WO2017216253A1 - Cd1a en tant que molécule cible thérapeutique et marqueur diagnostique pour maladies intestinales inflammatoires - Google Patents

Cd1a en tant que molécule cible thérapeutique et marqueur diagnostique pour maladies intestinales inflammatoires Download PDF

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WO2017216253A1
WO2017216253A1 PCT/EP2017/064597 EP2017064597W WO2017216253A1 WO 2017216253 A1 WO2017216253 A1 WO 2017216253A1 EP 2017064597 W EP2017064597 W EP 2017064597W WO 2017216253 A1 WO2017216253 A1 WO 2017216253A1
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cells
ibd
expressing
colitis
level
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Roswitha GROPP
Michael FÖHLINGER
Pia PALAMIDES
Matthias SIEBECK
Omar AL-AMODI
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Klinikum Der Universität München
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
    • G01N2333/705—Assays involving receptors, cell surface antigens or cell surface determinants
    • G01N2333/70503—Immunoglobulin superfamily, e.g. VCAMs, PECAM, LFA-3
    • G01N2333/70539—MHC-molecules, e.g. HLA-molecules
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/06—Gastro-intestinal diseases
    • G01N2800/065—Bowel diseases, e.g. Crohn, ulcerative colitis, IBS

Definitions

  • CD1a AS THERAPEUTIC TARGET MOLECULE AND DIAGNOSTIC MARKER FOR INFLAMMATORY BOWEL DISEASES
  • the present invention relates to a novel therapeutic target and diagnostic marker of chronic inflammatory bowel diseases, such as ulcerative colitis (UC) or Crohn's Disease (CD).
  • This diagnostic marker and therapeutic target is the lipid receptor CD1 a, which is a transmembrane glycoprotein present on antigen presenting cells (APC), and mediates the presentation of primarily lipid and glycolipid antigens to T-cells.
  • APC antigen presenting cells
  • IBD Inflammatory bowel disease
  • GI ulcerative colitis
  • Ulcerative colitis affects the colon and rectum and typically involves only the innermost lining or mucosa, manifesting as continuous areas of inflammation and ulceration.
  • Crohn's disease is a transmural inflammation affecting all layers of the intestine that can affect any portion of the digestive tract from mouth to anus, but is predominantly seen in the terminal ileum and/or colon. Intestinal inflammation and ulceration in Crohn's disease is asymmetrical and occurs in "patches,” with areas of healthy tissue interspersed, and extends deeply into the intestinal wall, forming granulomatous lesions. Several categories of Crohn's disease have been described, defined by the portion of the digestive tract involved and the presenting symptomatology. The symptoms are often more variable than ulcerative colitis depending on which part of the bowel is involved. Clinical symptoms of IBD include abdominal cramps and pain, (bloody) diarrhea fever, loss of appetite, weight loss and anemia.
  • the conventional diagnostic methods require invasive surgery or potentially damaging and costly radiological techniques, such as CT scans or MRI. Therefore there is a need for a less invasive and more cost-efficient method to diagnose an IBD in a subject.
  • therapeutic strategies can be employed to treat the disease.
  • efficacy of the applied therapeutic strategy is assessed by monitoring the symptoms and by evaluating the condition of the affected tissue, which is carried out through the use of endoscopic, radiological, and pathologic examinations, as is done for the initial diagnosis of the IBD.
  • a novel marker has been identified by the applicant of the present invention, which can be used to cost-efficiently diagnose an IBD in a subject suspected of suffering from an IBD. 15
  • the diagnostic method does not require an invasive surgical procedure, and can be carried out by measuring the level of the marker in a sample obtained from the patient.
  • the novel marker is CD1 a.
  • CD1 a has been known for decades as a phenotypic marker of human epidermal Langerhans cells (LC). Like other members of the CD1 family CD1 a presents lipids,
  • CD1 a activates T-cells resulting in the release of IL-22, IL-13 and IFNy (de Jong et al., 2010; de Jong et al., 2014).
  • CD1 a macrophages and monocytes have the capacity to activate cytotoxic CD8+ cells.
  • LC located in the epidermis and the corresponding lipid ligands concentrated
  • PLA2 releases fatty acids from phospholipids in the extracellular space where loading of CD1 a takes place.
  • CD1 a bearing macrophages or monocytes have not been detected in the colon, nor have they been known to be associated with the gut or with IBD, such as UC.
  • CD1 a expressing cell populations in particular at least 35 one of CD1 a expressing leukocytes, CD1 a expressing CD1 1 b+ macrophages, and CD1 a expressing CD14+ monocytes, are associated with IBD, such as UC.
  • IBD such as UC.
  • the ratio or percentage or proportion of CD1 a expressing cells within a specific cell population has been found to be a signal for IBD.
  • CD1 a level The ratio of the number of a specific type of CD1 a expressing cells in a sample to the total number of this type of cells measured in the sample is referred to as CD1 a level.
  • An association with IBD has been found if the ratio or percentage or proportion of CD1 a expressing cells in a population of these cells is above a threshold as defined below and in the claims. For example, if all leukocytes are counted in a sample, the CD1 a level is the ratio of the number of CD1 a expressing leukocytes to the total number of leukocytes measured.
  • the CD1 a level is the ratio of the number of CD1 a expressing CD1 1 + macrophages to the total number of CD1 1 + macrophages measured. If only the CD14+ monocytes are measured, then the CD1 a level is the ratio of the number of CD1 a expressing CD14+ monocytes to the total number of CD14+ monocytes measured.
  • the present invention in one embodiment is concerned with a method of diagnosing an inflammatory bowel disease (IBD) in a patient, wherein the CD1 a level is measured in a sample obtained from a patient suspected of suffering from an IBD.
  • IBD inflammatory bowel disease
  • a subject is diagnosed with having an IBD such as UC, if at least 3.3 % of the leukocytes in the sample obtained from the subject are expressing CD1 a. In further embodiments, at least 4 %, 5 %, 10 %, or 15 % of the leukocytes in the sample obtained from the subject are expressing CD1 a.
  • a subject is diagnosed with having an IBD such as UC, if at least 1 .7 % of the CD1 1 b+ macrophages in the sample obtained from the subject are expressing CD1 a. In further embodiments, at least 2 %, 3 %, 5 %, 10 %, or 15 % of the CD1 1 b+ macrophages in the sample obtained from the subject are expressing CD1 a.
  • a subject is diagnosed with having an IBD such as UC, if at least 77.5 % of the CD14+ monocytes in the sample obtained from the subject are expressing CD1 a. In further embodiments, at least 80 %, 85 %, 90 %, or 95 % of the CD14+ monocytes in the sample obtained from the subject are expressing CD1 a.
