WO2013155010A1 - Biomarqueurs non invasifs pour l'œsophagite éosinophile - Google Patents

Biomarqueurs non invasifs pour l'œsophagite éosinophile Download PDF

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WO2013155010A1
WO2013155010A1 PCT/US2013/035662 US2013035662W WO2013155010A1 WO 2013155010 A1 WO2013155010 A1 WO 2013155010A1 US 2013035662 W US2013035662 W US 2013035662W WO 2013155010 A1 WO2013155010 A1 WO 2013155010A1
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eoe
biomarkers associated
ige
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Anil Mishra
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Children's Hospital Medical Center
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/158—Expression markers

Definitions

  • the invention disclosed herein generally relates to diagnosis, treatment, and/or management of eosinophilic esophagitis and/or diseases, disorders, and/or conditions arising therefrom and/or related thereto.
  • Eosinophilic esophagitis is a painful and sometimes devastating inflammatory disease of the esophagus. EoE symptoms mimic gastroesophageal reflux disease (GERD) and include, but are not limited to, swallowing problems, vomiting, food refusal, food intolerance in infants, abdominal pain, dysphagia, and food impactions in adolescents and adults (see, e.g., Orenstein, S. et al. Am. J. Gastroenterol. 95: 1422-30 (2000); Walsh, S. et al. Am. J. Surg. Pathol. 23:390-6 (1999); Liacouras, C. and Ruchelli, E. Curr.
  • EoE is considered to be a global health mini-epidemic on multiple continents, as has been frequently noted (see, e.g., Orenstein, S. et al. Am. J. Gastroenterol. 95: 1422-30 (2000); Croese, J. et al. Gastrointest. Endosc. 58:516-22 (2003); Attwood, S. et al. Dig. Dis. Sci. 38: 109-16 (1993)); Cury, E. et al. J. Pediatr. Surg. 39:e4-7 (2004); Cantu, P. et al. Dig. Liver Dis. 37: 129-34 (2005); Fujiwara, H. et al. Ann.
  • Embodiments of the invention encompass methods of treating a patient with eosinophilic esophagitis (EoE), including: obtaining a sample from a patient, analyzing the sample to determine a level of one or more biomarkers associated with EoE, determining whether the level of the one or more biomarkers associated with EoE is up-regulated or down-regulated relative to a level of the one or more biomarkers measured in a normal individual, wherein the presence of an elevated or reduced level of one or more biomarkers associated with EoE can result in the patient being diagnosed with EoE, and treating the patient with an appropriate therapeutic strategy based upon the diagnosis.
  • EoE eosinophilic esophagitis
  • the one or more biomarkers associated with EoE can include, for example, one or more cytokines, IL- 15 -responsive iNKT, T, and B cell receptors, chemokines, mediators, and IgE receptors, or the like.
  • the one or more cytokines can include, for example, IL-5, IL-13, IL-15, INFy, and/or TGF- ⁇ , or the like.
  • the one or more IL-15 responsive iNKT, T, and B cell receptors can include, for example, IL-15Ra, ⁇ , ⁇ , CD Id, Va24, ⁇ 1, and/or Jal8, or the like.
  • the one or more chemokines can include, for example, CXCR6 and/or CXCL16, or the like.
  • the one or more mediators can include, for example, IgE, or the like.
  • the one or more IgE receptors can include, for example, FCeRI and/or FCeRII, or the like.
  • the one or more biomarkers associated with EoE can include, for example, IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CD Id, Va24, ⁇ 1, Jal8, CXCR6, CXCL16, IgE, FCeRI, and FCeRII, or the like.
  • the one or more biomarkers associated with EoE can include, for example, IL-15 and/or CXCL16, or the like.
  • the one or more biomarkers associated with EoE can include, for example, IL-15, or the like.
  • the presence of an elevated level of, for example, ⁇ , ⁇ , Va24, CXCR6, FCeRI, and/or FCeRII, or the like, can result in the patient being diagnosed with EoE.
  • the presence of a reduced level of, for example, ⁇ 1 and/or Jal8, or the like can result in the patient being diagnosed with EoE.
  • the mRNA level of the one or more biomarkers associated with EoE is determined.
  • the protein level of the one or more biomarkers associated with EoE is determined.
