over any other current treatment by improving host immune reactivity, epithelial barrier function, and pathogen colonization. Indole acts as an AhR ligand, activating transcription factors that control the degradation of endogenous and exogenous toxins, thereby preventing inflammatory damage and maintaining barrier integrity (Hubbard et al., 2015b). Furthermore, indole exerts diverse functions, including bidirectional communication with the microbiome to fine-tune the host immune response, immune tolerance, and metabolism (Stockinger et al., 2014). Indole administration to germ-free mice has been shown to increase the expression of epithelial tight junction proteins and attenuate indicators of inflammatory bowel disease (Shimada et al, 2013). Similarly, the therapeutic effect of indole-3-aldehyde (3-IAld) has been reported in a mouse model of dextran-induced colitis (Zelante et al, 2013).

[0033] As used herein, the terms “medicinal active ingredient” or “therapeutic agent (factor)” (“API”) refer to a biologically active compound.

[0034] The terms "patient" and "subject" are used interchangeably and refer to animals such as humans.

[0035] Those skilled in the art will recognize that when an amount of "a compound or a pharmaceutically acceptable salt thereof" is disclosed, the amount of the pharmaceutically acceptable salt form of that compound corresponds to an amount equivalent to the concentration of the free base of that compound. Note that the amounts of compounds or pharmaceutically acceptable salts thereof disclosed herein are based on their free base forms.

[0036] As used herein, the term “in combination with” when referring to two or more compounds, factors, or additional active pharmaceutical ingredients means administering those two or more compounds, factors, or additional active pharmaceutical ingredients to a patient prior to, simultaneously, or sequentially to each other.

[0037] CTLA-4 checkpoint-associated immunodeficiency, or primary CTLA-4 checkpoint-associated immunodeficiency, or immunodeficiency, is a disorder or disease in patients characterized by various combinations of enteropathy, hypogammaglobulinemia, recurrent respiratory infections, granulomatous lymphocytic interstitial lung disease, lymphocyte infiltration of non-lymphoid organs (intestine, lungs, brain, bone marrow, kidneys), autoimmune thrombocytopenia or neutropenia, autoimmune hemolytic anemia, and lymphadenomatosis. CTLA-4 checkpoint-associated immunodeficiency is diagnosed based on clinical symptoms, laboratory findings, and genetic testing. Patients with only one functional copy of the gene encoding CTLA-4 suffer from severe autoimmunity. These heterozygous mutations result in a novel phenotype, along with more typical signs of autoimmunity, including hyperactivated T and B cell infiltration into non-lymphoid organs such as the intestine, lungs, and brain. Some clinical symptoms in CTLA-4 haploinsufficiency are reminiscent of those observed on biopsies of inflamed organs. Patients with primary CTLA-4 checkpoint-associated immunodeficiency may also have recurrent respiratory infections, hypogammaglobulinemia, autoimmune cytopenia, autoimmune enteropathy, and granulomatous infiltrative lung disease.

[0038] Primary CTLA-4 checkpoint-associated immunodeficiency is diagnosed based on clinical symptoms, laboratory findings, and genetic testing. Patients with only one functional copy of the gene encoding CTLA-4 suffer from severe autoimmunity. These heterozygous mutations result in a novel phenotype, along with more typical signs of autoimmunity, involving hyperactivated T and B cell infiltration into non-lymphoid organs such as the intestines, lungs, and brain. Some clinical symptoms in CTLA-4 haploinsufficiency are reminiscent of those observed in biopsies of inflamed organs in patients receiving anti-CTLA-4 therapy. Patients with primary CTLA-4 checkpoint-associated immunodeficiency may also have recurrent respiratory infections, hypogammaglobulinemia, autoimmune cytopenia, autoimmune enteropathy, and granulomatous infiltrative lung disease.

[0039] CTLA-4 is an essential negative regulator of the T cell-mediated immune response. The essential role of CTLA-4 in lymphocyte homeostasis and tolerance is clearly demonstrated by ctla4 knockout mice, which rapidly develop lethal and destructive multi-organ lymphocyte infiltration. Autoimmunity originates from autoreactive T lymphocytes and / or B lymphocytes and underlies a wide range of conditions, from endocrine disorders to cytopenia. Genetic epidemiological studies have long suggested that many autoimmune conditions have a heritable component. Haploinsufficiency is defined by the occurrence of a certain phenotype in heterozygotes despite the absence of negative dominance of the mutated allele compared to the wild-type counterpart. Heterozygous CTLA-4 germline mutations impair the suppressive function of T cells, leading to immunodysregulation syndromes characterized by an activated immune system with autoimmune features and organ lesions caused by infiltrating effector T cells. Therefore, having a single working copy of CTLA-4 is not sufficient to produce enough CTLA-4 protein for a normal immune system. These heterozygous mutations result in a novel phenotype, accompanied by hyperactivated T and B cell infiltration into the gut, lungs, and brain, along with more typical signs of autoimmunity.

[0040] Located adjacent to the long arm of chromosome 2 are gene paralogs encoding the immunoglobulin family co-activating receptors CD28 and CTLA-4. While CD28 and CTLA-4 share the same B7 ligand, each provides opposing proliferation signals to T cells. CD28 delivers a positive signal to T cells, leading to T cell activation and effector cell differentiation, while ligation from CTLA-4 to CD80 and CD86 delivers a negative signal to T cells, limiting T cell IL-2 production, proliferation, and survival. CTLA-4 is inductively expressed on CD4+Foxp3 conventional T (Tconv) cells after activation and constitutively expressed on CD4+Foxp3+ regulatory T (Treg) cells. An important inhibitory function of CTLA-4 is CTLA-4-null (Ctla4 - / - This has been revealed by a rapidly developing, fatal inflammatory phenotype in mice, which spontaneously develop marked T-cell proliferation accompanied by multi-organ lymphocyte infiltration and tissue destruction (Tivol et al, 1995). The similarity between this phenotype and systemic autoimmune disease has prompted research into the role of CTLA-4 in T-cell tolerance and autoimmunity. CTLA-4 can act on Tconv cells and Treg cells and is a mediator of Treg cell suppression.

[0041] Human CTLA4 haploinsufficiency resulted in dysregulation of FoxP3+ regulatory T (Treg) cells, hyperactivation of effector T cells, and lymphocyte infiltration in target organs such as the gut, lungs, and brain, in addition to more classic signs of autoimmunity. Adaptive immune responses must balance the need to avoid damage to self-antigens and host tissues with the response to foreign antigens. At one end of the spectrum, inefficient activation of the immune response can lead to infectious lesions, while hyperactivation can promote autoimmune responses. While it may be expected that different genetic mutations underlie these seemingly opposing outcomes, it is paradoxically well recognized that autoimmunity and immunodeficiency can occur simultaneously in the same individual.

[0042] Since CTLA-4 inhibits the CD28 pathway, which plays a role in assisting T cells in the B cell response, CTLA-4 deficiency may be expected to enhance CD28 function and promote humoral immunity. One possible interpretation is that T cell hyperactivation may lead to infiltration and destruction of the bone marrow niche, impairing B cell development. This is consistent with the disruption of B cell lymphocyte synthesis in Treg-deficient mice. Alternatively, increased differentiation of CD28-dependent follicular helper T (TFH) cells may lead to chronic stimulation and exhaustion of B cells.

[0043] CTLA-4 deficiency is characterized by the infiltration of immune cells into the intestines, lungs, bone marrow, central nervous system, kidneys, and sometimes other organs. Most people with CTLA4 deficiency experience diarrhea or intestinal disorders. Enlargement of the lymph nodes, liver, and spleen is also common, as are respiratory infections. People with CTLA-4 deficiency often experience autoimmune problems that can affect various organs and tissues, including the blood, thyroid, skin, and joints. The disease may also slightly increase the risk of lymphoma, a type of immune cell carcinoma.

[0044] When the increased risk of cancer is combined with virus-related observations, the inventors hypothesize that the incomplete immune surveillance of chronically virus-infected cells and the reduced elimination of oncogenic viruses lead to uncontrolled cell proliferation. Reduced CTLA-4 expression results in uncontrolled T cell proliferation, as is known in HIV-infected individuals, potentially leading to abnormal proliferation of autoreactive clones beyond EBV-specific T cell clones, for example.

[0045] Notably, autosomal dominant CTLA-4 deficiency exhibits incomplete penetrance, resulting in some heterozygotes being asymptomatic. A delicate balance exists between self-tolerance and autoimmunity, which is known to be at least partially influenced by quantitative variations in CTLA-4 expression. As predicted by preclinical models, complex interactions between genetics and environment can determine the development of distinct phenotypes associated with CTLA-4 deficiency, or conversely, the maintenance of asymptomatic status (Kuehn et al, 2014).

[0046] CTLA-4 checkpoint-associated immunodeficiency syndromes result in a wide range of immunomodulatory disorders with extensive clinical manifestations. In many cases, the onset of clinical symptoms is not observed until adulthood. Some patients suffer from severe blood cell depletion. Patients with immune thrombocytopenic purpura are at risk of bleeding complications, and patients with immunohemolytic anemia may be at risk of complications secondary to hypoxia, e.g., acute renal failure and acute vascular events. Patients with pulmonary lymphocyte infiltration may experience recurrent lung infection episodes, which may be exacerbated by the presence of hypogammaglobulinemia and by combination immunosuppressive therapies established to reduce lymphocyte infiltration and autoinflammation. Some patients also suffer from gastrointestinal disorders. Those with gastric atrophy are at risk of ulcers, which is also due to a tendency to develop recurrent infections with Helicobacter pylori, which may progress to lymphoproliferative disorders (e.g., MALT lymphoma). Severe complications that can alter the quality and quantity of life include colitis, pancreatitis, and central nervous system infiltration. (Schubert et al, 2014; Kuehn et al, 2014).

[0047] The report by Kuehn et al., which explains the consequences of decreased CTLA-4 expression, shows striking similarities to, and also differences from, reports of inflammatory diseases associated with anti-CTLA-4 cancer therapy. Among 540 melanoma patients who received intermittent CTLA-4 blockade with ipilimumab, approximately 60% experienced immune-related adverse events, with 11% having severe symptoms. The most common were cutaneous (rash and vitiligo), gastrointestinal (colitis), and endocrine (hypothyroidism and hypophysitis) (Hodi et al, 2010). Less common inflammatory events included hepatitis, uveitis, neuropathy, and pneumonia (Attia et al, 2005). Most immune-related toxicities were easily managed with immunosuppressants, but some were fatal. Biopsies of inflamed organs showed mixed CD4+ and CD8+ T-cell infiltration. The elevated serum titers of autoantibodies observed in some patients were directed towards thyroid tissue, acetylcholine receptors, the pituitary gland, and other targets.

[0048] A significant correlation between severe immuno-related toxicity and major tumor regression has been described (Attia et al, 2005), suggesting a general biological mechanism and highlighting the precarious balance between self-tolerance and autoimmunity in malignant and normal tissues. Oncogenic predisposition in CTLA-4 dysfunction manifests as a result of immune activation due to chronic inflammation, uncontrolled oncogenic viruses or tumor cells by immunological means, and endogenous T-cell dysfunction stemming from underlying genetic defects (Salavoura et al, 2008). For these reasons, patients with CTLA-4 haploinsufficiency are more susceptible to malignancies, particularly gastric and lymphoma (Dhalla et al, 2011 - Cunningham - Rundles et al, 1999). Lymphoma is known to arise as a result of lymphopenia, which is present in most patients. Gastric cancer often arises from chronically inflammatory tissue, a commonly known risk factor (Compare et al, 2010). A recent study by Egg et al. showed that affected CTLA4 mutation carriers had a 12.9% higher risk of malignant tumors. Furthermore, when comparing cancer risk between the general population and affected CTLA4 mutation carriers, affected CTLA4 mutation carriers had a higher annual cancer incidence rate (Egg et al, 2018).

[0049] Therefore, patients with CTLA-4 checkpoint-associated immunodeficiency or primary CTLA-4 checkpoint-associated immunodeficiency may present with several different gastrointestinal diagnoses, syndromes, or symptoms, including but not limited to CHAI, LRBA, LATAIE, regulatory T cell deficiency, intestinal disease, enteritis, immune-mediated colitis (IMC), gastrointestinal disorders, gastric atrophy, colitis, and others that may not be fully recognized at present.

[0050] As used herein, “CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI)” is a CTLA-4 checkpoint-associated immunodeficiency disorder characterized by a heterozygous CTLA4 mutation(s) in humans associated with severe immunomodulatory impairment. CHAI disease is primarily characterized by lymphoproliferation, autoimmune cytopenia, enteropathy, interstitial lung disease, and recurrent infections.

[0051] As used herein, “Lipopolysaccharide-responsive beige-like anchor (LRBA)” is a CTLA-4 checkpoint-associated immunodeficiency disorder characterized by a gene carrying one or more genetically biallelemic mutations in a patient. Since LRBA deficiency also leads to secondary loss of CTLA-4, this recessive disorder is referred to as LRBA deficiency with autoantibodies, T-reg cell deficiency, autoimmune infiltration, and enteropathy (LATAIE). LATAIE also commonly presents with enteropathy, autoimmunity, lymphoproliferation, and respiratory infections. Indeed, CHAI patients have a higher rate of granulomas, malignancies, atopic dermatitis, skin disorders, and neurological disorders, while LATAIE patients are more commonly associated with life-threatening infections, pneumonia, ear and nose disorders, and organ hypertrophy, autoimmune enteropathy, and growth retardation. While the characteristics of CHAI and LATAIE are similar, a significant difference is the earlier age of onset in LATAIE, with the disease often becoming apparent in children under school age, whereas CHAI typically develops in older children or young adults.

[0052] As used herein, “regulatory T (Treg) cell deficiency” refers to any CTLA-4 checkpoint-related immunodeficiency disorder that may result in reduced elimination of chronically virus-infected cells and oncogenic viruses leading to uncontrolled cell proliferation. One example is reduced CTLA-4 expression, which may lead to uncontrolled T cell proliferation, such as the abnormal proliferation of autoreactive clones beyond EBV-specific T cell clones.

[0053] As used herein, “enteropathy disease” refers to a CTLA-4 checkpoint-related immunodeficiency disorder characterized by ongoing damage or irritation and swelling of the small intestine.