  • the present invention is concerned with a method of monitoring the stage and/or progress of an inflammatory bowel disease (IBD) in a patient, wherein the CD1 a level is measured in a first sample obtained from a patient suffering from an IBD at a first point of time; the CD1 a level is measured in a second sample obtained from a patient suffering from an IBD at a second point of time; and the CD1 a level measured in the first sample is compared to the CD1 a level measured in the second sample; wherein any significant increase of the CD1 a level measured in the second sample compared to the CD1 a level measured in the first sample means that the disease is progressing; and wherein a lack of a significant increase of the CD1 a level measured in the second sample compared to the CD1 a level measured in the first sample means that the disease is in a stable condition or in remission.
  • IBD inflammatory bowel disease
  • the methods of the present invention can also be used to evaluate and assess the efficacy of therapeutic strategy.
  • the second sample is taken after the patient has undergone treatment of the IBD for a period of time.
  • a lack of an increase of the CD1 a level measured in the second sample compared to the CD1 a level measured in the first sample means that the treatment is effective.
  • the sample used in any of the embodiments of the present invention is usually a sample of a body fluid and preferably is urine, serum, plasma, or blood, in particular blood.
  • the CD1 a level in the sample is determined by measuring the percentage of CD1 a expressing cells in the sample, preferably the percentage or proportion of CD1 a expressing cells in a population selected from at least one of leukocytes, monocytes, and/or macrophages, such as leukocytes and/or CD14+ monocytes and/or CD1 1 b+ macrophages.
  • the number of cells can be determined by any method and device known to the skilled person for this purpose, such as flow cytometric analysis.
  • IBD examples for IBD that can be diagnosed or monitored with the methods of the present invention are found in the group comprising Crohn's disease, ulcerative colitis (UC), collagenous colitis, lymphocytic colitis, ischemic colitis, diversion colitis, Behget's syndrome, infective colitis, and indeterminate colitis, among others.
  • the IBD to be diagnosed is Crohn's disease, or ulcerative colitis: In a most preferred embodiment the IBD is ulcerative colitis.
  • the invention is concerned with the use of CD1 a as a diagnostic marker which can be used to diagnose an inflammatory bowel disease and/or monitor progression of an inflammatory bowel disease, and/or assess treatment regime efficacy in a subject.
  • CD1 a is overexpressed in a patient suffering from an IBD, thus, CD1 a presents a novel therapeutic target molecule. Inhibition of CD1 a will decrease or slow the inflammatory processes present in IBDs. Therefore, in a further embodiment, the present invention is concerned with a CD1 a inhibitor for use in the treatment of an inflammatory bowel disease in a subject.
  • a CD1 a inhibitor for use in the present invention can be any compound or substance that inhibits or blocks binding of ligands to lipid receptor CD1 a and thereby interrupts activity elicited by binding or by a signaling cascade.
  • a CD1 a inhibitor can also be any substance that neutralizes or antagonizes the bioactivity of CD1 a.
  • Examples for a CD1 a inhibitor for use in the present invention are a CD1 a specific antibody, or a fragment or derivative thereof, or a CD1 a specific antagonist such as lipids, derivatives thereof, aptamers, aptamers or small molecules, inhibitory peptides, anti-sense RNAs, DNA and RNA sequences.
  • the present invention is concerned with a composition for use in the treatment of IBD, such as UC, comprising a CD1 a inhibitor and a pharmaceutically acceptable carrier.
  • the pharmaceutically acceptable carrier is preferably non pyrogenic.
  • the composition comprising a CD1 a inhibitor and a pharmaceutically acceptable carrier or the CD1 a inhibitor of the present invention can be administered alone or in combination with at least one other agent, such as a stabilizing compound, which can be administered in any sterile, biocompatible pharmaceutical carrier, including, but not limited to, saline, buffered saline, dextrose and water.
  • composition comprising the CD1 a inhibitor and a pharmaceutically acceptable carrier or the CD1 a inhibitor may contain pharmaceutically acceptable auxiliary substances as required.
  • Acceptable auxiliary substances preferably are nontoxic to recipients at the dosages and concentrations employed.
  • the pharmaceutical composition can contain auxiliary substances for modifying, maintaining, or preserving, for example the pH, osmolarity, viscosity, clarity, color, isotonicity, odor, sterility, stability, rate of dissolution and release, adsorption, or penetration of the composition.
  • Suitable formulation materials include, but are not limited to, amino acids, antimicrobials, antioxidants, buffers (such as borate, bicarbonate, Tris-HCI, citrates, phosphates or other organic acids), bulking agents, (such as mannitol or glycine), chelating agents (such as ethylendiamine tretraacetic acid (EDTA)), complexing agents, fillers, monosaccharides, disaccharides and other carbohydrates (such as glucose, mannose, or dextrins), proteins (such as serum albumin, gelatin or immunoglobulins), emulsifying agents, solvents (such as cetostearyl alcohol, propylene glycol, glycerin, propylene glycol, or polypropylene glycol), sugar alcohols, suspending agents, surfactant and wetting agents, stability enhancing agents, delivery vehicles, diluents, excipients and/or pharmaceutical adjuvants (see Remington's Pharmaceutical Sciences (22th Ed., Loy
  • composition comprising a CD1 a inhibitor and a pharmaceutically acceptable carrier or the CD1 a inhibitor for use can be provided for administration to a subject, wherein the subject has been diagnosed as having an increased level of CD1 a indicating the presence of an IBD, such as UC.
  • composition comprising a CD1 a inhibitor and a pharmaceutically acceptable carrier or the CD1 a inhibitor can also be provided for use in the treatment of IBD in patients for whom an increased level of CD1 a in a sample of the subject is not detectable.
  • administration of the composition comprising a CD1 a inhibitor and a pharmaceutically acceptable carrier or the CD1 a inhibitor is used to prevent a relapse, instead of to treat a relapsed IBD. This is also called maintenance therapy to prevent disease flares or disease progression in a subject.
  • composition comprising a CD1 a inhibitor and a pharmaceutically acceptable carrier, or the CD1 a inhibitor for use in the present invention can be combined with other compounds or substances that are used in the treatment of IBD.
  • compositions comprising a CD1 a inhibitor and a pharmaceutically acceptable carrier, or the CD1 a inhibitor for use in the present invention be combined with at least one further compound, substance or composition that is effective in treating IBD, such as 5- aminosalicylate (mesalazine), corticosteroids, immunomodulators, antibiotics and anti-tumor necrosis factor (TNF) agents, and anti-integrin ⁇ 4 ⁇ 7 agents, or a mixture thereof.