  • the determination of whether the level(s) of the one or more biomarkers associated with EoE are elevated or reduced relative to a level of the one or more biomarkers measured in a normal individual can be combined with a determination of a level(s) of one or more additional biomarkers associated with EoE.
  • the one or more additional biomarkers associated with EoE can include an mRNA biomarker.
  • the one or more additional biomarkers associated with EoE can include eotaxin-3.
  • the sample can be an esophageal tissue sample. In some embodiments, the sample can be a plasma or serum sample.
  • the appropriate therapeutic strategy for a patient diagnosed with EoE can include, for example, allergen removal, steroid treatment, dietary management, proton pump inhibitor (PPI) therapy, administration of one or more topical glucocorticoid, administration of one or more humanized antibody against one or more relevant cytokines and/or mediators, administration of one or more small molecule inhibitors of an eosinophil and/or allergic disease activation pathway, administration of one or more small molecule inhibitors capable of modulating levels of one or more biomarkers associated with EoE, and/or any combination thereof, or the like.
  • PPI proton pump inhibitor
  • the topical glucocorticoid can include, for example, fluticasone, budesonide, and/or ciclesonide, or the like.
  • the humanized antibody against one or more relevant cytokines and/or mediators can include, for example, an antibody targeting one or more biomarkers associated with EoE, or the like.
  • the one or more biomarkers associated with EoE can include, for example, IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ 1, Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII, or the like.
  • the humanized antibody against one or more relevant cytokines and/or mediators can be, for example,e anti-IL-15, anti-IgE, anti-CDld, anti-Va24Jal8, anti- CXCL16, and anti-IL-15Ra, or the like.
  • Embodiments of the invention also encompass methods of treating a patient with eosinophilic esophagitis (EoE), including: obtaining a sample from a patient, analyzing the sample to determine a level of one or more biomarkers associated with EoE, determining whether the level of the one or more biomarkers associated with EoE is up- regulated or down-regulated relative to a level of the one or more biomarkers measured in a normal individual, wherein the presence of an elevated or reduced level of one or more biomarkers associated with EoE results in the patient being diagnosed with EoE, and treating the patient with an appropriate therapeutic strategy based upon the diagnosis, further including a determination of eosinophilic esophagitis or GERD.
  • EoE eosinophilic esophagitis
  • the presence of reduced level of, for example, Vail and Jal8, or the like, in combination with a non-elevated or non-reduced level of one or more additional biomarkers associated with eosinophilic esophagitis can result in the patient being diagnosed with GERD.
  • Embodiments of the invention also encompass methods of diagnosing a patient with eosinophilic esophagitis (EoE), including: obtaining a sample from a patient, analyzing the sample to determine a level of one or more biomarkers associated with EoE, and determining whether the level of the one or more biomarkers are up-regulated or down- regulated relative to a level of the one or more biomarkers measured in a normal individual, wherein the presence of an elevated or reduced level of one or more biomarkers associated with EoE can result in the patient being diagnosed with EoE.
  • EoE eosinophilic esophagitis
  • Embodiments of the invention also encompass diagnostic kits, tests, or arrays, including materials for quantification of at least two analytes, wherein the at least two analytes can be, for example, biomarkers associated with eosinophilic esophagitis (EoE).
  • the at least two analytes can include, for example, one or more cytokines, IL-15 responsive iNKT, T, and B cell receptors, chemokines, mediators, and IgE receptors, or the like.
  • the one or more cytokines can include, for example, IL-5, IL-13, IL-15, INFy, and/or TGF- ⁇ , or the like.
  • the one or more IL-15 responsive iNKT, T, and B cell receptors can include, for example, IL-15Ra, ⁇ , ⁇ , CD Id, Va24, ⁇ ⁇ , and/or Jal8, or the like.
  • the one or more chemokines can include, for example, CXCR6 and/or CXCL16, or the like.
  • the one or more mediators can include, for example, IgE, or the like.
  • the one or more IgE receptors can include, for example, FCeRI and/or FCeRII, or the like.
  • the diagnostic kit, test, or array can include a gene chip.
  • the gene chip can include a low density array.
  • the diagnostic kit, test, or array can include a surface with a DNA array.
  • Figures 1A-D depict representative photomicrographs for the study of IL- 15 induction in human EoE.
  • Figures 1A-B demonstrate that there were no IL-15 -positive cells in the esophageal biopsies of non-EoE patients.