[0054] As used herein, "gut inflammation" refers to inflammation of the gastrointestinal (GI) tract and is a CTLA-4 checkpoint-related immunodeficiency condition. Prolonged inflammation can lead to damage to the GI tract.

[0055] As used herein, “immune-mediated colitis (IMC)” refers to a common immune-related adverse event associated with immune checkpoint inhibitors, characterized by abdominal pain, mucus or blood in the stool, and fever. Immune checkpoint inhibitors are immunotherapeutic drugs that work by preventing checkpoint proteins from binding to their partner proteins. This prevents the “off” signal from being sent, allowing T cells to kill cancer cells. Dysregulation of T cells can also contribute to IMC, in which these T cells attack GI ductal cells and tissues.

[0056] As used herein, “autoimmune infiltration” or “infiltration” refers to the diffusion or accumulation of foreign substances (in tissues or cells) in excessive amounts beyond normal. Infiltration may include lymphocyte infiltration of non-lymphoid organs such as the intestines, as in CHAI. Weakening of the intestinal lining can also lead to infiltration by other infiltrators within the intestines.

[0057] As used herein, “gastric atrophy” is a condition characterized by thinning of the inner lining of the stomach and / or intestinal wall, as well as the loss of the glandular cells of the inner lining that release substances that aid digestion. It may be caused by infection with Helicobacter pylori or certain autoimmune conditions.

[0058] "Cancer" refers to a broad group of diseases characterized by the uncontrolled proliferation of abnormal cells in the body. The uncontrolled division and proliferation of cells leads to the formation of malignant tumors, which may invade adjacent tissues and metastasize to distant parts of the body via the lymphatic system or bloodstream. "Cancer" or "cancer tissue" may include tumors. Examples of cancers that can be treated by the methods of the present invention include, but are not limited to, lymphoma, leukemia, and cancers of the immune system, including other leukocyte malignancies. In some embodiments, the methods of the present invention are used for, for example, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, cutaneous or intraocular malignant melanoma, uterine cancer, ovarian cancer, rectal cancer, anal cancer, gastric cancer, testicular cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, breast cancer, prostate cancer, lung cancer (e.g., non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC)), Hodgkin's disease, non-Hodgkin lymphoma, esophageal cancer, small intestine cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal gland cancer, soft tissue sarcoma, urethral cancer, penile cancer, chronic cancer It may be used to reduce the tumor size of tumors resulting from acute leukemia, acute myeloid leukemia, chronic myeloid leukemia, acute lymphoblastic leukemia, chronic lymphocytic leukemia, pediatric solid tumors, lymphocytic lymphoma, bladder cancer, kidney or ureteral cancer, renal pelvis carcinoma, central nervous system (CNS) neoplasms, primary CNS lymphoma, tumor angiogenesis, spinal axial tumors, brainstem gliomas, pituitary adenomas, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, T-cell lymphoma, environmentally induced cancers including those induced by asbestos, or any combination thereof. Certain cancers may respond to chemotherapy or radiotherapy, or they may be refractory. A refractory cancer is one that cannot be corrected by surgical intervention, and which either does not respond to chemotherapy or radiotherapy from the beginning, or becomes unresponsive over time.

[0059] Whereever an aspect is described using the word “including,” it is understood that other similar aspects are also provided, described using the terms “consisting of” and / or “essentially consisting of.”

[0060] As used herein, "cytokine" refers to a non-antibody protein released by one cell in response to contact with a specific antigen, which then interacts with a second cell to mediate a response in that second cell. Cytokines may be endogenously expressed in cells, added to cells in a culture, administered to a subject, or any combination thereof. Cytokines are released from immune cells, including macrophages, B cells, T cells, and mast cells, and can transmit immune responses. Cytokines can induce various reactions in recipient cells. Examples of cytokines include homeostatic cytokines, chemokines, pro-inflammatory cytokines, effectors, and acute-phase proteins. For example, homeostatic cytokines, including interleukin (IL) 7 and IL-15, can promote the survival and proliferation of immune cells, while pro-inflammatory cytokines can promote inflammatory responses. Examples of homeostatic cytokines include, but are not limited to, IL-2, IL-4, IL-5, IL-7, IL-10, IL-12p40, IL-12p70, IL-15, IL-21, and interferon (IFN) gamma. Examples of pro-inflammatory cytokines include, but are not limited to, IL-a, IL-lb, IL-6, IL-13, IL-17a, tumor necrosis factor (TNF)-alpha, TNF-beta, fibroblast growth factor (FGF)2, granulocyte-macrophage colony-stimulating factor (GM-CSF), soluble intercellular adhesion molecule 1 (sICAM-1), soluble vascular adhesion molecule 1 (sVCAM-1), vascular endothelial growth factor (VEGF), VEGF-C, VEGF-D, and placental growth factor (PLGF). Examples of effectors include, but are not limited to, granzyme A, granzyme B, soluble Fas ligand (sFasL), and perforin. Examples of acute-phase proteins include, but are not limited to, C-reactive protein (CRP) and serum amyloid A (SAA).

[0061] Chemokines are a type of cytokine that mediates chemotactic or directional cell movement. Examples of chemokines, but not limited to, include IL-8, IL-16, eotaxin, eotaxin-3, macrophage-derived chemokines (MDC or CCL22), monocyte chemotactic protein 1 (MCP-1 or CCL2), MCP-4, macrophage inflammatory protein la (MIP-la, MIP-la), MIP-Ib (MIP-lb), gamma-inducible protein 10 (IP-10), and thymic and activating regulatory chemokines (TARC or CCL17).

[0062] Other examples of cytokines include, but are not limited to, chemokine (CC motif) ligand (CCL) 1, CCL5, monocyte-specific chemokine 3 (MCP3 or CCL7), monocyte chemoattractant protein 2 (MCP-2 or CCL8), CCL13, IL-1, IL-3, IL-9, IL-11, IL-12, IL-14, IL-17, IL-20, IL-21, granulocyte colony-stimulating factor (G-CSF), leukemia suppressor (LIF), and oncostatin M. Examples include (OSM), CD154, lymphotoxin (LT) beta, 4-IBB ligand (4-1BBL), proliferation-inducing ligand (APRIL), CD70, CD153, CD178, glucocorticoid-inducing TNFR-related ligand (GITRL), tumor necrosis factor superfamily member 14 (TNFSF14), OX40L, TNF and ApoL-related leukocyte expression ligand 1 (TALL-1), or TNF-related apoptosis-inducing ligand (TRAIL).

[0063] "Immune response," as understood in the field, generally refers to the biological response within vertebrates to foreign factors or abnormal cells, such as cancerous cells, which protect the body from these factors and the diseases they cause. The immune response is mediated by the action of soluble macromolecules (including antibodies, cytokines, and complement) produced by one or more cells of the immune system (e.g., T lymphocytes, B lymphocytes, natural killer (NK) cells, macrophages, eosinophils, mast cells, dendritic cells, or neutrophils) and soluble molecules (including antibodies, cytokines, and complement) produced by any of these cells or the liver, resulting in selective targeting, binding to, damage to, destruction of, and / or elimination of invading pathogens, pathogen-infected cells or tissues, cancer or other abnormal cells, or, in the case of autoimmune or pathological inflammation, normal human cells or tissues. Examples of immune responses include T cells, e.g., effector T cells, Th cells, CD4 + cells, CD8 + This includes activation or inhibition of T cells or Treg cells, or activation or inhibition of any other cells of the immune system, such as NK cells. In some embodiments, the immune response refers to NK cell-mediated killing of foreign cells, such as allogeneic T cell therapy.

[0064] "Immunotherapy" refers to the treatment of individuals who have a disease or are at risk of developing or relapsing a disease, by means of inducing, enhancing, suppressing, or otherwise modifying the immune system or immune response.

[0065] As used herein, the terms “inactivation” or “inactivation” refer, for example, to a gene or protein, to a measure that can induce a reduction in protein expression. In some embodiments, inactivation may be achieved by deletion or mutation of all or part of the coding region of a gene, or all or part of the non-coding region of a gene, resulting in a decrease in the expression of that gene or the protein encoded by that gene. In some embodiments, inactivation is achieved by deletion of the entire coding region of a gene. In some embodiments, inactivation is achieved by partial deletion of the coding region of a gene. In some embodiments, inactivation is achieved by deletion of one or more regulatory elements that promote gene expression. In some embodiments, inactivation is achieved by mutation of one or more regulatory elements that result in a decrease or loss of gene expression. In some embodiments, inactivation is achieved by mutation of one or more nucleic acids resulting in the expression of a non-functional protein. In some embodiments, inactivation is achieved by missense mutations resulting in the expression of a non-functional protein. In some embodiments, inactivation is achieved by interference with the transcription or translation of a gene, resulting in a decrease in protein expression. In some embodiments, the decrease in expression is relative to the expression of the target gene in the cell before modification (e.g., deletion or mutation). In some embodiments, gene expression is measured before modification, then the cell is modified, and then gene expression is measured after modification.

[0066] As used herein, the term “lymphocyte” includes natural killer (NK) cells, T cells, or B cells. NK cells are a type of cytotoxic lymphocyte and a major component of the intrinsic immune system. NK cells reject tumor and virus-infected cells by inducing apoptosis or programmed cell death in target cells. They are called “natural killers” because NK cells do not require activation to kill target cells. T cells play a major role in cell-mediated immunity. T cell receptors (TCRs) are expressed on the surface of T cells and distinguish them from other lymphocyte types. The thymus, a specialized organ of the immune system, is primarily responsible for the maturation of T cells. There are six types of T cells: helper T cells (e.g., CD4+ cells); cytotoxic T cells (TCs, also known as cytotoxic T lymphocytes, CTLs, T killer cells, cytolytic T cells, CD8+ T cells, or killer T cells); and memory T cells ((i) stem memory TSCM cells, such as naive cells, are CD45RO-, CCR7+, CD45RA+, CD62L+ (L-selectin), CD27+, CD28+, and IL-7Rα+, but they also express large amounts of CD95, IL-2Rβ, CXCR3, and LFA-1, and are memory cells). - They exhibit a great many functional attributes specific to cells; (ii) Central memory TCM cells express L-selectin and CCR7 and secrete IL-2 but not IFNγ or IL-4, and (iii) Effector memory TEM cells do not express L-selectin or CCR7 but produce effector cytokines such as IFNγ and IL-4); regulatory T cells (Treg, suppressor T cells, or CD4+CD25+ regulatory T cells); natural killer T cells (NKT); and gamma-delta T cells.

[0067] B cells play a major role in humoral immunity (involving antibodies). B cells produce antibodies and antigens, act as antigen-presenting cells (APCs), and transform into memory B cells after activation through antigen interaction. In mammals, immature B cells are formed in the bone marrow, from which their name is derived.

[0068] Regulatory T cells (Tregs) are a specialized subpopulation of T cells that act to suppress the immune response, thereby maintaining homeostasis and self-tolerance. Tregs have been shown to inhibit T cell proliferation and cytokine production, playing a crucial role in preventing autoimmunity. Different subsets of Treg cells exist with various functions. Tregs can usually be identified by flow cytometry. The most specific marker for these cells is FoxP3, which is localized within the cell. Selected surface markers such as high CD25 density and low CD127 density can serve as surrogate markers for detecting Tregs in routine clinical practice. Dysregulation of Treg cell frequency or function can lead to the development of autoimmune diseases.

[0069] As used herein, “pharmaceutically acceptable carrier” includes any and all aqueous solvents (e.g., water, alcoholic / aqueous solutions, saline solutions, parenteral vehicles, e.g., sodium chloride, Ringer’s solution, etc.), non-aqueous solvents (e.g., propylene glycol, polyethylene glycol, vegetable oils, and injectable organic esters, e.g., ethyl oleate, etc.), dispersion media, polymers, coatings, surfactants, antioxidants, preservatives (e.g., antibacterial or antifungal agents, antioxidants, chelating agents, and inert gases), isotonic agents, absorption retarders, salts, drugs, drug stabilizers, gels, binders, excipients, disintegrants, lubricants, sweeteners, flavoring agents, dyes, fluids, and nutritional supplements, such similar materials and combinations thereof, as known to those skilled in the art. The pH and precise concentrations of various components in a pharmaceutical composition are adjusted by well-known parameters.

[0070] pharmaceutically acceptable carriers may include polymers or polymer mixes. One exemplary polymer mixture is Eudrgit® (Evonik Industries AG, Essen, Germany). EUDRAGIT® polymers are easily handled and processed at any scale and are available in multiple options for supply as aqueous dispersions, granules, organic solutions, powders, or ready-to-use powders. EUDRAGIT® polymers are compatible with all relevant process technologies, including film coating, melting, wet or dry granulation, hot melt extrusion, microencapsulation, and spray drying. Furthermore, all of our polymers are manufactured to consistent high quality, and established, audited facilities help provide global supply stability. EUDRAGIT® polymers may be used individually or in combination to suit substantially any target release profile, including immediate, delayed, sustained, pulsatile, accelerated, and zero-order release. Standard options for each coating layer include a single EUDRAGIT® polymer, or a combination of EUDRAGIT® polymers or other polymers, as well as certain other oral excipients and active pharmaceutical ingredients. Furthermore, Evonik's proprietary AEM® (Advanced Excipient Manufacturing Process) technology may be used to combine the functional advantages of different EUDRAGIT® polymers to create novel combination polymers that further enhance functionality and create new possibilities in formulation development and drug design. Pharmaceutically acceptable carriers may be designed for immediate release, delayed release, or sustained release.

[0071] As used herein, the term “pharmaceutically acceptable” may, in particular, indicate that a “pharmaceutically acceptable” compound or “pharmaceutically acceptable” composition is suitable for administration to a subject to achieve the treatment and / or prevention of a disease, a disorder or a condition (in particular at least one of CTLA-4 checkpoint-associated immunodeficiency disorders).

[0072] The pharmaceutical compositions of the present invention may be in solid or liquid form, and may, in particular, be one or more powders, one or more tablets, one or more fluids, especially one or more solutions, or one or more aerosols. The pharmaceutical compositions of the present invention may contain one or more further biological activators, e.g., activators, e.g., 3-IAld, for use in the treatment and / or prevention of at least one immunodeficiency disorder associated with the CTLA-4 checkpoint. Administration of the pharmaceutical compositions of the present invention may be selected from the group consisting of, for example, intraperitoneal, intravenous, parenteral, intrarenal, subcutaneous, topical, intrabronchial, intrapulmonary, and intranasal administration, and, if topical treatment is desired, intralesional administration. Intestinal administration may be, for example, oral administration, or other means of delivering 3-IAld to the intestines and GI ducts. The compositions of the present invention may also be administered directly to a target site by biorhythmic delivery to a target site, such as a specific organ affected by a disease, disorder, or condition.