  • these pharmaceutical compositions can contain suitable pharmaceutically acceptable carriers comprising excipients and auxiliaries that facilitate processing of the active compound into preparations which can be used pharmaceutically.
  • composition comprising a CD1 a inhibitor and a pharmaceutically acceptable carrier, or the CD1 a inhibitor for use in the present invention can be provided in any form as is known for this type of medicament, such as in a form for oral, intramuscular, intravenous, subcutaneous, rectal, and/or intraperitoneal administration.
  • CD1 a inhibitor of the present invention can be selected from the group comprising Crohn's disease, ulcerative colitis (UC), collagenous colitis, lymphocytic colitis, ischemic colitis, diversion colitis, Behget's syndrome, infective colitis, and indeterminate colitis.
  • the IBD is Crohn's disease, or ulcerative colitis: In a most preferred embodiment the IBD is ulcerative colitis.
  • Fig. 1 shows a bwplot of CD1 a+ expressing leukocytes in healthy subjects compared to patients with ulcerative colitis and Crohn's disease.
  • A leukocytes
  • B CD1 1 b+ macrophages
  • Statistical analysis was performed using Anova followed by TukeyHSD. Fig.
  • a cut off value of 3.28 % CD1 a+ leukocytes 89 % of UC patients are true positives and 77 % of healthy subjects are true negatives.
  • a cut off value of 1 .66 % CD1 a+ macrophages 85 % of UC patients are true positives and 97 % of healthy subjects are true negatives.
  • a cut off value of 77 % CD1 a+ monocytes 78 % of UC patients are true positives and 80% of healthy subjects are true negatives.
  • Fig. 5 shows the identification of CD1 1 b+ CD1 a+ macrophages and CD14+ CD1 a+ monocytes in colon of colectomized UC patients.
  • Fig. 7 shows that CD1 a+ CD14+ monocytes are increased upon challenge in the animal model.
  • Statistical analysis was performed using Anova followed by TukeyHSD.
  • Fig. 9 shows a comparison of frequency of leukocytes in colon samples of UC and Non UC patients.
  • a two-sided t-test and a significance level 0.05 was used to compare groups.
  • Fig. 10 shows a comparison of frequencies of leukocytes in human and murine colon samples.
  • Human leukocytes were isolated from colon of UC patients undergoing colectomy and mice reconstituted with PBMC from UC patients and challenged with ethanol and subjected to flow cytometric analysis.
  • Fig. 11 shows that challenge with ethanol results in development of colitis like symptoms in NSG mice engrafted with PMBC derived from a UC patient.
  • B Macrophotographs of colons at autopsy of NSG mice engrafted with PBMC derived from UC patients, (a) control; (b) challenged with ethanol.
  • Fig. 12 shows that challenge with ethanol resulted in edema, crypt abscesses, crypt loss, fibrosis, epithelial erosions and infiltration of inflammatory cells into the submucosa and lamina intestinal in NSG mice engrafted with PMBC derived from a UC patient.
  • H&E haematoxylin and eosin
  • Fig. 13 shows that challenge with ethanol induces influx of inflammatory cells into the colon in NSG mice reconstituted with PBMC from UC patients.
  • Human leukocytes were isolated from colons of mice and subjected to flow cytometric analysis.
  • B: Frequency of subtypes of CD1 1 b+ macrophages and CD14+ monocytes in the colon of challenged NSG mice. Sample size: n 6.
  • Fig. 14 shows the result of in vitro activation of PBMC by lipids.
  • PBMC derived from UC patients and healthy subjects were incubated in the presence or absence of phosphatidyl- DL-glycerol sodium salt (PGS).
  • PPS phosphatidyl- DL-glycerol sodium salt
  • PHA Phytohemagglutinin-M
  • Flow cytometric analysis revealed statistically significant increase of activated CD4+ T-cells and CD1 a expressing CD1 1 b+ macrophages and CD14+ monocytes.
  • Lipids induced the maturation of CD1 1 b+ macrophages and CD14+ monocytes as shown by the increase in frequencies of CD1 1 b+ CD80/86+ macrophages and CD14+ CD80/86+ monocytes.
  • Fig. 15 shows the result of in vitro inhibition of PBMC by an antibody directed against CD1 a.
  • PBMC derived from UC patients and healthy subjects were incubated without (control), in the presence of phosphatidyl-DL-glycerol sodium salt (PGS) or in the presence of PGS and an anti CD1 a antibody.
  • PGS phosphatidyl-DL-glycerol sodium salt
  • Flow cytometric analysis revealed statistically significant inhibition of activation of CD4+ T-cells and inhibition of maturation of CD14+ monocytes.
  • Fig. 16 shows the effect of an antibody directed against CD1 a on the clinical score of NSG mice reconstituted with PBMC from patients with UC, challenged with 10% ethanol at day 8, and 50% ethanol at days 15 and 18 and treated with 30 ⁇ g anti CD1 a antibody by intraperitoneal injection on day 7, 14, 17.
  • control control
  • ethanol challenged and untreated control group ethanol + isotype
  • ethanol challenged and treated group ethanol + anti CD1 a
  • Fig. 17 shows the effect of anti CD1 a antibodies on macroscopic appearance (A) and colon score of colons (B) of NSG mice reconstituted with PBMC from patients with UC, challenged with 10% ethanol at day 8, and 50% ethanol at days 15 and 18 and treated with 30 ⁇ g anti CD1 a antibody by intraperitoneal injection on day 7, 14, 17.
  • Fig. 18 shows the effect of anti CD1 a antibodies on histological alterations of colons of NSG mice reconstituted with PBMC from patients with UC, challenged with 10% ethanol at day 8, and 50% ethanol at days 15 and 18 and treated with 30 ⁇ g anti CD1 a antibody by intraperitoneal injection on day 7, 14, 17.
  • A Micrographs of H&E stained sections from distal parts of the colon. Arrow indicates influx of inflammatory cells and edema.
  • Fig. 19 shows the effect of anti CD1 a antibodies on CD1 a and TSLPR expressing monocytes and TSLPR expressing CD1 1 b+ macrophages isolated from colon of NSG mice reconstituted with PBMC from patients with UC, challenged with 10% ethanol at day 8, and 50% ethanol at days 15 and 18 and treated with 30 ⁇ g anti CD1 a antibody by intraperitoneal injection on day 7, 14, 17.