  • Figure 1A provides the pictomicrograph under the original magnification of lOx;
  • Figure IB provides the pictomicrograph under 400x magnification.
  • Figures 1C-D demonstrate that a number of IL- 15-positive cells were detected in EoE patient biopsies.
  • Figure 1C provides the pictomicrograph under the original magnification of lOx;
  • Figure ID provides the pictomicrograph under 400x magnification.
  • IL-15 -positive cells are indicated with arrows.
  • Figures 2A-D depict patterns resulting from staining a representative EoE patient esophageal biopsy with various media.
  • Figure 2A shows the cell nuclei through 4',6- diamidino-2-phenylindole (DAPI) staining.
  • Figure 2B shows T cells through anti-CD3 staining.
  • Figure 2C shows NK cells through anti-CD56 staining.
  • Figure 2D shows invariant natural killer T (iNKT) cells through co-localization of anti-CD3/anti-CD56 staining.
  • DAPI diamidino-2-phenylindole
  • Figure 3 depicts eosinophil counts and kinetics after exposing rtTA-CC 10- IL-15 bitransgenic mice to 4, 8, and 12-weeks of a normal diet or a diet of food impregnated with doxycycline (DOX).
  • DOX doxycycline
  • FIG. 4 depicts results from fluorescence-activated cell sorting (FACS) analysis conducted on freshly isolated bone marrow cells (BMCs) from naive mice cultured with and without IL-15 (20 ng/ml) and stem cell growth factor (SCF) (10 ng/ml) for 3 weeks at 37°C and 5% C0 2 ; around 65% of the anti-FcsRIa positive cell subsets of the culture developed into MCs (Fc8RI/C-kit + ), while 35.3% developed into basophils (FcsRI/C-kif).
  • FACS fluorescence-activated cell sorting
  • Figures 5A-B depict enzyme-linked immunosorbent assay (ELISA) analyses of rtTA-CC 10-IL- 15 bitransgenic mice treated with 6 weeks of a diet of normal food or a diet of food impregnated with DOX.
  • Figure 5A depicts esophageal B cell levels.
  • Figure 5B depicts serum IgE levels.
  • Figures 6A-B depict representative flow cytometric histograms after exposing purified B cells to different concentrations of IL-15 in vitro.
  • Figure 6 A depicts IL- 15 (20 ng/ml)-induced activation of B cells, analyzed using anti-CD69 antibody.
  • Figure 6B depicts dose-dependent B cell proliferation, analyzed by thymidine incorporation.
  • Figure 7 depicts a quantitative morphometric analysis of mast cell (MC) numbers in the esophagi of rtTA-CC 10-IL- 15 mice following an 8-week regimen of a diet of normal food or a diet of food impregnated with DOX.
  • Figures 8A-C depict results from bromodeoxyuridine (BrdU) incorporation analysis in saline- and allergen-challenged wild type (WT) and MC-deficient (WW V ) mice.
  • Figure 8A depicts BrdU + cells, as indicated by arrows, found in the epithelium of saline-challenged mice.
  • Figure 8B depicts BrdU + cells, as indicated by arrows, found in both the epithelium and muscularis mucosa of allergen-challenged WT mice; EP indicates the epithelium, and MM indicates the muscularis mucosa.
  • Figure 8C depicts a morphometric quantitative analysis indicating a significant increase of BrdU + cells in the muscularis mucosa of allergen-challenged WT mice compared to allergen-challenged WW V mice.
  • Figures 9A-C depict the detection of anti-CD 19 + B cells in the esophageal biopsies of EoE patients compared to normal individuals.
  • Figure 9A depicts a lack of B cells from esophageal biopsies of normal individuals.
  • Figure 9B depicts B cells, as indicated by arrows, and IgE + cells, as indicated by arrows, detected in the biopsies of EoE patients.
  • Figure 9C depicts the quantification of IgE + cells in the esophageal biopsies of normal (NL) and EoE patients; data are expressed as mean ⁇ standard deviation (SD), with p ⁇ 0.001.
  • SD standard deviation
  • Figure 10 depicts a representative photograph showing esophageal stricture in mice following stomach IL-5 transgene-induced EoE, as detected by feeding the mice barium, then subsequently analyzing the gastrointestinal tract x-ray; esophageal stricture is indicated by the rectangle.