[0073] In particular, the aforementioned administration may be carried out by injection and / or infusion and / or delivery, for example, by intravenous or intraperitoneal injection or infusion. The pharmaceutical composition may exist in the form of an injectable dosage form or a dosage form for administration by infusion, in particular an injectable dosage form for intravenous or intraperitoneal injection, or an infusion dosage form for intravenous or intraperitoneal administration, or a solid dose delivered by enteric coating by oral administration such as tablet or capsule formation, or in the form of other solid or semi-solid formulations such as a dry chewable or aqueous-based gel or softgel, or a gum formulation.

[0074] The pharmaceutical compositions according to the present invention can be administered to a subject in an appropriate dose. The dosage regimen may be determined, for example, by the attending physician. As is well known in the art, the dose for a given patient may depend on many factors, such as the patient's size, body surface area, age, weight, whether the administration is for prophylactic or therapeutic purposes, the indication of the target, the specific compound being administered, the patient's overall condition, and other drugs being administered concurrently. According to one embodiment, at least one antibody of the present invention.

[0075] According to one embodiment, the pharmaceutical composition of the present invention may be a pharmaceutical composition comprising 3-IAld and optionally at least one pharmaceutically acceptable carrier.

[0076] Furthermore, doses of 3-IAld and optionally at least one pharmaceutically acceptable carrier of the present invention in doses below or above the exemplary range shown above may be administered, for example, to treat and / or prevent at least one of the CTLA-4 checkpoint-associated immunodeficiency disorders. The pharmaceutical compositions of the present invention may be formulated to be short-acting, fast-release, long-acting, or sustained-release.

[0077] Furthermore, the pharmaceutical composition of the present invention may contain additional biologically active factors depending on the intended use of the pharmaceutical composition.

[0078] As used herein, the terms “reduced expression” and “increased expression” refer to the expression of a particular gene or protein in cells compared to a control, for example, the expression of a particular gene in modified cells compared to the expression of that gene in wild-type (unmodified) cells. Relative expression may be based on mRNA and / or protein levels. Any means of measuring mRNA and / or protein levels, including but not limited to immunohistochemistry and PCR-based techniques, may be used to determine whether gene or protein expression is reduced or increased.

[0079] As used herein, the terms “subject” and “patient” are interchangeable and refer to either a human or a non-human such as a primate, mammal, or vertebrate. In certain embodiments, the subject is a human.

[0080] As used herein, the terms “therapeutic benefit” or “therapeutic effectiveness” refer to anything that promotes or enhances the health condition of the subject in relation to the medical treatment of that condition. These include, but are not limited to, a reduction in the frequency or severity of signs or symptoms of the disease.

[0081] The terms “effective dose,” “therapeutic dose,” or “therapeutably effective dose” refer to the dose or concentration of a drug that elicits a positive biological response. For the purposes of this disclosure, embodiments of an effective dose may result in the alleviation of symptoms of inflammation, infiltration, weight loss, irritation, diarrhea, bloody stools, mucus in the stool, and tissue and cell damage.

[0082] As used herein, the terms “to treat, manage” or “to manage, manage” a disease or condition mean performing a protocol that may involve administering one or more therapies to a patient in an attempt to alleviate the signs or symptoms of the disease. In some embodiments, the therapy may slow the rate of disease progression, restore or alleviate the disease state, and / or promote remission or improved prognosis. Alleviation may occur before the signs or symptoms of the disease or condition appear, as well as after their appearance. Thus, in some embodiments, “to treat, manage” or “to manage, manage” includes “prevention” or “prevention” of the disease or undesirable condition. However, “to treat, manage” or “to manage, manage” may specifically include protocols that do not require the complete relief of all signs and / or symptoms, do not require a cure, and have only a minor effect on the patient.

[0083] In various forms, those who require it may be treated for the disease or to alleviate symptoms associated with CTLA-4 checkpoint-associated immunodeficiency.

[0084] As used herein, the terms "ug" and "uM" are interchangeable with "μg" and "μM," respectively.

[0085] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as those generally understood by those skilled in the art in the field to which this disclosure relates. For example, the Concise Dictionary of Biomedicine and Molecular Biology, Juo, Pei-Show, 2nd ed., 2002, CRC Press; The Dictionary of Cell and Molecular Biology, 3rd ed., 1999, Academic Press; and the Oxford Dictionary of Biochemistry and Molecular Biology, Revised, 2000, Oxford University Press provide many general dictionaries of the terms used herein.

[0086] Units, prefixes, and symbols are shown in their approved forms within the International System of Units (SI). Numerical ranges encompass the numbers that define the range. As described herein, any concentration range, percentage range, ratio range, or integer range should be understood to include any integer values ​​within the listed ranges, and, where appropriate, fractions thereof (such as 1 / 10 and 1 / 100 of an integer), unless otherwise specified.

[0087] Abbreviations used herein are defined throughout this disclosure. Various aspects of this disclosure are described in further detail in the following subsections.

[0088] The various embodiments described herein are described in further detail in the following subsections.

[0089] II. Compositions of the Disclosure Some aspects of this disclosure relate to 3-IAlds. Other aspects of this disclosure relate to pharmaceutical compositions comprising 3-IAlds and pharmaceutically acceptable carriers.

[0090] 1H-indole-3-carboxyaldehyde (3-IAld, MF:C9H7NO, IUPAC name: 1H-indole-3-carbaldehyde), also known as indole-3-aldehyde or 3-formylindole, belongs to the classification of organic compounds known as indole. Indole is a compound containing an indole moiety consisting of a pyrrole ring that condenses with benzene to form 2,3-benzopyrrole (Figure 1). 1H-indole-3-carboxyaldehyde exists as a solid, slightly soluble in water, and is a very weakly acidic compound (based on its pKa). 3-IAld is a metabolite of dietary L-tryptophan synthesized by human digestive bacteria, particularly species of the genus Lactobacillus (Zelante et al, 2013 - Lamas et al, 2016).

[0091] 3-IAld is an agonist of aryl hydrocarbon receptors (AhRs), ligand-activated transcription factors involved in a wide range of physiological activities, including maintaining mucosal homeostasis in barrier organs (Zelante et al, 2013; Zhang et al, 2016; Stockinger et al, 2014).

[0092] AhR is a ligand-dependent basic helix-loop-helix transcription factor that is evolutionarily highly conserved and expressed in most immune cell types and human tissues (Stockinger et al, 2014). Traditionally, AhR has been considered for its ability to metabolize harmful substances through the activation of cytochrome P450 drug-metabolizing enzymes, but its multiple functions, including developmental biology and interactions with the microbiome (crosstalk) to regulate host immunity, resistance, and metabolism, are increasingly being recognized. In particular, due to AhR's ability to bind to Th17 cells for antimicrobial activity, induce IL-22 production for epithelial cell repair and protection, and activate regulatory T cells to control inflammation, the intestinal and respiratory barriers are highly sensitive to AhR activity and activation (Esser et al, 2015). Therefore, AhR ligands such as 3-IAlD are promising compounds for drug discovery to treat inflammatory lesions on mucosal surfaces. Several studies have highlighted AhR's ability to respond to indole and indol metabolites, and therefore AhR is positioned as a candidate indole receptor (Hubbard et al, 2015a). Indole represents a wide range of enterobacteria-derived compounds produced from tryptophan, exerting important biological effects and potentially contributing to the pathogenesis of cardiovascular, metabolic, and psychiatric disorders (Konopelski et al, 2018). However, most research in this area is limited to experimental studies, which may be explained by AhR's context- and ligand-dependent activity (Safe et al, 2020). Therefore, activation of AhR by 3-IALD holds great therapeutic potential, provided that ligands such as 3-IALD, which have an optimal efficacy / safety profile, are accurately delivered to the target organ via appropriate biopharmaceutical formulations.As a result, when 3-IAld was formulated as enteric-coated microparticles for intestinal release, inflammatory histopathology and barrier function were improved, as evidenced by increased IL-22 production, increased expression of tight junction zonula occludens (ZO)1, proliferation of intestinal Lgr5+ (leucine-rich repeat-containing G protein-binding receptor 5) cells (a stem cell marker of the intestinal crypts) (Kumar et al, 2014), decreased intestinal leakage and Nfil3 expression (Yu et al, 2014), and transcription factors that induce the development of innate lymphoid cells 3, known to maintain epithelial barrier integrity (Puccetti et al., 2021).

[0093] Some aspects of this disclosure relate to 1-methylindole-3-carboxylic acid. Other aspects of this disclosure relate to pharmaceutical compositions comprising 1-methylindole-3-carboxylic acid and a pharmaceutically acceptable carrier.

[0094] 1-Methylindole-3-carboxylic acid, also known as 1-methyl-3-indolecarboxylic acid or 1-methyl-1H-indole-3-carboxylic acid (MECA) (1ME3CA) (MF:C10H9NO2, IUPAC name: 1-methyl-1H-indole-3-carboxylic acid), belongs to the classification of organic compounds known as methylindole. 1-Methylindole-3-carboxylic acid contains a methylindole moiety, which consists of a 1-methylpyrrole ring that condenses with benzene to form 1-methylindole (Figure 16).

[0095] III. Treatment Methods Several aspects of this disclosure relate to methods for treating a disease or condition of interest that requires such treatment, comprising administering the compositions disclosed herein to the subject. In some aspects, the method comprises administering 3-IAld alone or with a pharmaceutically acceptable carrier. In one aspect, the present invention relates to an improved method for preventing and / or treating inflammation and other symptoms associated with patients with CTLA-4 checkpoint-associated immunodeficiency using indole-3-aldehyde (3-IAld). In another aspect, the present invention relates to a method for preventing and / or treating inflammation, improving intestinal health and the health of the lining of the GI tubule, and reducing intestinal symptoms associated with the lining of the intestinal tubule and other GI, gastric cell infiltration, and patients with CTLA-4 checkpoint-associated immunodeficiency using an AhR ligand having biosimilar activity. In some aspects, the AhR ligand is indole-3-acetaldehyde (IAAld), indole-3-acetic acid (IAA, indoleacetic acid), or indole-3-lactic acid (ILA, indolelactic acid).

[0096] In some embodiments, the method involves administering 1-methylindole-3-carboxylic acid alone or with a pharmaceutically acceptable carrier. In one embodiment, the present invention relates to an improved method for preventing and / or treating inflammation and other symptoms associated with patients with CTLA-4 checkpoint-associated immunodeficiency using 1-methylindole-3-carboxylic acid. In another embodiment, the present invention relates to a method for preventing and / or treating inflammation, improving intestinal health and the health of the lining of the GI tubule, and reducing the lining of the intestinal tubule and other GI, gastric cell infiltration, and intestinal symptoms associated with patients with CTLA-4 checkpoint-associated immunodeficiency using an AhR ligand having biosimilar activity. In some embodiments, the AhR ligand is 1-methylindole-3-carboxylic acid.

[0097] In some aspects, this disease or condition includes CTLA-4 checkpoint-associated immunodeficiency, characterized by various combinations of enteropathy, hypogammaglobulinemia, recurrent respiratory infections, granulomatous lymphocytic interstitial lung disease, lymphocyte infiltration of non-lymphoid organs (intestine, lungs, brain, bone marrow, kidneys), autoimmune thrombocytopenia or neutropenia, autoimmune hemolytic anemia, and lymphadenopathy. CTLA-4 checkpoint-associated immunodeficiency is diagnosed based on clinical symptoms, laboratory findings, and genetic testing. Patients with only one functional copy of the gene encoding CTLA-4 suffer from severe autoimmunity. These heterozygous mutations result in a novel phenotype, accompanied by hyperactivated T and B cell infiltration into non-lymphoid organs such as the intestine, lungs, and brain, along with more typical signs of autoimmunity. Some clinical symptoms in CTLA-4 haploinsufficiency are reminiscent of those observed on biopsies of inflamed organs. Patients with primary CTLA-4 checkpoint-associated immunodeficiency may have recurrent respiratory infections, hypogammaglobulinemia, autoimmune cytopenia, autoimmune enteropathy, and granulomatous infiltrative lung disease.

[0098] One example of CTLA-4 checkpoint-associated immunodeficiency is "CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI)," a CTLA-4 checkpoint-associated immunodeficiency disorder characterized by heterozygous CTLA4 mutations in humans associated with severe immunomodulation. Another example of CTLA-4 checkpoint-associated immunodeficiency is lipopolysaccharide-responsive beige-like anchor (LRBA), a disorder characterized by a gene carrying one or more genetically biallelemic mutations in the patient. LRBA deficiency also leads to secondary loss of CTLA-4, so this recessive disorder is called LRBA deficiency with autoantibodies, T-reg cell deficiency, autoimmune infiltration, and enteropathy (LATAIE).

[0099] The description of asymptomatic adults with heterozygous CTLA4 deficiency (Kuehn et al, 2014 - Schubert et al, 2014), and the wide range of ages of onset among symptomatic individuals, suggests that additional interacting factors are required to cross the autoimmune threshold. These factors may include other genetic or epigenetic events and environmental influences (microbes or others). Exposure to the microbiome of certain frequently affected organs (skin and gut) suggests that this environmental factor may contribute to the development of autoimmunity in patients receiving the CTLA4 blocker ipilimumab. Interestingly, CTLA4 - / - Even when mice are returned to a sterile environment, they die within two weeks of age, suggesting the importance of autoantigens in driving inflammatory phenotypes (Tivol et al, 1995).

[0100] Microbial indole is a highly attractive postbiotic because it has been shown to extend the health span across a wide range of evolutionarily diverse species from different phyla (Descamps et al, 2019). The gastrointestinal tract contains many species capable of synthesizing indole and indole-containing compounds, which may act as contributing factors to imbalances in the microbiome by influencing host immunoresponsiveness, epithelial barrier function, and pathogen colonization (Roager et al, 2018). By acting as a ligand for AhR, a transcription factor that controls the biodegradation of endogenous and exogenous toxins, it prevents inflammatory damage and provides barrier integrity (Hubbard et al, 2015 b). Indole exerts multiple functions, including bidirectional communication with the microbiome to fine-tune host immunity, tolerance, and metabolism (Stockinger, 2014). Indole administration to germ-free mice has been shown to increase the expression of epithelial tight junction proteins and attenuate indicators of inflammatory bowel disease (Shimada et al, 2013). Similarly, the therapeutic effect of dextran-induced colitis in a mouse model has been reported in 3-IAld (Zelante et al, 2013).