  • Fig. 20 shows the effect of anti CD1 a antibodies on HGF, mTARC, TGFM and hlFNy mRNA expression in distal parts of the colon of NSG mice reconstituted with PBMC from patients with UC, challenged with 10% ethanol at day 8, and 50% ethanol at days 15 and 18 and treated with 30 ⁇ g anti CD1 a antibody (Clone OKT-6) by intraperitoneal injection on day 7, 14, 17.
  • Untreated and unchallenged control group control
  • ethanol challenged and untreated control group ethanol + isotype
  • ethanol challenged and treated group ethanol + anti CD1 a
  • Fig. 21 shows the effect of anti CD1 a antibodies on frequencies of CD14+ CD1 a+ monocytes and NK T cells isolated from spleen of NSG mice reconstituted with PBMC from patients with UC, challenged with 10% ethanol at day 8, and 50% ethanol at days 15 and 18 and treated with 30 ⁇ g anti CD1 a antibody by intraperitoneal injection on day 7, 14, 17.
  • control control
  • ethanol challenged and untreated control group ethanol + isotype
  • ethanol challenged and treated group ethanol + anti CD1 a
  • Fig. 22 shows the correlation of CD14+ CD1 a+ and central memory CD8+ T cells (A), NK T cells (B), switched (C) and unswitched B cells (D), regulatory T cells (Treg; E ) and activated CD4+ T cells (F).
  • A CD14+ CD1 a+ and central memory CD8+ T cells
  • B NK T cells
  • C switched
  • D unswitched B cells
  • D regulatory T cells
  • Reg regulatory T cells
  • F activated CD4+ T cells
  • Fig. 23 shows the correlation of CD14+ CD1 a monocytes with serum triglycerides (TAG) and cholesterol levels in mice challenged with ethanol or control mice.
  • Human leukocytes were isolated from spleen of mice and subjected to flow cytometric analysis. TAG and cholesterol levels were determined in sera of mice. For analysis a spearman correlation was performed.
  • IBD Inflammatory bowel diseases
  • IBD Inflammatory bowel diseases
  • UC Inflammatory bowel diseases
  • a combination of genetic, environmental and microbiotic factors is responsible for an uncontrolled immune response characterized and driven by the typical Th2 cytokine IL-13.
  • Most studies to characterize inflammatory processes in IBDs such as UC are based on defining roles of single cell types or interleukins, chemokines or growth factors in the human disease and to prove their roles in an animal model. This approach has tremendously improved the understanding of inflammatory mechanism underlying the disease and to develop therapeutics targeting previously identified as crucial components.
  • cells of the adaptive immunity such as plasma cells, CD4+ T-cells and subtypes thereof, CD40L+ or CD25+ activated CD4+ cells, and mucosal CD4+ cells, CD8+ T-cells and central memory CD8+ cells, cells of the innate immunity acting as mediators for the adaptive immunity such as dendritic cells, CD1 1 b+ macrophages and CD14+ monocytes and subtypes thereof, CD1 1 b+ CD1 a+, CD14+ TSLPR+, CD14+ CD1 a+, CD14+ CD163+, and cells of the innate immunity such as NKT- and NK-cells.
  • cells of the adaptive immunity such as plasma cells, CD4+ T-cells and subtypes thereof, CD40L+ or CD25+ activated CD4+ cells, and mucosal CD4+ cells, CD8+ T-cells and central memory CD8+ cells, cells of the innate immunity acting as mediators for the adaptive immunity such as dendritic cells, CD1
  • HGF and ⁇ which also have been described as elevated, displayed a significant increase in serum levels. With the exception of NKT-cells age did not affect these cell types.
  • the mesalazine treated group was characterized by a decline in frequencies of CD1 a+ CD16+ patrolling monocytes, dendritic cells and the subtype of NK T-cells which are thought to have an inhibitory function.
  • the group treated with immuno-suppressors was similar to the glucocorticoid treated group. As some of these patients were irresponsive to treatment, the observed effect might be due to the inflammatory condition and not the response to treatment.
  • CD1 a expressing leukocytes CD1 a expressing CD1 1 b+ macrophages and CD1 a expressing CD14+ monocytes, especially.
  • CD1a expressing leukocytes CD1 a expressing leukocytes
  • CD1 a expressing leukocytes and CD1 a expressing CD1 1 b+ macrophages and CD1 a expressing CD14+ monocytes specifically, emerged as cell types significantly associated with UC. These cell types differentiated between UC and Non UC donors with high discriminatory potential as shown by an AUC value of 0.81 , 0.87 and 0.76, respectively (see Fig. 1 - 4).
  • the present application is the first disclosure showing an association of CD1 a expressing leukocytes, and specifically CD1 a expressing monocytes and macrophages with the gut or with IDBs such as UC.
  • TNFoblockers Treatment with TNFoblockers induced a condition signified by CD14+ TSLPR+ which correlated positively with innate NKT-cells and ⁇ and periostin, indicating that this condition represent a 'remodeling' inflammatory condition characterized by remodeling of the colon architecture (see Table 1 ). The fact that most patients in this group responded to treatment and were considered to be in remission further supported the idea.
  • the second inflammatory condition could be more characteristic of an acute inflammation signified by TSLPR+ CD1 1 b+ macrophages that correlated positively with the SCCAI-Score, cells of the adaptive immunity, HGF and TARC.
  • TSLPR+ CD1 1 b+ and TSLPR+ CD14+ monocytes correlated negatively with each other, suggesting that treatment with TNFoblockers clearly suppresses the acute condition while promoting the remodeling condition.
  • Identification of potentially significant markers in the blood of UC patients is based on the assumption that the influx of inflammatory cells into the colonic mucosa is not a one way route. Due to disrupted endothelial layers or an active transport mechanism, cell trafficking occurs to and from both compartments. If this assumption is correct one has to find the cells identified as relevant in the blood also in the colon of UC patient.
  • CD1 a+ expressing monocytes were significantly elevated in UC patients as compared to normal tissue derived from cancer patients undergoing colectomy (see Fig. 5 and 6).
  • lipids on CD4+ T-cells and CD1 a expressing CD14+ monocytes and CD1 a expressing CD1 1 b+ macrophages was assessed in an in vitro assay.
  • PBMC derived from UC patients and healthy subjects were incubated in the presence or absence of phosphatidyl-DL-glycerol sodium salt (PGS).
  • PPS phosphatidyl-DL-glycerol sodium salt
  • PHA Phytohemagglutinin-M
  • Flow cytometric analysis revealed statistically significant increase of activated CD4+ T-cells and CD1 a expressing CD1 1 b+ macrophages and CD14+ monocytes.