  • Figure 11 depicts eosinophil numbers analyzed in the esophagi of IL- 15Ra-deficient mice following anti-myelin basic protein (MBP) immunostaining.
  • MBP myelin basic protein
  • Figure 12 depicts eosinophil numbers analyzed in the esophagi of CD Id null mice following anti-MBP immunostaining.
  • Figures 13 A-B depict the protein and mRNA levels of IL- 15 and CXCL 16 in the blood and blood leukocytes of EoE patients compared to those of normal individuals.
  • Figure 13A depicts blood IL-15 protein levels.
  • Figure 13B depicts blood mRNA expression levels of iNKT-specific chemokine CXCL 16.
  • Figures 14A-J depict real-time PCR analyses to detect receptor mRNA levels in the blood of normal, EoE, and GERD patients.
  • Figure 14A depicts the relative expression of FcsRI mRNA levels.
  • Figure 14B depicts the relative expression of FcsRII mRNA levels.
  • Figure 14C depicts the relative expression of ⁇ T cell receptor mRNA levels.
  • Figure 14D depicts the relative expression of ⁇ T cell receptor mRNA levels.
  • Figure 14E depicts the relative expression of a T cell receptor mRNA levels.
  • Figure 14F depicts the relative expression of ⁇ T cell receptor mRNA levels.
  • Figure 14G depicts the relative expression of CXCR6 mRNA levels.
  • Figure 14H depicts the relative expression of ⁇ ⁇ mRNA levels.
  • Figure 141 depicts the relative expression of J l8 mRNA levels.
  • Figure 14J depicts the relative expression of Va24 mRNA levels. Data are normalized to glyceraldehyde 3 -phosphate dehydrogenase (GAPDH).
  • GPDH glyceraldehyde 3 -phosphate dehydrogenase
  • non-invasive biomarker refers to a molecule measured in blood whose concentration reflects the severity or presence of some disease state.
  • blood can include, for example, plasma, serum, whole blood, blood lysates, and the like.
  • assessing includes any form of measurement, and includes determining if an element is present or not.
  • determining includes determining if an element is present or not.
  • evaluating includes determining if an element is present or not.
  • assessing includes determining if an element is present or not.
  • determining includes determining if an element is present or not.
  • evaluating includes determining if an element is present or not.
  • assessing includes determining if an element is present or not.
  • assaying can be used interchangeably and can include quantitative and/or qualitative determinations.
  • diagnosis or monitoring with reference to a disease state or condition refers to a method or process of determining if a subject has or does not have a particular disease state or condition or determining the severity or degree of the particular disease state or condition.
  • treatment refers to obtaining a desired pharmacologic and/or physiologic effect.
  • the effect can be prophylactic in terms of completely or partially preventing a disease or symptom thereof and/or can be therapeutic in terms of a partial or complete cure for a disease and/or adverse effect attributable to the disease.
  • Treatment covers any treatment of a disease in a subject, particularly in a human, and includes: (a) preventing the disease from occurring in a subject which may be predisposed to the disease but has not yet been diagnosed as having it; (b) inhibiting the disease, i.e., arresting its development; and (c) relieving the disease, i.e., causing regression of the disease and/or relieving one or more disease symptoms.
  • Treatment can also encompass delivery of an agent or administration of a therapy in order to provide for a pharmacologic effect, even in the absence of a disease or condition.
  • treatment is used in some embodiments to refer to administration of a compound of the present invention to mitigate a disease or a disorder in a host, preferably in a mammalian subject, more preferably in humans.
  • treatment can include includes: preventing a disorder from occurring in a host, particularly when the host is predisposed to acquiring the disease, but has not yet been diagnosed with the disease; inhibiting the disorder; and/or alleviating or reversing the disorder.
  • the term “prevent” does not require that the disease state be completely thwarted (see Webster's Ninth Collegiate Dictionary).
  • the term preventing refers to the ability of the skilled artisan to identify a population that is susceptible to disorders, such that administration of the compounds of the present invention can occur prior to onset of a disease. The term does not mean that the disease state must be completely avoided.
  • modulated or modulation can refer to both up regulation (i.e., activation or stimulation, e.g., by agonizing or potentiating) and down regulation (i.e., inhibition or suppression, e.g., by antagonizing, decreasing or inhibiting), unless otherwise specified or clear from the context of a specific usage.