[0101] Activating innate lymphoid cells type 3 to produce IL-22, 3-IAld enhanced barrier integrity and antimicrobial peptide production in mouse models of colitis, gastrointestinal, and vaginal candidiasis (Zelante et al, 2013; Borghi et al, 2019; Puccetti et al, 2021). Treatment with 3-IAld also limited intestinal epithelial damage, reduced transepithelial bacterial migration, and decreased inflammatory cytokine production, which reduced graft-versus-host disease (GvHD) (a systemic inflammatory state initiated by donor T cells causing colitis) in mouse models. 3-IAld treatment also resulted in recipient-specific tolerance of transplanted T cells. Transcriptional profiling and gene ontology analysis showed that 3-IAld administration upregulated genes associated with the type I interferon response, which is known to protect against radiation-induced bowel injury (Swimm et al, 2018). Therefore, 3-IAld may limit intestinal inflammation and damage by acting through different downstream effector pathways, potentially providing a treatment option for patients at risk of gastrointestinal tract inflammation-induced damage. This increases the potential for developing microbiome-derived indoles or their derivatives to promote human epithelial barrier function.

[0102] Given that CTLA-4 checkpoint-associated immunodeficiency shares similarities with the underlying mechanisms of anti-CTLA-4 therapy (Bakacs et al, 2015), the therapeutic capacity of 3-IAlds may extend beyond CTLA-4 checkpoint-associated immunodeficiency to include the prevention of adverse immune events associated with cancer treatment with immune checkpoint inhibitors, including colitis and enteritis (Karamchandani et al, 2018; Marin-Acevedo et al, 2018). While cancer immunotherapy using immune checkpoint inhibitors has been highly successful, their therapeutic benefits are limited by various resistance mechanisms (Schoenfeld et al, 2020) or associated toxic effects, including frequent gastrointestinal, endocrine, and cutaneous toxicity, as well as potentially fatal neurotoxicity and cardiotoxicity (Choi et al, 2020). Therefore, novel therapeutic strategies that offer manageable side effects to existing immunotherapies would enhance and expand their therapeutic efficacy and applications. Studies have shown that the presence of commensal bacteria is necessary for the effectiveness of immunotherapy against various tumors (Iida et al, 2013). Clinical trials have supported these findings with compelling evidence that microbial abundance and diversity are associated with a durable response to immunotherapy using gut microbiota signatures that predict toxicity associated with the combination of CTLA-4 and PD-1 blockade (Andrews et al, 2021).

[0103] Another example of CTLA-4 checkpoint-related immunodeficiency is regulatory T (Treg) cell deficiency, which refers to any defective immune surveillance response that can lead to reduced elimination of chronically virus-infected cells and oncogenic viruses, resulting in uncontrolled cell proliferation. One example is decreased CTLA-4 expression, which can lead to uncontrolled T cell proliferation, potentially resulting in abnormal proliferation of autoreactive clones beyond EBV-specific T cell clones, for example.

[0104] Another example of CTLA-4 checkpoint-related immunodeficiency is intestinal disease, which refers to ongoing damage or irritation and swelling of the small intestine. Another example of CTLA-4 checkpoint-related immunodeficiency is enteritis, which refers to inflammation of the gastrointestinal (GI) tract. If the inflammation persists, damage to the GI tract can occur.

[0105] Another example of CTLA-4 checkpoint-related immunodeficiency disorder is immune-mediated colitis (IMC), a common immune-related adverse event associated with immune checkpoint inhibitors, characterized by abdominal pain, mucus or blood in the stool, and fever. Immune checkpoint inhibitors are immunotherapy drugs that work by preventing checkpoint proteins from binding to their partner proteins. This prevents the "off" signal from being sent, allowing T cells to kill cancer cells. Dysregulation of T cells can also contribute to IMC, in which these T cells attack GI ductal cells and tissues.

[0106] Therapeutic compositions, such as 3-IAld, may be administered in doses based on the body weight or mass of the individual to whom the therapeutic composition is administered. Body weight-adjusted doses of the therapeutic composition can be administered at approximately 1 mg / kg to approximately 25 mg / kg. Body weight-adjusted doses of the therapeutic composition can be administered at approximately 3 mg / kg to approximately 25 mg / kg, approximately 10 mg / kg to approximately 25 mg / kg, approximately 15 mg / kg to approximately 20 mg / kg, or approximately 15 mg / kg to approximately 18 mg / kg. Doses of 3-IAld and / or 3-IAld pharmaceutical compositions may be delivered in immediate-release, delayed-release, or continuous-release formulations.

[0107] Therapeutic compositions, such as 1-methylindole-3-carboxylic acid, may be administered in doses based on the body weight or mass of the individual to whom the therapeutic composition is administered. Body weight-adjusted doses of the therapeutic composition can be administered at approximately 1 mg / kg to approximately 25 mg / kg. Body weight-adjusted doses of the therapeutic composition can also be administered at approximately 3 mg / kg to approximately 25 mg / kg, approximately 10 mg / kg to approximately 25 mg / kg, approximately 15 mg / kg to approximately 20 mg / kg, or approximately 15 mg / kg to approximately 18 mg / kg. Doses of 1-methylindole-3-carboxylic acid and / or 1-methylindole-3-carboxylic acid pharmaceutical compositions may be delivered in immediate-release, delayed-release, or continuous-release formulations.

[0108] IV. Pharmaceutical Compositions The compounds of the present invention may be incorporated into pharmaceutical compositions suitable for administration to a target. Such compositions typically include the active compound and a pharmaceutically acceptable carrier. As used herein, the term "pharmaceutically acceptable carrier" includes any solvent, dispersion medium, coating, isotonic agent, and absorption retarder that is compatible with the administration of the drug. The use of such media and agents (factors) for pharmaceutically active substances is well known in the art. Unless any conventional media or agent (factor) is incompatible with the active compound, its use in the composition is intended. As described above, auxiliary active compounds may also be incorporated into the composition.

[0109] The pharmaceutical compositions of the present invention are formulated to suit their intended route of administration. Examples of routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g., swallowing), transdermal (topical), transmucosal, and rectal administration. In one embodiment, the active compound is prepared with a carrier that protects the compound from rapid elimination from the body, such as a controlled-release formulation including implants and microencapsulation delivery systems. Biodegradable biocompatible polymers such as ethylene vinyl acetate, polyanhydride, polyglycolic acid, collagen, polyorthoesters, and polylactic acid may be used. Methods for preparing such formulations will be apparent to those skilled in the art.

[0110] The pharmaceutical composition may be included in a container, pack, or dispenser along with the instructions for administration.

[0111] Exemplary aspects provided herein In one embodiment (Aspect 1; A1), the Specified provides a method for treating CTLA-4 checkpoint-associated immunodeficiency in a patient, comprising administering a therapeutically effective amount of a pharmaceutical composition containing 1H-indole-3-carboxyaldehyde (3-IAld) to a patient in need thereof.

[0112] In one aspect of A1, namely A2, CTLA-4 checkpoint-associated immunodeficiency is selected from the following group: CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI), lipopolysaccharide-responsive beige-like anchor (LRBA) deficiency (LATAIE), regulatory T (Treg) cell deficiency, autoimmune infiltration, intestinal disease, enteritis, immune-mediated colitis, gastrointestinal disorders, and gastric atrophy.

[0113] In one aspect of A1 or A2, namely A3, CTLA-4 checkpoint-associated immunodeficiency is immune-mediated colitis.

[0114] In any one embodiment of A1 to A3, namely A4, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier.

[0115] In one embodiment of A4, namely A5, the pharmaceutically acceptable carrier is at least one polymer.

[0116] In one aspect of A4, namely A6, the pharmaceutically acceptable carriers are a group of polymers.

[0117] In one embodiment of A6, namely A7, the group of polymers is Eudragit® polymer.

[0118] In any one embodiment from A1 to A7, namely A8, the pharmaceutical composition is formulated for intestinal delivery.

[0119] In any one embodiment of A1 to A8, namely A9, the pharmaceutical composition is administered orally.

[0120] In any one embodiment of A1 to A9, namely A10, the pharmaceutical composition is in a form selected from capsules, tablets, gel tablets, gel capsules, gels, liquids, and gums.

[0121] In any one embodiment of A10, namely A11, in some embodiments, the pharmaceutical composition is in the form of a tablet or a capsule.

[0122] In any one embodiment of A1 to A11, namely A12, the pharmaceutical composition is administered at intervals of every other day (qod).

[0123] In any one embodiment of A1 to A12, namely A13, the pharmaceutical composition is administered in a 3-IAld dose of at least about 3 mg / kg, at least about 4 mg / kg, at least about 5 mg / kg, at least about 6 mg / kg, at least about 7 mg / kg, at least about 8 mg / kg, at least about 9 mg / kg, at least about 10 mg / kg, at least about 11 mg / kg, at least about 12 mg / kg, at least about 13 mg / kg, at least about 14 mg / kg, at least about 15 mg / kg, at least about 16 mg / kg, at least about 17 mg / kg, or at least about 18 mg / kg.

[0124] In one embodiment of A13, namely A14, the 3-IAld dose is approximately 18 mg / kg.

[0125] In one embodiment, namely A15, the Specified provides a method for treating CTLA-4 checkpoint-associated immunodeficiency in a patient, comprising administering a therapeutically effective amount of a pharmaceutical composition comprising 1-methylindole-3-carboxylic acid to a patient in need thereof.

[0126] In one aspect of A15, namely A16, CTLA-4 checkpoint-associated immunodeficiency is selected from the following group: CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI), lipopolysaccharide-responsive beige-like anchor (LRBA) deficiency (LATAIE), regulatory T (Treg) cell deficiency, autoimmune infiltration, intestinal disease, enteritis, immune-mediated colitis, gastrointestinal disorders, and gastric atrophy.

[0127] In one aspect of A15 or A16, namely A17, CTLA-4 checkpoint-associated immunodeficiency is immune-mediated colitis.

[0128] In any one embodiment of A15 to A17, namely A18, the pharmaceutical composition includes a pharmaceutically acceptable carrier.

[0129] In one embodiment of A18, namely A19, the pharmaceutically acceptable carrier is at least one polymer.

[0130] In one embodiment of A18, namely A20, the pharmaceutically acceptable carriers are a group of polymers.

[0131] In one embodiment of A20, namely A21, the group of polymers is Eudragit® polymer.

[0132] In any one embodiment of A15 to A21, namely A22, the pharmaceutical composition is formulated for intestinal delivery.

[0133] In one embodiment of A15 to A22, namely A23, the pharmaceutical composition is administered orally.

[0134] In any one embodiment of A15 to A23, namely A24, the pharmaceutical composition is in a form selected from capsules, tablets, gel tablets, gel capsules, gels, liquids, and gums.

[0135] In one aspect of A24, namely A25, the pharmaceutical composition is in the form of a tablet or a capsule.

[0136] In any one embodiment of A15 to A25, namely A26, the pharmaceutical composition is administered at intervals of every other day (qod).

[0137] In any one embodiment of A15 to A23, namely A27, the pharmaceutical composition is administered in doses of 1-methylindole-3-carboxylic acid of at least about 2 mg / kg, at least about 3 mg / kg, at least about 4 mg / kg, at least about 5 mg / kg, at least about 6 mg / kg, at least about 7 mg / kg, at least about 8 mg / kg, at least about 9 mg / kg, at least about 10 mg / kg, at least about 11 mg / kg, at least about 12 mg / kg, at least about 13 mg / kg, at least about 14 mg / kg, at least about 15 mg / kg, at least about 16 mg / kg, at least about 17 mg / kg, or at least about 18 mg / kg.

[0138] In any one embodiment of A15 to A23, namely A28, the pharmaceutical composition is administered at a dose of 1-methylindole-3-carboxylic acid of approximately 2.25 mg / kg.

[0139] The present invention will be further described by the following embodiments, but this should not be construed as a further limitation. The contents of all references cited throughout this application are expressly incorporated herein by reference.

[0140] Unless otherwise indicated, the implementation of this disclosure will employ conventional techniques in cell biology, cell culture, molecular biology, transgenic biology, microbiology, recombinant DNA, and immunology within the scope of the art. Such techniques are adequately described in the literature. For example, Sambrook et al., ed. (1989) Molecular Cloning A Laboratory Manual (2nd ed.; Cold Spring Harbor Laboratory Press); Sambrook et al., ed. (1992) Molecular Cloning: A Laboratory Manual, (Cold Springs Harbor Laboratory, NY); DNGlover ed., (1985) DNA Cloning, Volumes I and II; Gait, ed. Synthesis; Mullis et al. U.S. Patent No. 4,683,195; Hames and Higgins, eds. (1984) Nucleic Acid Hybridization; Hames and Higgins, eds. (1984) Transcription And Translation; Freshney (1987) Culture Of Animal Cells (Alan R. Liss, Inc.); Immobilized Cells And Enzymes (IRL Press)(1986);Perbal(1984)A Practical Guide To Molecular Cloning;the treatise,Methods In Enzymology(Academic Press,Inc.,NY);Miller and Calos eds.(1987)Gene Transfer Vectors For Mammalian Cells,(Cold Spring Harbor Laboratory);Wu et al.,eds.,Methods In Enzymology,Vols.154 and 155;Mayer and Walker,eds.(1987) Immunochemical Methods In Cell And Molecular Biology (Academic Press, London); Weir and Blackwell, eds., (1986) Handbook Of Experimental Immunology, Volumes I-IV; Manipulating the Mouse Embryo, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, (1986)); Crooke, Antisense drug Technology: Principles, Strategies and Applications,2. nd See Ed. CRC Press (2007) and Ausubel et al. (1989) Current Protocols in Molecular Biology (John Wiley and Sons, Baltimore, Md.).

[0141] All of the references cited above, and all references cited herein, are incorporated herein by reference in their entirety.