  • mice were reconstituted with PBMC derived from UC patients, and is also termed NSG-UC model herein.
  • mice Upon challenge with ethanol, mice developed symptoms similar to conventional mouse models such as weight loss, diarrhea, influx of lymphocytes into the mucosa and altered colon architecture characterized by edema, crypt loss, crypt abscesses, fibrosis and epithelial hyperplasia.
  • symptoms similar to conventional mouse models such as weight loss, diarrhea, influx of lymphocytes into the mucosa and altered colon architecture characterized by edema, crypt loss, crypt abscesses, fibrosis and epithelial hyperplasia.
  • edema edema
  • crypt loss crypt abscesses
  • fibrosis fibrosis and epithelial hyperplasia.
  • mice reflect a chronic disease sometimes ongoing for decades and which do not bear the entire inflammatory cell repertoire, do not host the same microbiota and provide different defense mechanism with regard to defensin
  • CD1 a expressing CD1 1 b+ macrophages and CD14+ monocytes are highly correlated with the clinical activity score in UC patients and that both cellular populations were found to be elevated in the colon of UC patients. Moreover, both populations had been identified as inflammatory markers in the mouse model, which is based on NSG mice reconstituted with
  • mice were reconstituted with PBMC derived from three different UC patients, two of which were treated with adalimumab and exhibited moderate simple clinical colitis activity index
  • SCCAI SCCAI
  • the challenged control group was treated with 30 ⁇ g isotype control antibodies. Symptoms of colitis were induced by intrarectal challenge of 10% ethanol on day eight, followed by
  • mice were monitored throughout the experiment and symptoms were classified according to a clinical score as described in Materials and Methods. As shown in Fig. 16 mice developed a clinical activity score upon challenge with ethanol which was slightly decreased in the study group (for complete data set see Table 11 ).
  • mice were sacrificed on day 24, the colon was removed, subjected to visual inspection and classified according to a colon score.
  • Fig. 17 A colons of mice in the challenged group revealed lack of pellets, soft pellets and dilatation whereas the appearance of the colon of the study group was similar to the unchallenged control group.
  • Colons were classified according to a colon score as depicted in Fig. 17 B and described in material and methods. The analysis of the colon score supported the results from the clinical score indicating a beneficial effect of anti CD1 a antibodies. The score in the study group reached levels observed in the unchallenged control group.
  • Fig. 18 A the challenged control group revealed edema and influx of inflammatory cells whereas the unchallenged control and the treated study group had a normal appearance. This was also reflected in the histological score (see Fig. 18 B).
  • HGF, IFNy, ⁇ and TARC have been identified as markers associated with the acute or remodeling condition of inflammation in NSG-UC mice or UC patients, mRNA expression levels were determined.
  • CD14+ CD1 a+ monocytes correlated positively with serum levels of triglycerides and cholesterol, indicating that CD14+ CD1 a+ monocytes act as sensors to metabolic switches induced by inflammation. These results support the idea of a pro-inflammatory phenotype of CD1 a+ CD14+ monocytes. In addition, CD14+ CD1 a+ monocytes may act as sensors of inflammation. Mice benefitted from treatment with anti CD1 a antibodies and thus confirm CD1 a as a novel promising target to address IBD.
  • CD1 a has been identified an validated as a therapeutic target
  • LPMC lamina limbal mononuclear cells
  • the tissue was predigested for 4 x 15 minutes in 15 ml predigestion solution containing 1 x HBSS (Thermo Scientific, Darmstadt, Germany), 5mM EDTA, 5% FCS, 100 U/ml Pencillin-Streptomycin (Sigma-Aldrich Co., St. Louise USA) in an orbital shaker with slow rotation (40g) at 37° Celsius.
  • HBSS Thermo Scientific, Darmstadt, Germany
  • 5mM EDTA 5% FCS
  • Pencillin-Streptomycin Sigma-Aldrich Co., St. Louise USA
  • Isolated LPMC were collected by centrifugation with 177 g for 10 minutes and resuspended for FACS analysis. Cell suspensions were filtrated one more time using a 35 ⁇ cell strainer for further purification before labelling the cells for flow cytometry analysis.
  • spleens were minced and cells filtrated through a 70 ⁇ cell strainer followed by centrifugation at 1400 g for 5 minutes and resuspended in FACS buffer (1 x PBS, 2mM EDTA, 2 %FCS). For further purification cell suspensions were filtrated using a 35 ⁇ cell strainer and then labelled for flow cytometry analysis.
  • Isolated human leukocytes were centrifuged at 1400 g for 5 minutes and resuspended in FACS buffer. Cell suspensions were filtrated one more time using a 35 ⁇ cell strainer for further purification before labelling the cells for flow cytometry analysis. Cell populations were defined as described in Table 5. Flow cytometric analysis
  • CD294 (CRTH2) APC BM 16
  • CD1 a biotin
  • 4x10 6 PBMC were incubated in 1 .5 ml RPMI, 10 % FCS; 1 % sodium pyruvate, 1 % penicillin/streptomycin; 1 % glutamine (Sigma, Deisenhofen, Germany) for 48h in 24-well flat-bottom cell culture plates (Greiner Bio-One) with 50 ⁇ g PGS or 10 ⁇ g PHA (Sigma, Deisenhofen, Germany) in a humidified incubator at 37°C with 5% C0 2 .
  • NOD.cg-Prkdc SCID N2rg tm1Wjl /Szj mice (abbreviated as NOD IL-2RY nu ") were obtained from Charles River Laboratories (Sulzfeld, Germany). Mice were kept under specific pathogen free conditions in individually ventilated cages. The facility is controlled according to the Federation of Laboratory Animal Science Association (FELASA) guidelines. Following engraftment (day 1 ) mice were presensitized by rectal application of 150 ⁇ of 10 % Ethanol on day 8 using a 1 mm cat catheter (Henry Schein, Hamburg, Germany). The catheter was lubricated with Xylocain ⁇ Gel 2% (AstraZeneca, Wedel).
  • Rectal application was performed under general anesthesia using 4% Isofluran. Post application mice were kept at an angle of 30° to avoid ethanol dripping. On day 15 and 18 mice were challenged by rectal application of 50% ethanol following the protocol of day 8. Mice were sacrificed on day 21 .
  • the anti-CD1 a antibodies or IgG isotype control (30 ⁇ g in 200 ⁇ PBS) (BioXcell, West Riverside NH, USA, Clone OKT-6) were applied on day 7 and 14.