  • up regulation i.e., activation or stimulation, e.g., by agonizing or potentiating
  • down regulation i.e., inhibition or suppression, e.g., by antagonizing, decreasing or inhibiting
  • the term "marker” or “biomarker” refers to a biological molecule, such as, for example, a nucleic acid, peptide, protein, hormone, and the like, whose presence or concentration can be detected and correlated with a known condition, such as a disease state. It can also be used to refer to a differentially expressed gene whose expression pattern can be utilized as part of a predictive, prognostic or diagnostic process in healthy conditions or a disease state, or which, alternatively, can be used in methods for identifying a useful treatment or prevention therapy.
  • the term "expression levels” refers, for example, to a determined level of biomarker expression.
  • pattern of expression levels refers to a determined level of biomarker expression compared either to a reference (e.g. a housekeeping gene or inversely regulated genes, or other reference biomarker) or to a computed average expression value (e.g. in DNA-chip analyses).
  • a pattern is not limited to the comparison of two biomarkers but is more related to multiple comparisons of biomarkers to reference biomarkers or samples.
  • a certain "pattern of expression levels” can also result and be determined by comparison and measurement of several biomarkers as disclosed herein and display the relative abundance of these transcripts to each other.
  • a "reference pattern of expression levels” refers to any pattern of expression levels that can be used for the comparison to another pattern of expression levels.
  • a reference pattern of expression levels is, for example, an average pattern of expression levels observed in a group of healthy or diseased individuals, serving as a reference group.
  • Eosinophilic esophagitis is an allergen-induced T cell-mediated disease and is differentiated from reflux esophagitis (gastroesophageal reflux disease, or GERD) by the magnitude of mucosal eosinophilia, intraepithelial eosinophil counts, epithelial cell hyperplasia, and the lack of response to acid suppression therapy (Furuta, G. et al. Gastroenterology 133:1342-63 (2007); Rothenberg, M. et al. J. Allergy Clin. Immunol. 108:891-4 (2007)).
  • GERD reflux esophagitis
  • EoE is a chronic disease; however, little has been documented on its natural history (Straumann, A. et al. Gastroenterology 125: 1660-9 (2003)). Despite the increased global incidence of EoE (Orenstein, S. et al. Am. J. Gastroenterol. 95: 1422-30 (2000); Croese, J. et al. Gastrointest. Endosc. 58:516-22 (2003); Attwood, S. et al. Dig. Dis. Sci. 38: 109-16 (1993); Cury, E. et al. J. Pediatr. Surg. 39:e4-7 (2004); Cantu, P. et al. Dig. Liver Dis.
  • EoE therapies include the use of proton pump inhibitors (PPIs), topical glucocorticoids, such as fluticasone or budesonide, humanized antibodies against relevant cytokines, such as eotaxin-3, IL-13, and IL-5, and small molecule inhibitors of an eosinophil and/or allergic disease activation pathway, such as a prostaglandin D2, IL-4, or IL-13 antagonist.
  • PPIs proton pump inhibitors
  • topical glucocorticoids such as fluticasone or budesonide
  • humanized antibodies against relevant cytokines such as eotaxin-3, IL-13, and IL-5
  • small molecule inhibitors of an eosinophil and/or allergic disease activation pathway such as a prostaglandin D2, IL-4, or IL-13 antagonist.
  • PPIs proton pump inhibitors
  • topical glucocorticoids such as fluticasone or budesonide
  • An antigen elimination approach in sensitized individuals is typically unsatisfactory or practically difficult, such as when patients are sensitized to many allergens.
  • a diet consisting exclusively of an elemental (amino acid-based) formula frequently improves symptoms and normalizes esophageal pathology (Teng, M. et al. J. Immunol. 183: 1911-20 (2009); Teng, M. et al. J. Immunol. 182:3366-3371 (2009); Jiang, X. et al. J. Immunol. 175:2051-5 (2005); Liacouras, C. et al. Clin. Gastroenterol.
  • Glucocorticoid treatment shows a significant effect in reducing esophageal eosinophillia, and newer glucocorticoids with decreased systemic effects can improve the care of EoE patients.
  • some treatments have been shown to be effective in EoE, the molecular mechanisms involved in the remission have heretofore not been established and remain unclear.