[0142] The following examples are provided for illustrative purposes only and are not limiting. [Examples]

[0143] Example 1 A 3-IAld+ anti-CTLA-4 induced mouse colitis model in dextran sulfate sodium (DSS) prevents inflammatory bowel lesions.

[0144] Given that CTLA-4 checkpoint-associated immunodeficiency shares similarities with the underlying mechanisms of anti-CTLA-4 therapy (Bakacs et al, 2015), it was hypothesized that the therapeutic capacity of 3-IAlds may exist not only in CTLA-4 checkpoint-associated immunodeficiency but also in the prevention of adverse immune events associated with cancer treatment with immune checkpoint inhibitors, including colitis and enteritis, e.g., intestinal disease or gastritis (Karamchandani et al, 2018; Marin-Acevedo et al, 2018). It is known that a mouse model of colitis, e.g., pathology caused by CTLA4 blockade in C57BL / 6 mice (Scott et al, 2020), mimics the pathogenic effects of hereditary human CTLA-4 haploinsufficiency in the gut (Bakacs et al, 2015).

[0145] In the study, C57BL / 6 mice were given 3% dextran sulfate sodium (DSS) in drinking water, and 100 μg of anti-CTLA-4 monoclonal antibody (mAb) or isotype control antibody was administered intraperitoneally at 0, 4, and 8 days after DSS administration, as shown in Figure 2A. DSS is a water-soluble, negatively charged sulfated polysaccharide with a highly variable molecular weight ranging from 5 to 1400 kDa. Sulfated polysaccharides do not directly induce intestinal inflammation, but rather act as a chemotoxin directly on the colonic epithelium, causing damage to epithelial cells. Eudragit preparation 3-IAld (Puccetti et al, 2018) was administered intragastrally at a dose of 18 mg / kg every other day, as illustrated in Figure 2A. The animals were observed daily for diarrhea, bloody stools, weight loss, and overall survival. One week after DSS treatment (14 days after the start of treatment), at the point when the model replicates human disease (Manicassamy et al, 2014), surviving mice were sacrificed, the colon was resected, and macroscopic damage and local immune parameters were evaluated. 3-IAld treatment increased survival rate (Figure 2B) and body weight (Figure 2C) and decreased disease activity index (Figure 1D). Surviving mice showed improvement in macroscopic pathology (Figure 2E), as well as i) improvement in the normal structural composition of the colon (Figures 2F and 2G), ii) improvement in epithelial barrier function as revealed by ZO-1 expression (Figure 2H), and iii) improvement in epithelial cell proliferation and regeneration as revealed by Ki-67 staining (Figure 2H). These results demonstrate that 3-IAld has the ability to prevent CTLA4 blocker-induced intestinal inflammatory lesions in a mouse model of colitis.

[0146] 3-IAld also promoted the expression of Lgr5, Nfil3, and Muc1, which are cell surface mucins that function as barriers against infection and regulators of inflammation.

[0147] After confirming the functional recovery of C57BL / 6 mice as shown in Figure 2, it was found that 3-IAld also promotes the expression of Lgr5, Nfil3, and Muc1, cell surface mucins that function as barriers against infection and regulators of inflammation (Dhar et al, 2019) (Figure 3A). Consequently, the passage of dextran-FITC across the intestinal barrier was reduced (Figure 3B), and the level of soluble CD14, a marker of intestinal permeability, also decreased (Figure 3C). These changes were paralleled by a shift to an anti-inflammatory profile with decreased levels of TNF-α, IL-1β, and IL-17A, as well as an increase in IL-10 (Figure 3D). Consistent with the anti-inflammatory profile, the level of calprochitin also decreased (Figure 3E). Considering that 3-IAld is deficient in mice with colitis (Alexeev et al, 2018), these results suggest that 3-IAld supplementation can protect against DSS + anti-CTLA-4 induced colitis by maintaining epithelial barrier integrity and suppressing inflammatory responses. This activity is consistent with the AhR agonist activity (Zelante et al, 2013-Puccetti et al. 2021) of the antimicrobial peptide Reg3γ (Figure 3F), the AhR-dependent genes Cyp1a1 and Ahrr (Figure 3F), and, importantly, of IL-22 (Figure 3G, both gene and protein), a key mediator of mucosal functional activity in response to 3-IAld (Renga et al. 2022), as indicated by its AhR agonist activity.

[0148] Example 2 3-IAld in an immune-mediated colitis model

[0149] The activity of 3-IAld was evaluated in an immune-mediated model of colitis, which, unlike chemically induced colitis models, is thought to best reproduce the pathophysiology of immune-dependent colitis as it occurs in patients with CTLA-4 haploinsufficiency (Consnt et al, 2022) or those treated with checkpoint inhibitors (Westdorp et al, 2021). Since CTLA-4 deletion in mice leads to marked lymphoproliferation and fatal multi-organ tissue destruction, causing early lethality (Tivol et al, 1995), an alternative reference model of immune-mediated colitis (Kiesler et al, 2015) was used.

[0150] Humanized immunodeficiency NSG mice were injected with human peripheral blood mononuclear cells from a healthy donor and treated with an anti-CTLA-4 monoclonal antibody. 8-10 week old NOD.Cg-Prkdcscid Il2rgtm1Wjl / SzJ(NSG) mice were injected with newly isolated human peripheral blood mononuclear cells from a healthy donor (10 7 The drug was administered intraperitoneally, and 100 μg of anti-CTLA-4 mAb or human IgG was used as an antibody control for intraperitoneal treatment on days 0, 4, 8, 12, and 16.

[0151] 3-IAld (18 mg / kg) was administered intragastricly every other day, starting from the day of cell infusion. Mice were observed daily for overall mortality and weight loss, and sacrificed on day 21. These results demonstrated that treatment with 3-IAld increased survival (Figure 4A), reduced weight loss (Figure 4B), improved inflammatory histopathology (Figure 4C) and histological scores (Figure 4D), and restored barrier function, as evidenced by decreased ZO-1 expression (Figure 4E) and inflammatory cytokine gene expression (Figure 4F).

[0152] 3-IAld limits the progression of immune-mediated colitis in RAG1-deficient mice.

[0153] Immunodeficiency Rag1 - / - Mouse Naive CD4 +Reconstituted with T cells and treated with anti-CTLA-4 monoclonal antibody. Rag1- / - mice (mice homozygous for the Rag1tm1Mom mutation) do not produce mature T cells or B cells. Their phenotype can be described as "non-leaky" immunodeficiency. 8- to 10-week-old Rag1- / - mice were injected intraperitoneally with 4 × 10 5 naive CD4 + T cells purified from the spleen of C57BL / 6 mice, and on the day following T cell reconstitution, they were then treated with anti-CTLA-4 mAb or control IgG on days 0, 4, 8, 12, and 16.

[0154] 3-IAld (18 mg / kg) was administered intragastrically every other day starting from the day of cell injection. Mice were observed daily for overall mortality and weight loss and sacrificed on day 21. Similar to what was observed in NSG mice, treatment with 3-IAld reduced weight loss (Figure 5A), and both gross pathology (Figure 5B) and colonic histopathology (Figures 5C and 5D) were improved.

[0155] IL-10-deficient (Il10 - / - ) mice treated short-term or long-term with anti-CTLA-4 monoclonal antibody, with or without DSS.

[0156] Due to the spontaneous onset of post-weaning intestinal inflammation and the pathological changes first seen in 3-week-old mice, IL-10-deficient mice (Il10 - / - ) are an excellent model reflecting the etiology of immune-mediated colitis (Keubler et al, 2015), characterized by discontinuous and transmural inflammatory lesions, cell infiltration, mainly lymphocyte infiltration into the lamina propria and submucosa of the mucosa, epithelial hyperplasia, mucin depletion, ulcers, and thickening of the intestinal wall (Berg et al, 1996; Gomes-Santos et al, 2012). Il10 - / - mice develop mild colitis by 10 weeks of age, and the severity reaches a plateau at 16 weeks of age (Gomes-Santos et al, 2012). The inventors used Il10 of different ages (i.e., 8 - 10 weeks old or 16 weeks old) - / -The activity of 3-IAld after short-term or long-term treatment with anti-CTLA-4 mAb in mice, with or without 1% DSS, was evaluated. The model and results are described below.

[0157] Il10 at 8-10 weeks old - / - DSS + anti-CTLA-4 induced colitis in mice.

[0158] Il10 - / - Mice were intraperitoneally administered 1% DSS in drinking water and 100 μg of anti-CTLA-4 monoclonal antibody on days 0, 4, and 8 after DSS administration. 3-IAld-Eudragit was administered intragastricly every other day throughout the experiment, starting two days prior to treatment (18 mg / kg 3-IAld). Animals were monitored daily for body weight and colonic inflammatory lesions until 14 days after DSS administration. The results showed weight loss (Figure 6A) and increased epithelial thickening (Figures 6B and C) in 8-10 week old IL-10 deficient mice; both features worsened after DSS + anti-CTLA-4 treatment, with further weight loss, worsening colonic pathology, and signs of epithelial destruction and transmural infiltration. 3-IAld treatment prevented weight loss (Figure 6A) and reduced inflammatory lesions and disease activity index (Figures 6B and C).

[0159] Il10 - / - Short-term anti-CTLA-4 treatment in mice at 8-10 weeks of age

[0160] Il10 - / - Mice were treated with anti-CTLA-4 monoclonal antibody and 3-IAld alone, without DSS, as described above (Figure 6). Animals were monitored daily for body weight and inflammatory lesions of the colon, and sacrificed 14 days after the initial administration of anti-CTLA-4 monoclonal antibody. The results showed that treatment with anti-CTLA-4 alone resulted in weight loss (Figure 7A) and accelerated epithelial thickening associated with marked submucosal infiltration in 8-10 week old IL-10 deficient mice, in contrast to wild-type mice (Figure 7B). 3-IAld treatment prevented weight loss (Figure 7A) and improved inflammatory lesions (Figure 7B), particularly by reducing large-scale cell infiltration.

[0161] Il10 - / - Long-term anti-CTLA-4 treatment in mice at 16 weeks of age.

[0162] Il10 - / - Mice were treated with 100 μg of anti-CTLA-4 monoclonal antibody and 3-IAld alone, without DSS, as shown in Figure 8A. Animals were monitored daily for body weight, disease activity index, clinical laboratory tests, inflammatory pathology of the ileum and colon, epithelial barrier function, and immunological parameters of inflammation. The results showed that long-term treatment with anti-CTLA-4 alone resulted in decreased body weight (Figure 8B) and disease activity index (Figure 8C), increased clinical (Figure 8D) and histological lesions in the ileum and colon (Figures 8E and 8F), accelerated epithelial damage (as evidenced by decreased ZO-1 expression) and dysfunction (as evidenced by staining with bromodeoxyuridine (BrdU), a thymidine analog that incorporates DNA for cell division during the S phase of the cell cycle), increased levels of inflammatory calprotein (Figure 8G) and IL-6 (Figure 8H), while decreased expression of the antimicrobial peptide Reg3γ (Figure 8H). Treatment with 3-IAld was effective in counteracting the cytotoxic effects induced by CTLA-4 blockade. Consistent with the large-scale T-cell infiltration in various organs observed in CTLA-4 haploinsufficiency (Kuehn et al, 2014; Schubert et al, 2014; Schwab et al, 2018), co-administration of 3-IAld significantly reduced large-scale CD3 infiltration in the ileum and colon (Figure 9A), lungs and liver (Figure 9B) of mice treated with anti-CTLA-4, as quantified in Figure 9. + The inventors discovered T cell infiltration. No lymphocyte infiltration or immune lesions were detected in organs such as the spleen and kidneys (data not shown).

[0163] 3-IAld treatment slows disease progression in 16-week-old IL-10-deficient mice.

[0164] As shown in Figure 10A, 16-week-old IL-10-deficient mice (Il10 - / -The mice were treated with 3-IAld every other day for 4 weeks. As a mimic of disease exacerbation, mice were administered 1% DSS after 3-IAld treatment, and then evaluated for body weight changes, intestinal lesions, and epithelial integrity. The results showed that 3-IAld treatment clearly delayed body weight loss (Figure 10B) and colon inflammatory histopathology (Figure 10D), and maintained epithelial integrity and regeneration (Figure 10D).

[0165] Example 3 3-IAld promotes beneficial microbiomes.

[0166] Considering the role of the AhR / IL-22 axis in maintaining the balance of the microbiome, the effects of 3-IAld on fecal microbial composition were evaluated. The activity of the 3-IAld-modified microbiome in colitis was assessed using fecal microbiome transplantation. Feces were collected from untreated or 3-IAld-treated mice for 6 days and transplanted into recipient mice at the onset of colitis (FMT). In contrast to fecal transplantation from untreated mice, fecal transplantation from 3-IAld-treated mice prevented weight loss in DSS-treated mice (Figure 11A) and improved macroscopic and histopathological aspects of the colon (Figures 11B-11D). Similar results were obtained in DSS + anti-CTLA-4-treated mice, where FMT from 3-IAld-treated mice also prevented weight loss (Figure 11E) and induced IL-10-producing regulatory T cells (Treg), as evidenced by the reversal of DNA hypermethylation of the Foxp3 promoter (Figure 11F). Overall, these results suggest that the beneficial activity of 3-IAld can arise through various pathways, including enhancement of the intestinal barrier via the AhR / IL-22 axis, modification of the composition and function of the microbiome, and regulation of inflammation via Treg cells.

[0167] Example 4 3-IAld does not interfere with the development of antitumor immunity.

[0168] Potential applications of 3-IAld require that its immunomodulatory activity not interfere with tumor immune surveillance. For this purpose, the effects of 3-IAld were evaluated in an anti-CTLA-4 responsive B16 melanoma model (Renga et al, 2022). 3-IAld, consistent with increased expression of the leukocyte recruitment chemokine Cxcl9 and effector perforin (Figure 12E), did not alter tumor growth, nor interfere with the therapeutic effect of anti-CTLA-4 antibodies (Figures 12A-12B), and CD4 + and CD8 + It also did not affect the recruitment of tumor-infiltrating lymphocytes (Figures 12C-12D). Similarly, 3-IAld did not interfere with the therapeutic effect of anti-PD-1 antibodies in a model of Lewis lung cancer (LLC). In fact, 3-IAld increased survival (Figure 12F), reduced tumor growth (Figure 12G), improved macroscopic pathology (Figure 12H), and reduced Foxp3 in the lung. + CD25 + It did not interfere with the ability of anti-PD-1 antibodies to reduce the recruitment of Treg cells (Figures 12I-12J). Therefore, the beneficial activity of 3-IAld does not interfere with tumor immune surveillance.