  • Infliximab (6 mg/kg (Remicade ⁇ , Janssen, Netherlands) was applied on day 7, 14 and 17.
  • An isotype antibody human lgG1 , kindly provided by, MorphoSys AG
  • All treatments were applied intraperitoneally.
  • Inflammation was scored as follows: infiltration of a few inflammatory cells into the Lamina propria (1 ), major infiltration of inflammatory cells into the Lamina propria (2), confluent infiltration of inflammatory cells into the Lamina propria (3), infiltration of inflammatory cells including tunica muscularis (4).
  • Fibrosis was scored as follows: focal fibrosis (1 ), multifocal fibrosis and crypt atrophy (2). The presence of oedema, hyperaemia and crypt abscess was scored with 1 additional point in each case. The scores for each criteria were added into a total score ranging from 0 to 12. Images were taken with a Zeiss AxioVert 40 CFL camera. Figures show representative longitudinal sections in original magnification. In Adobe Photoshop CS6 a tonal correction was used in order to enhance contrasts within the pictures. Colon Score
  • the colon score was assessed at necropsy according to the following scoring parameters: Consistency of pellets: formed (0), pasty (1 ), fluid (2). Length: ⁇ 10 cm (0), 8-10 cm (1 ), ⁇ 8 cm (2). Hyperemia: no (0), yes (2). Dilatation: no (0), low (1 ), major (2). Necrosis: no (0), yes (2). RNA analysis
  • RNA extraction Approximately 1 cm from distal parts of the colon was disrupted and homogenized using a TissueLyser LT (Qiagen, Hilden, Germany) followed by total RNA extraction according to the manufacturer's instruction using RNeasy Plus Universal Mini Kit (Qiagen, Hilden, Germany) and Chloroform (Sigma-Aldrich, St. Lousi, MO, USA). No further treatment with DNase was needed since gDNA Eliminator Solution was included in the kit.
  • RNA samples were diluted with RNase free water.
  • RNA and cDNA purity was assessed using a Nanodrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA).
  • Quantitative real-time PCR was performed according to the TaqMan Fast Advanced Master Mix protocol (Thermo Fisher Scientific, Waltham, MA, USA) using the Applied Biosystems StepOnePlus real-time PCR system (Thermo Fisher Scientific, Waltham, MA, USA).
  • Single Tube TaqMan Gene Expression Assays included primers for the housekeeping genes GAPDH (Mm99999915_g1 ) and GUSB (Mm00446953_m1 ) as well as TGF3 (Mm01 178820_m1 ), HGF (Hs04329698_m1 ), CCL17 (Mm01244826_g1 ) and IFN ⁇ (Hs00989291_m1 ). Analysis was performed using StepOnePlusTM Software v2.3.
  • the mean cycle threshold (CT) value was calculated for the housekeeping genes. Relative expression values for the analyzed genes were calculated as the difference between the mean cycle threshold (CT) of the housekeeping genes and the analyzed gene (delta CT) and depicted as the logarithmic value. Detection of cholesterol and triglyceride serum levels
  • TAG triglycerides
  • PBMC Peripheral blood mononuclear cells
  • samples were collected as described in Example 1 and samples were subjected to flow cytometric analysis to determine the frequencies using surface markers of B-cells, T-cells, macrophages, monocytes, dendritic cells, NKT and NK cells and subtypes thereof (Table 5).
  • Table 5 Cellular markers used in phenotyping of UC patients and used to define human immune cells in colon of NSG mice reconstituted with PBMC from UC patient
  • CD4+ CD44+ CD62L- Antigen experienced CD4+ T- cell
  • CD4+ CD25+ activated CD4+ T-, regulatory T cells, Th2 cells
  • CD14+ CCR2+ MC tissue penetrating, inflammatory
  • CD14+ CD163+ M2 MC CD14+ CD163+ M2 MC, scavenging cells
  • CD14+ CD163+ CD206+ M2 MC CD14+ CD163+ CD206+ M2 MC, scavenging cells
  • CD16+ MC non classical, patrolling
  • CD16+ CD80/86+ MC non classical, patrolling, mature
  • CD16+ TSLPR+ MC non classical, patrolling, TSLPR expressing
  • CD16+ CD163+ MC non classical, patrolling, M2?
  • CD1 1 b+ HLADR+ cDC1 presenting
  • CD11 b+ CD1a+ cDC1 CD1 a expressing
  • CD4+ T-cells As shown in Table 6 below, a significant decline in frequencies was found for plasma cells, CD4+ T-cells, antigen experienced lymph homing CD4+ T-cells and activated CD25+ CD4+ T-cells, central memory CD8+ T-cells, CD1 1 b+ macrophages, CD1 1 c+ dendritic cells, CD16+ resident monocytes, KIR expressing NK T-cells in the patient group as compared to the Non UC group.
  • CD40L+ activated CD4+ T-cells, and mucosal regulatory CD4+ and CD8+ T-cells, CD1 a expressing CD1 1 b+ macrophages, TSLPR-, CD1 a- and CD163 expressing CD14+ monocytes, KIR- and CD94 expressing NK cells, NKT cells and inhibitory CD94 expressing NKT cells were observed.
  • serum levels of HGF and ⁇ were increased in the UC group.
  • patients treated with TNFoblockers displayed lower frequencies of cells of the adaptive immune system to include plasma cells, TSLPR+ and mature CD1 1 b+ macrophages and dendritic cells along with a decline in HGF and TARC whereas frequencies of TSLPR+ CD14+ monocytes and serum levels of ⁇ increased. Opposing effects were observed in the patient group treated with glucocorticoids. In this group, cells of the adaptive immune system were not suppressed. Frequencies of B cells, switched memory B-cells, plasma cells and TSLPR expressing CD1 1 b+ macrophages displayed increased frequencies along with and HGF and TARC serum levels.
  • CD1 a+ leukocytes and CD1 1 b+ CD1 a+ and CD14+ CD1 a+ leukocytes, specifically, seemed to be cell populations discriminating between Non UC and UC and seemed to be unaffected by therapeutic interventions, a ROC analysis was performed to evaluate the quality of CD1 a as a marker.
  • the analysis revealed a high specificity and sensitivity for CD1 a expressing leukocytes, and CD1 a expressing CD1 1 b+ macrophages, and CD1 a expressing CD14+ monocytes (see Fig. 1 - 4).
  • Table 6 shows that immunological profiling leads to identification of specific cellular markers for UC and to markers signifying therapeutic responses.
  • PBMC and serum from UC patients and Non UC patients were isolated and subjected to flow cytometric analysis.