[0169] This study demonstrates the potential of bacterial metabolites such as 3-IAld as biologics that can mitigate anti-CTLA-4 induced enteropathy without interfering with tumor surveillance.

[0170] This suggests that 3-IAld is a unique molecule capable of disrupting the dynamic feedforward loop of inflammation in the gut, thereby suggesting that the tissue-destructive effects of infiltrating lymphocytes in the CTLA-4 deficiency state can lead to a microbial imbalance that further promotes inflammation and lesions. By acting dually on both the host and microbial sides, 3-IAld is particularly suitable for breaking this vicious cycle of pathogenicity and for calming inflammation.

[0171] Example 5 Considering that hereditary human CTLA-4 haploinsufficiency shares similarities with the underlying mechanisms of anti-CTLA-4 therapy (Bakacs et al, 2015), a relevant model for demonstrating the efficacy of 3-IAld is a mouse model of immune checkpoint inhibitor-induced colitis in combination with anti-CTLA-4 and dextran sulfate sodium (DSS) (Wang et al, 2018-Wang et al, 2019; Perez Riuz et al, 2019). To further supplement the medical validity of our findings, we evaluated the activity of 3-IAld in an immune-mediated model of colitis that, while different from chemically induced colitis models, is considered to best reproduce the pathophysiology of immune-dependent colitis as it occurs in patients with CTLA-4 haploinsufficiency (Constant et al, 2022) or in patients treated with checkpoint inhibitors (Westdorp et al, 2021).

[0172] Pharmacology Major pharmacodynamic studies were conducted. The mechanism of action of 3-IAld is based on binding to AhR, both in vitro and in vivo. In vitro, 3-IAld induced luciferase activity in H1L1.1c2 cell lines containing stably transfected AhR-responsive firefly luciferase in a dose range of 0.1–100 mM (Zelante et al, 2013). In vivo, intragastric administration of 3-IAld induced IL-22 production in an AhR-dependent manner in a mouse model of mucosal candidiasis and DSS-induced colitis (Zelante et al, 2013). Preliminary pharmacokinetic data were obtained by administering Eudragit-formulated 3-IAld to a composition of 90% 3-IAld / 10% 13C8-labeled 3-IAld, and performing targeted / untargeted mass spectrometry analysis in serum at different time points (30 min, 1 h, 2 hours, 4.5 hours, 6 hours, 24 hours). Analysis of unlabeled 3-IAld revealed small fluctuations above the basic endogenous level of 3-IAld at different time points, as shown in Figure 13A. However, analysis of labeled 3-IAld showed a clear peak between 30 min and 1 hour after administration, with a sharp decrease in levels at 2 hours, while trace amounts were detected at subsequent time points (Figure 13B). These data demonstrate the rapid metabolic transformation of 3-IAld. Further analysis of potential metabolites of unlabeled 3-IAld revealed a peak level of the oxidized form of 3-IAld (indole-3-formic acid or Ox-3-IAld) at 30 min at an average of 80 μM, rapidly decreasing to 20 μM after 1 hour, and then becoming detectable trace amounts at subsequent time points (Figure 13C). Lower levels of methylated forms of indole-3-formic acid, namely 1-methylindole-2-carboxylic acid and methyl indole-3-carboxylate, were also detected at peaks of approximately 150 and 25 nM at 30 min, respectively, with decreasing levels at 1 hour, thus following the same kinetics as indole-3-formic acid (Figure 13D). The presence of indole-3-formic acid was also confirmed with labeled 3-IAld (Figure 13E), showing similar kinetics and a peak level of approximately 25 μM at 30 min.

[0173] Further in vitro experiments demonstrated that indole-3-formic acid is pharmacologically active and therefore may contribute to the pharmacodynamics of 3-IAld. To demonstrate this, we exposed human cell line HepG2, a liver cancer cell line, to either the ligand indole-3-formic acid or Ox-3-IAld at different concentrations. Indole-3-formic acid and / or Ox-3-IAld were able to induce the AhR activation marker Cyp1A1 in a dose-dependent manner in the range of 1 to 100 μM, as shown in Figure 14.

[0174] toxicology Eudragit formulation 3-IAld (Puccetti et al, 2018) was administered intragastricly to naive C57BL / 6 and AhR- / - mice at the indicated dose every three weeks. At the end of the treatment, the mice were sacrificed, and tissue lesions were examined blindly in different organs after hematoxylin and eosin (H&E) staining. As shown in Figure 15, no visible tissue lesions were observed in any organ examined at the two different concentrations.

[0175] It should be understood that the interpretation of the claims is intended to be based on the section describing the modes for carrying out the invention, rather than the section describing the summary and abstract of the invention. The summary and abstract section may describe one, but not all, exemplary embodiments of the disclosure as contemplated by the inventors(s) and is therefore not intended to limit the scope of the disclosure and the accompanying claims in any way.

[0176] The foregoing description relating to specific embodiments is intended to fully illustrate the general nature of the disclosure, and others can readily modify and / or adapt such specific embodiments to various uses without excessive experimentation and without departing from the general concepts of the disclosure, by applying knowledge within the scope of the art in that field. Therefore, such adaptations and modifications are intended to be within the meaning and scope of equivalents of the embodiments of the disclosure, based on the teachings and guidance presented herein. The language or terminology used herein is for illustrative purposes only and not limiting purposes, and it should be understood that the language or terminology used herein should be interpreted by those skilled in the art in light of the teachings and guidance.

[0177] The scope and width of this disclosure should not be limited by any of the exemplary embodiments described above, but should be defined solely in accordance with the following claims and their equivalents.

[0178] All cited references throughout this application (including references, U.S. or foreign patents or patent applications, and websites) are expressly incorporated herein by reference as if they were included in their entirety for any purpose. In the event of any conflict, the content disclosed herein in writing shall prevail.

[0179] While various specific embodiments are illustrated and described, the above specifications are not limiting. It will be understood that various modifications can be made without departing from the spirit and scope of the invention(s). Many modifications will become apparent to those skilled in the art upon further examination of this specification.

[0180] Example 6 1-methylindole-3-carboxylic acid for use in the prevention of intestinal lesions in cases of CTLA-4 blockade or dysfunction.

[0181] As shown in Example 5, rapid metabolic conversion of 3-IAld occurs after administration. Specifically, oxidized 3-IAld (indole-3-formic acid) and methylated indole-3-formic acid were detected after administration of labeled 3-IAld. For example, 1-methylindole-3-carboxylic acid was detected at the same reaction rate as indole-3-formic acid, with peak levels obtained at 30 minutes (Figures 13C and 13D). Several studies have evaluated the biological activity of methylindole and methoxyindole as aryl hydrocarbon receptor (AhR) activators. Preliminary data obtained in vitro in cell lines have shown that 1-methylindole-3-carboxylic acid dose-dependently activates AhR-dependent genes (data not shown). Given 3-IAld's ability to induce AhR-dependent genes and protect the mucosal barrier from damage, it was hypothesized that 1-methylindole-3-carboxylic acid may have similar therapeutic potential to prevent CTLA-4 blockade-induced inflammatory bowel lesions in a mouse model of colitis, which is known to mimic the pathogenic effects of hereditary human CTLA-4 haploinsufficiency in the gut (Bakacs et al, 2015).

[0182] In the study, C57BL / 6 mice were given 3% dextran sulfate sodium (DSS) in drinking water, and 100 μg of anti-CTLA-4 monoclonal antibody (mAb) or isotype control antibody was administered intraperitoneally at 0, 4, and 8 days after DSS administration, as shown in Figure 17A. Eudragit-formulated 1-methylindole-3-carboxylic acid was administered intragastricly every other day at scaling doses of 0.09, 0.18, or 0.36 mg / mouse (Figure 17A). Animals were monitored daily for the appearance of diarrhea, bloody stools, and weight loss. One week after DSS treatment (14 days after the start of treatment), when the mouse model replicated human disease (Manicassamy et al, 2014), surviving mice were sacrificed, the colon was resected, and macroscopic injury and local immune parameters were evaluated. Mice treated with 1-methylindole-3-carboxylic acid showed significantly reduced weight loss (Figure 17B), decreased disease activity index (Figure 17C), and protection from clinical morbidity and rectal bleeding (Figure 17D). Positive effects were observed at each dose of 1-methylindole-3-carboxylic acid, including 0.09 mg / mouse dose. Mice treated with 1-methylindole-3-carboxylic acid showed i) restoration of normal architectural structure of the colon and ileum (Figures 17E-17F), as evidenced by ZO-1 expression, and ii) maintenance of epithelial barrier function (Figures 17E-17F). CTLA-4 haploinsufficiency typically presents with widespread CD3+ T cell infiltration across different organs (Kuehn et al, 2014; Schubert et al, 2014; Schwab et al, 2018). Extensive CD3+ T cell infiltration in the ileum and colon of mice treated with anti-CTLA-4 was significantly reduced by concomitant administration of 1-methylindole-3-carboxylic acid (Figures 17E-17F). Administration of 1-methylindole-3-carboxylic acid was also effective in reducing CD3+ T cell infiltration in the lungs (Figure 18). These results indicate that 1-netylindole-3-carboxylic acid, a metabolite of 3-IAld, has the ability to prevent CTLA4-blocker-induced intestinal inflammatory lesions in a mouse model of colitis, similar to treatment with 3-IAld, as shown in Figure 2.

[0183] 1-Methylindole-3-carboxylic acid is a potent activator of AhR-dependent genes.

[0184] After confirming functional recovery in C57BL / 6 mice as shown in Figure 17, 1-methylindole-3-carboxylic acid was also found to promote an anti-inflammatory profile in the colon, reducing Il1b expression levels and increasing Il10 expression levels (Figure 19). These results indicate that supplementation with 1-methylindole-3-carboxylic acid may protect against DSS + anti-CTLA-4 induced colitis by maintaining epithelial barrier integrity and suppressing inflammatory responses. Consistent with its in vitro AhR agonist activity (data not shown), 1-methylindole-3-carboxylic acid induced the expression of the AhR-dependent gene Cyp1a1, the antimicrobial peptide Reg3g, and Il22 (Figure 19), a key mediator of AhR-dependent mucosal protective activity (Stockinger et al, 2021). These results demonstrate the potent activity of 1-methylindole-3-carboxylic acid in neutralizing the intestinal morbidity associated with CTLA-4 deficiency during CLTA-4 blockade by activating an at least partially protective AhR / IL-22-dependent pathway.

[0185] Example 7 Pharmacology of 3-IAld / 1ME3CA

[0186] Pharmacokinetics A single-dose pharmacokinetic (PK) study was conducted in healthy C57BL / 6 mice to compare 1ME3CA and 3-IAld at a topo dose of 0.36 mg. Untreated naive mice were used as controls. Four animals were sacrificed at 0.5, 2, 6, and 24 hours. Blood and organs (brain, lungs, intestines, liver, and kidneys) were evaluated by cardiac puncture and collected in EDTA-containing tubes. All samples were stored at -80°C until use. To assess the functional activity of 1ME3CA and 3-IAld, AhR downstream gene expression was also evaluated in these mice. [Table 1]

[0187] Pharmacokinetic data were obtained after a single oral administration of 18 mg / kg of 1ME3CA or 3-IAld (both formulated in enteric-coated microparticles) to C57BL / 6 mice, and targeted analysis was performed by mass spectrometry at different time points (30 minutes, 2 hours, 6 hours, and 24 hours) in the intestines, serum, lungs, liver, brain, and kidneys. As shown in Figure 20A, 3-IAld was rapidly eliminated in most tissues (after 2 hours), and its terminal half-life ranged from 3 to 5 hours depending on the tissue, as reported (Puccetti et al, Int J Pharm. 2021, 602:120610). Analysis of potential metabolites of 3-IAld revealed that indole-3-carboxylic acid was the major metabolite in serum and other tissues, and that 1ME3CA was present at a lower level, with a tmax of 30 minutes observed in all tissues tested, including the brain, and rapid clearance from all organs examined. Compared to 3-IAld, the concentration of 1ME3CA was basal in the intestines, serum, and kidneys, while it was significantly higher in the lungs, liver, and brain, at 10³–10⁵ nmol / kg. Such observations suggest the formation of 1ME3CA as a secondary metabolite of 3IAld and / or indole-3-carboxylic acid. Upon reaching tmax, 3-IAld rapidly disappeared in most tissues (after 2 hours), and the terminal half-lives were in the same range of 3 for all compounds, suggesting the extremely rapid in vivo transformation of 3-IAld, as well as indole-3-carboxylic acid.

[0188] The above considerations were confirmed by examining the analysis of 1ME3CA PK after enteric-coated administration (Figure 20B). Indeed, 3-IAld and indole-3-carboxylic acid concentrations were basal in all organs, confirming the role of 1ME3CA as a metabolite of 3-IAld and / or indole-3-carboxylic acid. The tmax values ​​(30 mins) were the same, and the Cmax values ​​were remarkably similar in all organs and serum. The profiles were similar despite 1ME3CA being undetectable in serum, lungs, and kidneys at 6 hours. Furthermore, in mice administered with 3-IAld, 1ME3CA levels were still higher than in controls 24 hours after administration, when 1ME3CA had been eliminated (at least in some organs, intestines, liver, and brain). Systemic levels of 1ME3CA after oral administration appeared to be more persistent and, as expected, significantly higher than those observed after oral administration of 3-IAld.

[0189] Pharmacodynamics - in vitro activity 3-IAld has been shown to induce luciferase activity in H1L1.1c2 cell lines containing stably transfected AhR-responsive firefly luciferase in a dose range of 0.1–100 μM (Zelante et al. Immunity. 2013;39:372-85). The ability of 1ME3CA was comparatively evaluated here in mouse hepatocarcinoma cells (H1L6.1c3) donated by Allison K. Ehrlich (Meyer Hall, University of California, Davis, United States) containing a stably integrated AhR xenobiotic reactive element driven by the firefly luciferase reporter plasmid, pGudLuc6.167. Cells were plated in MAMA (Gibco) in 24-well plates, supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin solution, and stimulated with either 3-IAld or 1ME3CA at different concentrations. Luciferase activity, calculated as relative light units (RLU) per microgram of protein and expressed as an induction factor, was retained after 2-hour or 24-hour exposure. These results clearly demonstrated that 1ME3CA is a more potent inducer of luciferase activity than 3-IAld, as observed by higher RLU at 2 hours and persisting up to 24 hours at a 100 mM concentration (Figure 21).

[0190] To further characterize the AhR activating properties of 1ME3CA, the ability of 3-IAld and 1ME3CA to activate AhR-downstream target genes (Cyp1a1, Cyp2a1, and AhRR) was comparatively evaluated in A549 cell lines derived from adenocarcinoma human alveolar basal epithelium, Calu-3 cell lines derived from adenocarcinoma human bronchiolar epithelial cells, CaCo-2 cell lines derived from human colon cancer, and HepG32 human liver cancer cell lines. After exposing cells to 1, 10, 100, or 1000 μM of a molecule or DMSO at 37°C for 4 hours or overnight (on), the expression of the above genes was evaluated by RT-PCR. The results showed the following: i) 1ME3CA promotes the expression of AhR-dependent Cyp1a1, Cyp2a1, and AhRR genes in the tested cell lines to a degree comparable to, or even superior to, that of 3-IAld at reference AhR ligands ITE or FICZ and similar concentrations, i.e., 100 μM; ii) both molecules act within the optimal concentration range of 10–100 μM; at higher concentrations of 1000 μM, inconsistent results were obtained, specifically that AhR activity increased in CaCo-2 and HepG32 cell lines but not in Calu-3 cells; iii) this activity appeared to occur early, after 4 hours of exposure, and then be maintained (Figures 22A–D). No activity was observed in the expression of IDO1 and IDO2 genes, which were evaluated for their ability to activate AhR. In summary, these data suggest that the AhR agonist activity of 1ME3CA is comparable to that of 3-IAld, even if it does not exceed that of 3-IAld.

[0191] Pharmacodynamics - Dose-dependent in vitro activity In DSS + anti-CTLA-4 induced colitis, mice were administered intragastric Eudragit every other day, starting 4 days before DSS treatment and continuing until the mice were sacrificed. Disease activity was evaluated in mice in terms of histopathology, barrier permeability parameters, and intestinal inflammation.

[0192] The scaling doses of 0.09–0.045–0.022 mg / mouse (corresponding to 4.5, 2.25, and 1.12 mg / kg) in a mouse model of immune-mediated colitis (DSS + anti-CTLA-4) were evaluated as shown in Table 2 below. [Table 2]

[0193] C57BL / 6 mice were treated with DSS in drinking water for one week, followed by a one-week recovery period, and then administered 100 μg of anti-CTLA-4 mAb and 1ME3CA as shown in the experimental schedule in Figure 23. Mice were evaluated for body weight change (Figure 23B), disease activity index (Figure 23C), clinical morbidity and rectal bleeding (Figure 23D), histology of the colon and ileum (PAS staining) (Figure 23E), and ×40 magnification (scale bar, 100 μm). Each in vivo experiment included 4-6 mice per group (20 mice in each experiment).

[0194] To confirm the effect of 1ME3CA on lymphocyte infiltration, mice were treated as before, and histological changes (PAS staining) were evaluated. High-resolution microscope images were taken at 10x magnification (scale bar, 400 μm) for the lungs and spleen, and at 20x magnification (scale bar, 200 μm) for the liver and kidneys. As shown in Figure 24, 1ME3CA dose-dependently reduced lymphocyte infiltration in anti-CTLA-4 treated mice, with an effective dose of 3-IAld being 18 mg / kg and an effective dose of 1ME3CA being 2.25 mg / kg.

[0195] toxicology To evaluate toxicity, C57BL / 6 mice were administered intragastric doses of 3-IAld or 1ME3CA, formulated in enteric-coated microparticles, at increasing doses of 0.36–0.18–0.09 mg / mouse, for up to 3 weeks, as shown in Table 3 below. At the end of treatment, the mice were sacrificed, and tissue lesions were examined blindly in different organs after PAS staining of various sections derived from each organ. A portion of the organs of four mice in each group were evaluated separately. The results (Figures 25A and 25B) show no visible tissue lesions in any of the organs examined at the three different concentrations. [Table 3] References

[0196] Alexeev EE, Lanis JM, Kao DJ, et al.Microbiota-Derived indole metabolites promote human and murine intestinal homeostasis through regulation of interleukin-10 receptor. Am J Pathol 2018;188:1183-94.

[0197] Andrews MC, Duong CPM, Gopalakrishnan V, Iebba V, Chen WS, Derosa L, et al. Gut microbiota signatures are associated with toxicity to combined CTLA-4 and PD-1 blockade. Nat Med.2021 Aug;27(8):1432-41.doi:10.1038 / s41591-021-01406-6.

[0198] Attia P,et al.Autoimmunity correlates with tumor regression in patients with metastatic melanoma treated with anti-cytotoxic T-lymphocyte antigen-4. J Clin Oncol.2005;23(25):6043-53.doi:10.1200 / JCO.2005.06.205.

[0199] Ayrignac X,Goulabchand R,Jeziorski E,et al.Two neurologic facets of CTLA4-related haploinsufficiency. Neurol Neuroimmunol Neuroinflamm.2020;7(4):e751.

[0200] Bakacs T,Mehrishi JN. Anti-CTLA-4 therapy may have mechanisms similar to those occurring in inherited human CTLA4 haploinsufficiency. Immunobiology.2015;220(5):624-5.

[0201] Bakhtiar S,Gamez-Diaz L,Jarisch A,Soerensen J,Grimbacher B,Belohradsky B,Keller KM,Rietschel C,Klingebiel T,Koletzko S,Albert MH,Bader P(2017)Treatment of Infantile Inflammatory Bowel Disease and Autoimmunity by Allogeneic Stem Cell Transplantation in LPS-Responsive Beige-Like Anchor Deficiency. Frontiers in immunology 8:52.

[0202] Berg DJ,Davidson N,Kuhn R,et al.Enterocolitis and colon cancer in interleukin-10-deficient mice are associated with aberrant cytokine production and CD4(+)TH1-like responses. J Clin Invest.1996;98:1010-1020.

[0203] Besnard C,Levy E,Aladjidi N,Stolzenberg MC,et al.Pediatric-onset Evans syndrome:Heterogeneous presentation and high frequency of monogenic disorders including LRBA and CTLA4 mutations. Clin Immunol.2018;188:52-57.doi:10.1016 / j.clim.2017.12.009.

[0204] Borghi M,et al.Targeting the Aryl Hydrocarbon Receptor with Indole-3-Aldehyde Protects from Vulvovaginal Candidiasis via the IL-22-IL-18 Cross-Talk. Front Immunol.2019;10:2364.

[0205] Bratanic N,Kovac J,Pohar K,Trebusak Podkrajsek K,Ihan A,Battelino T,Avbelj Stefanija M.Multifocal gastric adenocarcinoma in a patient with LRBA deficiency. Orphanet J Rare Dis.2017 Jul 18;12(1):131.

[0206] Choi J,Lee SY. Clinical Characteristics and Treatment of Immune-Related Adverse Events of Immune Checkpoint Inhibitors. Immune Netw.2020;20(1):e9.

[0207] Compare D,Rocco A,Nardone G.Risk factors in gastric cancer. Eur Rev Med Pharmacol Sci.2010;14(4):302-8.

[0208] Constant BD,Dutmer CM,Arnold MA,Hall C,Abbott JK,de Zoeten E.Cytotoxic T-Lymphocyte-Associated Antigen 4 Haploinsufficiency Mimics Difficult-to-Treat Inflammatory Bowel Disease. Clinical Gastroenterology and Hepatology 2022;20:e696-e702.

[0209] Cunningham-Rundles C,Bodian C.Common variable immunodeficiency:clinical and immunological features of 248 patients. Clin Immunol. Jul 1999;92(1):34-48.doi:10.1006 / clim.1999.4725.

[0210] Dai X,Zhu BT. Indoleamine 2,3-dioxygenase tissue distribution and cellular localization in mice:implications for its biological functions. J Histochem Cytochem.2010;58(1):17-28.

[0211] Descamps HC,Herrmann B,Wiredu D,Thaiss CA. The path toward using microbial metabolites as therapies. EBioMedicine.2019;44:747-754.

[0212] Dhalla F,da Silva SP,Lucas M,Travis S,Chapel H.Review of gastric cancer risk factors in patients with common variable immunodeficiency disorders,resulting in a proposal for a surveillance programme. Clin Exp Immunol.2011;165(1):1-7.

[0213] Dhar P,McAuley J.The role of the cell surface mucin MUC1 as a barrier to infection and regulator of inflammation. Front Cell Infect Microbiol 2019;9:117.

[0214] Egg D,et al.Increased Risk for Malignancies in 131 Affected CTLA4 Mutation Carriers. Front Immunol.2018;9:2012.

[0215] Egg D,et al.Therapeutic options for CTLA-4 insufficiency. J Allergy Clin Immunol.2021;S0091-6749(21)00891-5.

[0216] Esser C,Rannug A.The aryl hydrocarbon receptor in barrier organ physiology,immunology,and toxicology. Pharmacol Rev.2015;67(2):259-79.

[0217] Gamez-Diaz L,August D,Stepensky P,Revel-Vilk S,Seidel MG,Noriko M,Morio T,Worth AJJ,Blessing J,Van de Veerdonk F,Feuchtinger T,Kanariou M,Schmitt-Graeff A,Jung S,Seneviratne S,Burns S,Belohradsky BH,Rezaei N,Bakhtiar S,Speckmann C,Jordan M,Grimbacher B(2016)The extended phenotype of LPS-responsive beige-like anchor protein(LRBA)deficiency. The Journal of allergy and clinical immunology 137(1):223-230.

[0218] Garcia-Perez JE,Baxter RM,Kong DS,Tobin R,McCarter M,Routes JM,Verbsky J,Jordan MB,Dutmer CM,Hsieh EWY. CTLA4 Message Reflects Pathway Disruption in Monogenic Disorders and Under Therapeutic Blockade. Front Immunol.2019;10:998.

[0219] Gomes-Santos AC,Garcias Moreira T,Barbosa Castro-Junior A,Coelho Horta B,Lemos L,Nogueira Cruz D,Freitas Guimaraes MA,Cara DM,McCafferty D-M,and Caetano Fari AM. New Insights into the Immunological Changes in IL-10-Deficient Mice during the Course of Spontaneous Inflammation in the Gut Mucosa. Clin Dev Immunol 2012;2012:560817.

[0220] Hashimoto T,et al.ACE2 links amino acid malnutrition to microbial ecology and intestinal inflammation. Nature. Jul 25 2012;487(7408):477-81.

[0221] He Y,Li X,Yu H,Ge Y,Liu Y,Qin X,Jiang M,Wang X.The Functional Role of Fecal Microbiota Transplantation on Dextran Sulfate Sodium-Induced Colitis in Mice. Front Cell Infect Microbiol.2019;9:393.

[0222] Hodi FS,et al.Improved survival with ipilimumab in patients with metastatic melanoma. N Engl J Med.2010;363(8):711-23.

[0223] Hoshino A,Toyofuku E,Mitsuiki N,et al.Clinical Courses of IKAROS and CTLA4 Deficiencies:A Systematic Literature Review and Retrospective Longitudinal Study. Front Immunol.2022;12:784901.

[0224] Hubbard TD,Murray IA,Bisson WH,Lahoti TS,Gowda K,Amin SG,Patterson AD,Perdew GH.Adaptation of the human aryl hydrocarbon receptor to sense microbiota-derived indoles. Sci Rep.2015;5:12689. (Hubbard et al 2015 a).

[0225] Hubbard TD,Murray IA,Perdew GH.Indole and Tryptophan Metabolism:Endogenous and Dietary Routes to Ah Receptor Activation. Drug Metab Dispos.2015;43(10):1522-35. (Hubbard et al 2015 b).

[0226] Iida N,Dzutsev A,Stewart CA,Smith L,Bouladoux N,Weingarten RA,et al.Commensal bacteria control cancer response to therapy by modulating the tumor microenvironment. Science.2013;342(6161):967-70.

[0227] Jamee M,Hosseinzadeh S,Sharifinejad N,Zaki-Dizaji M,Matloubi M,Hasani M,Baris S,Alsabbagh M,Lo B,Azizi G.Comprehensive comparison between 222 CTLA-4 haploinsufficiency and 212 LRBA deficiency patients:a systematic review. Clin Exp Immunol.2021;205(1):28-43.

[0228] Karamchandani DM,Chetty R.Immune checkpoint inhibitor-induced gastrointestinal and hepatic injury:pathologists’ perspective. J Clin Pathol.2018;71(8):665-671.

[0229] Keubler LM,Buettner M, Hager C, Bleich A.A Multihit Model:Colitis Lessons from the Interleukin-10-deficient Mouse.Inflamm Bowel Dis 2015;21:1967-1975.

[0230] Kiesler P,Fuss IJ,Strober W.Experimental Models of Inflammatory Bowel Diseases. Cell Mol Gastroenterol Hepatol.2015 Mar 1;1(2):154-170.

[0231] Konopelski P,Ufnal M.Indoles-Gut Bacteria Metabolites of Tryptophan with Pharmacotherapeutic Potential.Curr Drug Metab.2018;19(10):883-890.

[0232] Kuehn HS,et al.Immune dysregulation in human subjects with heterozygous germline mutations in CTLA4. Science.2014;345(6204):1623-1627.

[0233] Kumar KK,Burgess AW,Gulbis JM. Structure and function of LGR5:an enigmatic G-protein coupled receptor marking stem cells Protein Sci 2014;23(5):551-65.

[0234] Lamas B,et al.CARD9 impacts colitis by altering gut microbiota metabolism of tryptophan into aryl hydrocarbon receptor ligands. Nat Med.2016;22(6):598-605.

[0235] Lanz AL,Riester M,Peters P,Schwerd T,et al.Abatacept for treatment-refractory pediatric CTLA4-haploinsufficiency. Clin Immunol.2021;229:108779.

[0236] Lee JS,et al.AHR drives the development of gut ILC22 cells and postnatal lymphoid tissues via pathways dependent on and independent of Notch. Nat Immunol.2011;13(2):144-51.