  • the table displays differences of mean values of frequency of immune cells in the blood and serum levels of factors.
  • UC patients versus Non UC donors UC patients treated with TNFa blocker, glucocorticoid, mesalazine and immunosuppressive compared to all other patients.
  • the profiling and cluster analysis suggested that mainly two immunological profiles prevailed in the UC patient group a correlation analysis was performed.
  • the cluster analysis included the clinical activity score (SCCAI- Score) and HGF, TARC, ⁇ and periostin).
  • Table 7 shows that correlation analysis reveals two distinct inflammatory conditions.
  • the SCCAI-Score positively correlated with switched memory B-, plasma-, CD25+ activated and experienced CD44+ CD4+ T-cells, TSLPR+ macrophages and mature CD16+ and M2 resident monocytes, suppressive NK cells, HGF and TARC whereas it negatively correlated with unswitched memory B-, mucosal memory CD8+ cells, mature CD14+ monocytes, TSLPR expressing CD16+ monocytes, and periostin, suggesting that the adaptive immune cells along with TSLPR expressing macrophages and mature CD16+ monocytes play an important role during the active form of the disease and that HGF and TARC are hallmarks of this condition.
  • HGF, TARC, ⁇ and periostin clearly fell into two groups: In most cases where HGF and TARC displayed positive correlation coefficients, ⁇ and periostin displayed negative ones and vice versa. HGF and TARC were positively correlated with each other and both factors were negatively correlated with ⁇ and periostin. Likewise, ⁇ and periostin were positively correlated with each other. HGF and TARC displayed a similar correlation pattern as the SCCAI-Score: Both positively correlated with switched memory B- and plasma cells, CD25+ activated CD4+ T-cells, TSLPR expressing 1 1 b+ macrophages, mature CD16+ monocytes and suppressing NK cells.
  • TARC and HGF were negatively correlated with unswitched memory B-cells, mature CD14+ monocytes, and TSLPR expressing CD16+ monocytes.
  • Table 8 shows that correlation analysis of CD1 a expressing monocytes
  • colon samples of UC patients undergoing colectomy were analyzed with regard to the presence of leukocytes and compared to unaffected areas of colon samples from cancer patients.
  • Leukocytes were isolated as described above and subjected to the same flow cytometric analysis as the leukocytes from blood samples. Although frequencies of all cell types were increased in UC samples as compared to Non UC samples, a significant increase could only be determined for CD14+ monocytes when CD1 1 b+ macrophages, CD1 1 c+ dendritic cells, CD14+ monocytes and CD4+ and CD8+ cells were analyzed (Fig. 9A).
  • NSG mice developed similar symptoms to wildtype mice upon challenge with oxazolone. Unexpectedly, however, similar albeit milder effects were observed in the control mice challenged with the carrier ethanol. These effects were much stronger when PBMC derived from a relapsing UC patient were used for reconstitution.
  • response to challenge was analyzed with regard to the development of a clinical and histological score, macroscopic changes of the colon, the frequency of leukocytes isolated from spleen and colon and cytokine and growth factor expression in the colon.
  • SCCAI simple clinical colitis activity indices
  • Donor 3 differed also from the other donors with regard to CD1 a+ monocytes where all other donors displayed elevated levels as compared to Non UC subjects. The highest variability between donors was observed in CD1 a expressing Cd1 1 b+ macrophages. Seven days post reconstitution the mice were divided into two groups: one was left unchallenged and the other was challenged by rectal application of ethanol. Each group contained four animals. On day seven mice were presensitized by rectal application of 10% ethanol followed by rectal application of 50% ethanol at day 15 and 18. The onset of the disease was monitored by body weight and visual inspection of stools and mice. Symptoms were classified according to a clinical activity score as described in Materials and Methods.
  • mice Upon challenge with ethanol mice stools became soft or liquid, the animals lost weight and the activity was reduced. Control animals displayed hardly any symptoms. Symptoms peaked at day 16 and challenged animals recovered two days post challenge. All animals survived. The development of symptoms was reflected in the clinical score of the challenged group which was significantly higher as compared to the unchallenged group (Fig. 11 A). Control animals remained unaffected. A high variability between different donors was observed. On day 21 mice were sacrificed, the colon was visually inspected, colon samples from the distal part of the colon were collected for histological and mRNA expression analysis, and leukocytes were isolated from spleen and colon. Visual inspection of the colon corroborated the observed clinical scores. As shown in Fig.
  • Fig. 12A increased basophilia at the base of the crypts indicated epithelial proliferation.
  • Fig. 12B The morphological changes were classified according to a histological score as described in Material and Methods. As observed with the clinical score the degree of pathological manifestation varied between experiments and indicated a donor to donor variability. As shown in Fig 12B the histological score in the challenged group was significantly higher as compared to the control.
  • human leukocytes isolated from murine spleens were subjected to flow cytometric analysis.
  • CD14+ monocytes and CD1 1 b+ macrophages were the most abundant populations and exceeded CD4+ and CD8+ T-cells.
  • the variability of frequencies was high and reflected the donor to donor variability.
  • the most abundant subsets of monocytes and macrophages were CD1 a expressing, TSLPR expressing, and mature macrophage and monocytes (see Fig. 13B).
  • Example 6 Comparison of UC and the NSG mouse model
  • NSG NOD-scid IL2R ⁇ ⁇ "
  • PBMS PBMS derived from UC patients developed colitis like symptoms upon challenge with ethanol.
  • this model is of the human disease presence and frequencies of human leukocytes were compared in colon samples from UC patients undergoing colectomy from and mouse colon challenged with ethanol.
  • mice were reconstituted with 4 x 10 6 PBMC from six different donors as described above. Following reconstitution on day 1 mice were presensitized by rectal application of 10% ethanol on day 8 followed by challenge with 50% ethanol on day 15 and 18. On day 21 mice were sacrificed and human leukocytes were isolated from pooled colons as described in Example 1 and subjected to flow cytometric analysis.
  • Example 7 In vitro activation and stimulation of CD4+ T-cells and CD1 a+ CD14+ monocytes and CD1 a + CD11 b+ macrophages
  • Example 8 In vitro inhibition of activation of CD4+ T-cells and maturation of CD14+ monocytes and CD11 b+ macrophages.
  • Example 9 In vivo efficacy of an anti CD1 a antibody in the NSG-UC mouse model
  • mice were reconstituted with PBMC derived from three different UC patients, two of which were treated with adalimumab and exhibited moderate simple clinical colitis activity index (SCCAI) of three and five, respectively, and one patient was treated with vedolizumab and exhibited a high SCCAI score of nine.