[0237] Levy E,Stolzenberg MC,Bruneau J,Breton S,Neven B,Sauvion S,Zarhrate M,Nitschke P,Fischer A,Magerus-Chatinet A,Quartier P,Rieux-Laucat F.LRBA deficiency with autoimmunity and early onset chronic erosive polyarthritis. Clin Immunol.2016 Jul;168:88-93.

[0238] Lo B,Fritz JM,Su HC,Uzel G,Jordan MB,Lenardo MJ.CHAI and LATAIE:new genetic diseases of CTLA-4 checkpoint insufficiency. Blood.2016 Aug 25;128(8):1037-42.

[0239] Manicassamy S,Manoharan I.Mouse models of acute and chronic colitis. Methods Mol Biol.2014;1194:437-48.

[0240] Marin-Acevedo JA,Dholaria B,Soyano AE,Knutson KL,Chumsri S,Lou Y.Next generation of immune checkpoint therapy in cancer:new developments and challenges. J Hematol Oncol.2018;11(1):39.

[0241] Moraes-Fontes MF,Hsu AP,Caramalho I,Martins C,Araujo AC,Lourenco F,Taulaigo AV,Llado A,Holland SM,Uzel G.Fatal CTLA-4 heterozygosity with autoimmunity and recurrent infections:a de novo mutation. Clin Case Rep.2017;5(12):2066-2070.

[0242] Opitz CA,et al.An endogenous tumour-promoting ligand of the human aryl hydrocarbon receptor. Nature.2011;478(7368):197-203.

[0243] Perez-Ruiz E,et al.Prophylactic TNF blockade uncouples efficacy and toxicity in dual CTLA-4 and PD-1 immunotherapy. Nature.2019;569(7756):428-432.

[0244] Puccetti M,Giovagnoli S,Zelante T,Romani L,Ricci M.Development of Novel Indole-3-Aldehyde-Loaded Gastro-Resistant Spray-Dried Microparticles for Postbiotic Small Intestine Local Delivery. J Pharm Sci.2018;107(9):2341-2353.

[0245] Puccetti M,Pariano M,Borghi M,Barola C,Moretti S,Galarini R,Mosci P,Ricci M,Costantini C,Giovagnoli S.Enteric formulated indole-3-carboxaldehyde targets the aryl hydrocarbon receptor for protection in a murine model of metabolic syndrome. Int J Pharm.2021;602:120610.

[0246] Qiu J,et al.The aryl hydrocarbon receptor regulates gut immunity through modulation of innate lymphoid cells. Immunity.2012;36(1):92-104.

[0247] Renga G,Bellet MM,Pariano M,Gargaro M,Stincardini C,D’Onofrio F,et al.Thymosin alpha1protects from CTLA-4 intestinal immunopathology. Life Sci Alliance.2020;3(10).

[0248] Renga G,Nunzi E,Pariano M,Puccetti M,Bellet MM,Pieraccini G,D’Onofrio F,et al.Optimizing therapeutic outcomes of immune checkpoint blockade by a microbial tryptophan metabolite. J Immunother Cancer.2022;(3):e003725.

[0249] Roager HM,Licht TR. Microbial tryptophan catabolites in health and disease. Nat Commun.2018;9(1):3294.

[0250] Safe S,Jayaraman A,Chapkin RS. Ah receptor ligands and their impacts on gut resilience:structure-activity effects. Crit Rev Toxicol.2020;50(6):463-473.

[0251] Salavoura K,Kolialexi A,Tsangaris G,Mavrou A.Development of cancer in patients with primary immunodeficiencies. Anticancer Res.2008;28(2B):1263-9.

[0252] Schoenfeld AJ,Hellmann MD. Acquired Resistance to Immune Checkpoint Inhibitors. Cancer Cell.2020;37(4):443-55.

[0253] Schubert D,et al.Autosomal dominant immune dysregulation syndrome in humans with CTLA4 mutations. Nat Med.2014;20(12):1410-1416.

[0254] Schwab C,Gabrysch A,Olbrich P,et al.Phenotype,penetrance,and treatment of 133 cytotoxic T-lymphocyte antigen 4-insufficient subjects. J Allergy Clin Immunol.2018;142(6):1932-1946.doi:10.1016 / j.jaci.2018.02.055.

[0255] Scott SA,Fu J,Chang PV. Microbial tryptophan metabolites regulate gut barrier function via the aryl hydrocarbon receptor. Proc Natl Acad Sci U S A.2020;117(32):19376-19387.doi:10.1073 / pnas.2000047117.

[0256] Semo Oz R,M ST(2019)Arthritis in children with LRBA deficiency-case report and literature review. Pediatric rheumatology online journal 17(1):82.

[0257] Shimada Y,et al.Commensal bacteria-dependent indole production enhances epithelial barrier function in the colon. PLoS One.2013;8(11):e80604.

[0258] Soler-Palacin P,Garcia-Prat M,Martin-Nalda A,Franco-Jarava C,Riviere JG,Plaja A,Bezdan D,Bosio M,Martinez-Gallo M,Ossowski S,Colobran R(2018)LRBA Deficiency in a Patient With a Novel Homozygous Mutation Due to Chromosome 4 Segmental Uniparental Isodisomy. Frontiers in immunology 9:2397.

[0259] Spits H,et al.Innate lymphoid cells--a proposal for uniform nomenclature. Nat Rev Immunol.2013;13(2):145-9.

[0260] Stockinger B,Di Meglio P,Gialitakis M,Duarte JH.The aryl hydrocarbon receptor:multitasking in the immune system. Annu Rev Immunol.2014;32:403-32.

[0261] Stockinger B,Shah K,Wincent E.AHR in the intertinal microenvironment:safeguarding barrier function. Nat Rev Gastroenterol Hepatol.2021;18(8):559-570.

[0262] Swimm A,et al.Indoles derived from intestinal microbiota act via type I interferon signaling to limit graft-versus-host disease. Blood.2018;132(23):2506-2519. Tesch VK,Abolhassani H,Shadur B,Zobel J,Mareika Y,Sharapova S,Karakoc-Aydiner E,Riviere JG,Garcia-Prat M,Moes N,Haerynck F,Gonzales-Granado LI,Santos Perez JL,Mukhina A,Shcherbina A,Aghammarohammadi L,Agustrom F,Haskologlu S,Ikinciogullari AI,Kostel Bal S,Baris S,Kilic SS,Karaca NE,Kutukculer N,Girschick H,Kolios A,Keles S,Uygun V,Stepensky P,Worth A,van Montfrans JM,Peters AMJ,Mayts I,Adeli M,Mardem,Adzoh,Khozah,Chozah,N,Khazah Z,Avbelj Stefanija M,Bakhtiar S,Florkin B,Meeths M,Gamez L,Grimbacher B,Seppanen MRJ,Lankester A,Gennery AR,Seidel MG(2020)Long-term outcome of LRBA deficiency in 76 patients after various modal treatment by immune dysregulation and evaluation activity(IDDA)score. The Journal of allergy and clinical immunology 145(5):1452-1463.

[0263] Tesi B,Priftakis P,Lindgren F,Chiang SC,Kartalis N,Lofstedt A,Lorinc E,Henter JI,Winiarski J,Bryceson YT,Meeths M.Successful Hematopoietic Stem Cell Transplantation in a Patient with LPS-Responsive Beige-Like Anchor(LRBA)Gene Mutation. J Clin Immunol.2016 Jul;36(5):480-9.

[0264] Tivol EA,Borriello F,Schweitzer AN,Lynch WP,Bluestone JA,Sharpe AH.Loss of CTLA-4 leads to massive lymphoproliferation and fatal multiorgan tissue destruction,revealing a critical negative regulatory role of CTLA-4. Immunity.1995;3(5):541-7.

[0265] Van Leeuwen EM,Cuadrado E,Gerrits AM,Witteveen E,de Bree GJ.Treatment of Intracerebral Lesions with Abatacept in a CTLA4-Haploinsufficient Patient. J Clin Immunol.2018;38(4):464-467.

[0266] Wang F,Yin Q,Chen L,Davis MM. Bifidobacterium can mitigate intestinal immunopathology in the context of CTLA-4 blockade. Proc Natl Acad Sci U S A.2018;115(1):157-161.

[0267] Wang T,et al.Probiotics Lactobacillus reuteri Abrogates Immune Checkpoint Blockade-Associated Colitis by Inhibiting Group 3 Innate Lymphoid Cells. Front Immunol.2019;10:1235.

[0268] Westdorp et al.,Mechanisms of Immune Checkpoint Inhibitor-Mediated Colitis Front Immunol.2021;12:768957.

[0269] Wikoff WR,et al.Metabolomics analysis reveals large effects of gut microflora on mammalian blood metabolites. Proc Natl Acad Sci U S A.2009;106(10):3698-703.

[0270] Yu X,Wang Y,Deng M,et al.The basic leucine zipper transcription factor NFIL3 directs the development of a common innate lymphoid cell precursor. Elife 2014;3 10.7554 / eLife.04406.

[0271] Zelante T,et al.Tryptophan catabolites from microbiota engage aryl hydrocarbon receptor and balance mucosal reactivity via interleukin-22. Immunity.2013;39(2):372-85.

[0272] Zhang LS,Davies SS. Microbial metabolism of dietary components to bioactive metabolites:opportunities for new therapeutic interventions. Genome Med.2016;8(1):46.

Claims

1. A method for treating CTLA-4 checkpoint-associated immunodeficiency in a patient, comprising administering a therapeutically effective amount of a pharmaceutical composition containing 1H-indole-3-carboxyaldehyde (3-IAld) to a patient in need thereof.

2. The method according to claim 1, wherein the CTLA-4 checkpoint-related immunodeficiency is selected from the group consisting of CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI), lipopolysaccharide-responsive beige-like anchor (LRBA) deficiency (LATAIE), regulatory T (Treg) cell deficiency, autoimmune infiltration, intestinal disease, enteritis, immune-mediated colitis, gastrointestinal disorders, and gastric atrophy.

3. The method according to any one of claims 1 or 2, wherein the CTLA-4 checkpoint-associated immunodeficiency is immune-mediated colitis.

4. The method according to any one of claims 1 to 3, wherein the pharmaceutical composition comprises a pharmaceutically acceptable carrier.

5. The method according to claim 4, wherein the pharmaceutically acceptable carrier is at least one polymer.

6. The method according to claim 4, wherein the pharmaceutically acceptable carrier is a group of polymers.

7. The method according to claim 6, wherein the group of polymers is Eudragit® polymer.

8. The method according to any one of claims 1 to 7, wherein the pharmaceutical composition is formulated for intestinal delivery.

9. The method according to any one of claims 1 to 8, wherein the pharmaceutical composition is administered orally.

10. The method according to any one of claims 1 to 9, wherein the pharmaceutical composition is in a form selected from capsules, tablets, gel tablets, gel capsules, gels, liquids, and gums.

11. The method according to claim 10, wherein the pharmaceutical composition is in the form of a tablet or a capsule.

12. The method according to any one of claims 1 to 11, wherein the pharmaceutical composition is administered at intervals of one day (q.o.d).

13. The method according to any one of claims 1 to 12, wherein the pharmaceutical composition is administered in a 3-IAld dose of at least about 3 mg / kg, at least about 4 mg / kg, at least about 5 mg / kg, at least about 6 mg / kg, at least about 7 mg / kg, at least about 8 mg / kg, at least about 9 mg / kg, at least about 10 mg / kg, at least about 11 mg / kg, at least about 12 mg / kg, at least about 13 mg / kg, at least about 14 mg / kg, at least about 15 mg / kg, at least about 16 mg / kg, at least about 17 mg / kg, or at least about 18 mg / kg.

14. The method according to claim 13, wherein the dose of 3-Iald is approximately 18 mg / kg.

15. A method for treating CTLA-4 checkpoint-associated immunodeficiency in a patient, comprising administering a therapeutically effective amount of a pharmaceutical composition containing 1-methylindole-3-carboxylic acid to the patient in need thereof.

16. The method according to claim 15, wherein the CTLA-4 checkpoint-related immunodeficiency is selected from the group consisting of CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI), lipopolysaccharide-responsive beige-like anchor (LRBA) deficiency (LATAIE), regulatory T (Treg) cell deficiency, autoimmune infiltration, intestinal disease, enteritis, immune-mediated colitis, gastrointestinal disorders, and gastric atrophy.

17. The method according to any one of claims 15 or 16, wherein the CTLA-4 checkpoint-associated immunodeficiency is immune-mediated colitis.

18. The method according to any one of claims 15 to 17, wherein the pharmaceutical composition comprises a pharmaceutically acceptable carrier.

19. The method according to claim 18, wherein the pharmaceutically acceptable carrier is at least one polymer.

20. The method according to claim 18, wherein the pharmaceutically acceptable carrier is a group of polymers.

21. The method according to claim 20, wherein the group of polymers is Eudragit® polymer.

22. The method according to any one of claims 15 to 21, wherein the pharmaceutical composition is formulated for intestinal delivery.

23. The method according to any one of claims 15 to 22, wherein the pharmaceutical composition is administered orally.

24. The method according to any one of claims 15 to 23, wherein the pharmaceutical composition is in a form selected from capsules, tablets, gel tablets, gel capsules, gels, liquids, and gums.

25. The method according to claim 24, wherein the pharmaceutical composition is in the form of a tablet or a capsule.

26. The method according to any one of claims 15 to 25, wherein the pharmaceutical composition is administered at intervals of one day (q.o.d).

27. The method according to any one of claims 15 to 23, wherein the pharmaceutical composition is administered in a dose of 1-methylindole-3-carboxylic acid of at least about 2 mg / kg, at least 3 mg / kg, at least about 4 mg / kg, at least about 5 mg / kg, at least about 6 mg / kg, at least about 7 mg / kg, at least about 8 mg / kg, at least about 9 mg / kg, at least about 10 mg / kg, at least about 11 mg / kg, at least about 12 mg / kg, at least about 13 mg / kg, at least about 14 mg / kg, at least about 15 mg / kg, at least about 16 mg / kg, at least about 17 mg / kg, or at least about 18 mg / kg.

28. The method according to any one of claims 15 to 23, wherein the pharmaceutical composition is administered at a dose of about 2.25 mg / kg of 1-methylindole-3-carboxylic acid.

Images