  • SCCAI simple clinical colitis activity index
  • the challenged control group was treated with 30 ⁇ g of isotype control antibodies. Symptoms of colitis were induced by intrarectal challenge of 10% ethanol on day 8, followed by 50% ethanol on days 15 and 18. Mice were monitored throughout the experiment and symptoms were classified according to a clinical score. As shown in Fig.
  • mice developed a clinical activity score upon challenge with ethanol which was slightly decreased in the study group (for complete data set see Table 11 ).
  • Mice were sacrificed on day 21 , the colon was removed, subjected to visual inspection and classified according to a colon score.
  • Fig. 17A colons of mice in the challenged group revealed lack of pellets, soft pellets and dilatation whereas the appearance of the colon of the study group was similar to the unchallenged control group.
  • Colons were classified according to a colon score as depicted in Fig. 17B and described in Materials and Methods. The analysis of the colon score supported the results from the clinical score indicating a beneficial effect of anti CD1 a antibodies.
  • the score in the study group reached levels observed in the unchallenged control group.
  • Fig. 18A To further analyze the effect of anti CD1 a antibodies distal parts of the colon were subjected to histological analysis. As shown in Fig. 18A the challenged control group revealed edema and influx of inflammatory cells whereas the unchallenged control and the treated study group exhibited reduced influx of inflammatory cells and less edema. Some of them, however, appeared to have augmented fibrosis (data not shown). This was also reflected in the histological score (Fig. 18B).
  • HGF As HGF, IFNy, ⁇ and TARC had been identified as markers associated with the acute or remodeling condition of inflammation in NSG-UC mice or UC patients, mRNA expression levels were determined. As shown in Fig. 20 levels of mTARC and ⁇ increased in response to treatment suggesting that inflammation was not completely suppressed but that anti CD1 a antibodies favored a remodeling condition. In contrast, treatment resulted in decreased HGF mRNA indicating that the acute inflammatory condition suppressed. Treatment had no effect on IFNy mRNA levels.
  • CD1 1 b+ macrophages correlated positively with activated CD4+ T (CD69+) cells suggesting an entirely different mode of action of these two cell types.
  • CD14+CD1 a+ monocytes correlated positively with TAG and cholesterol (Fig. 23), suggesting that CD14+ CD1 a+ monocytes sense inflammation.
  • CD1 a-autoreactive T cells are a normal component of the human alphabeta T cell repertoire. Nat Immunol 11 , 1 102-1 109.
  • CD1 a- autoreactive T cells recognize natural skin oils that function as headless antigens. Nat Immunol 15, 177-185. Zadeh-Khorasani, M., Nolte, T., Mueller, T. D., Pechlivanis, M., Rueff, F., Wollenberg, A., Fricker, G., Wolf, E., Siebeck, M. and Gropp, R. (2013). NOD-scid IL2R gammanull mice engrafted with human peripheral blood mononuclear cells as a model to test therapeutics targeting human signaling pathways. J TransI Med 11 , 4.

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Abstract

La présente invention concerne l'identification d'une nouvelle cible thérapeutique et d'un marqueur diagnostique de maladies intestinales inflammatoires chroniques, telles que la rectocolite hémorragique (UC) ou la maladie de Crohn (CD). Le nouveau marqueur diagnostique peut être utilisé pour surveiller l'efficacité du traitement et/ou le stade de la maladie. Ce marqueur est le récepteur de lipide CD1a, qui est une glycoprotéine transmembranaire présente sur des cellules présentatrices d'antigène (CPA), et médie la présentation d'antigènes principalement lipidiques et glycolipidiques à des lymphocytes T. CD1a peut également être utilisé en tant que molécule cible thérapeutique, étant donné que le blocage de ce récepteur conduira à une diminution du processus inflammatoire médié par les lymphocytes T activés par la présentation de l'antigène par les CPA. Ce blocage peut, par exemple, être obtenu par des anticorps anti-CD1a spécifiques, ou par l'utilisation de lipides inhibiteurs ou de petites molécules inhibitrices.
PCT/EP2017/064597 2016-06-15 2017-06-14 Cd1a en tant que molécule cible thérapeutique et marqueur diagnostique pour maladies intestinales inflammatoires WO2017216253A1 (fr)

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Title
"Thermo Fisher Scientific", WALTHAM, MA, USA
BOURGEOIS, E. A.; SUBRAMANIAM, S.; CHENG, T. Y.; DE JONG, A.; LAYRE, E.; LY, D.; SALIMI, M.; LEGASPI, A.; MODLIN, R. L.; SALIO, M: "Bee venom processes human skin lipids for presentation by CD1 a", J EXP MED, vol. 212, 2015, pages 149 - 163
DE JONG, A.; CHENG, T. Y.; HUANG, S., GRAS, S.; BIRKINSHAW, R. W.; KASMAR, A. G.; VAN RHIJN, I.; PENA-CRUZ, V.; RUAN, D. T.; ALTMA: "CD1 a-autoreactive T cells recognize natural skin oils that function as headless antigens", NAT IMMUNOL, vol. 15, 2014, pages 177 - 185
DE JONG, A.; PENA-CRUZ, V.; CHENG, T. Y.; CLARK, R. A.; VAN RHIJN, I; MOODY, D. B: "CD1a-autoreactive T cells are a normal component of the human alphabeta T cell repertoire", NAT IMMUNOL, vol. 11, 2010, pages 1102 - 1109, XP055328951, DOI: doi:10.1038/ni.1956
LOYD V ALLEN, JR: "Remington's Pharmaceutical Sciences", 2012, MACK PUBLISHING COMPANY
MICHAEL J. PAGE ET AL: "Cd1d-Restricted Cellular Lysis by Peripheral Blood Lymphocytes: Relevance to the Inflammatory Bowel Diseases", JOURNAL OF SURGICAL RESEARCH., vol. 92, no. 2, 1 August 2000 (2000-08-01), US, pages 214 - 221, XP055300975, ISSN: 0022-4804, DOI: 10.1006/jsre.2000.5940 *
ZADEH-KHORASANI, M.; NOLTE, T.; MUELLER, T. D.; PECHLIVANIS, M.; RUEFF, F.; WOLLENBERG, A.; FRICKER, G.; WOLF, E.; SIEBECK, M; GRO: "NOD-scid IL2R gammanull mice engrafted with human peripheral blood mononuclear cells as a model to test therapeutics targeting human signaling pathways", J TRANSL MED, vol. 11, 2013, pages 4

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