  • IL-5 and IL-13 are secondary products of activated T cell subsets, as opposed to products that contribute to the growth and survival of T cell subsets like, invariant natural killer T (iNKT) or ⁇ T cells; these secondary products elevate, rather than initiate, EoE pathogenesis.
  • EoE diagnosis criteria have been recommended by an expert panel established as part of the First International Group of EoE researchers (FIGERS) (Liacouras, C. et al. J. Allergy Clin. Immunol. 128:3-20e6 (2011)).
  • EoE diagnosis requires endoscopy, which is an invasive and inconvenient procedure, followed by biopsy analysis to record the characteristic histological features of esophageal mucosal eosinophilia and epithelial proliferative changes.
  • There are few diagnostic biomarkers for EoE other than determination of eosinophil counts/high power field (hpf) from esophageal biopsies; there is also no proven permanent EoE treatment strategy.
  • Non-invasive techniques for the diagnosis of EoE such as biomarker detection methods, would be preferable to endoscopic techniques. Determination of target molecules that have potential use in EoE diagnosis and future therapy would therefore be beneficial, as would innovative fundamental studies which uncover new possibilities for diagnostic and therapeutic interventions.
  • the identification of reliable, non-invasive EoE biomarkers would advance EoE treatment, as it would allow for more accurate and timely detection of changes resulting from therapy administration.
  • the ability to avoid repeated endoscopies will not only have a positive impact in the medical care of the patients, but it will also reduce the possibility of complications related to the invasive nature of the procedure and the cost of patient care.
  • the discovery of a biomarker for EoE would therefore change the approach to the diagnosis and management of patients with EoE and would have broad applicability on clinical practice and patient health.
  • biomarkers for EoE A number of invasive diagnostic and monitoring molecules have been proposed as biomarkers for EoE. However, most biomarkers proposed for EoE diagnosis rely on endoscopic and histological analysis of patient esophageal biopsies, and a number of questions remain regarding their ability to diagnose EoE. In addition, none of the heretofore established non-invasive biomarkers has been proven to differentiate EoE from GERD, a closely related esophageal disease.
  • EoE patients can have eosinophil counts below the FIGURE recommended eosinophil levels ( ⁇ 15 eosinophils/hpf), making diagnosis difficult; there are also clear cases of GERD with biopsies that are difficult to differentiate from EoE, as described in a recent group of PPI-responsive EoE patients (Liacouras, C. et al. J. Allergy Clin. Immunol. 128:3-20e6 (2011)). These observations can lead to a delay in diagnosis or, in the worst-case scenario, to the inappropriate use of therapy.
  • mRNA levels of T cell and B cell receptors namely CXR6, Va24, ⁇ , ⁇ , FceRI, and FceRII, were measured in the blood of EoE patients.
  • the experiments described herein demonstrate that these compounds can be used as non-invasive biomarkers for EoE and can differentiate EoE from GERD.
  • Eosinophils and IgE-associated mast cell (MC)/basophil activation are involved in various steps of EoE pathogenesis.
  • immunoglobulin (Ig) class switching has been shown to be involved in the pathogenesis of allergic diseases (Shimoda, K. et al. Nature 380:630-3 (1996); Punnonen, j. et al. Proc. Natl. Acad. Sci. U.S.A. 90:3730-4 (1993))
  • IL-15- induced IgE therefore can be involved in promoting esophageal pathological abnormalities in EoE.
  • IL-13 mRNA has been demonstrated to be induced in EoE patients compared with normal individuals (Blanchard, C. et al. J. Allergy Clin. Immunol. 120: 1292- 300 (2007)), and murine models have also demonstrated that IL-13 promotes esophageal eosinophilia (Mishra, A. and Rothenberg, M. Gastroenterology 125: 1419-27 (2003)) and remodeling (Zuo, L. et al. J. Immunol. 185:660-9 (2010)) in mice. These data elucidate the relationship between IL-13 and human EoE and demonstrate that murine IL-13 -induced esophageal transcripts resemble the human EoE transcriptome.
  • eotaxin chemokines induce eosinophil trafficking and accumulation by promoting chemoattraction (Rothenberg, M. J. Allergy Clin. Immunol. 113: 11-28 (2004)). Additionally, IL-15 and eotaxin-3 levels were found to correlate with disease severity and distinguished EoE from control individuals (Zhu, X. et al. Gastroenterology 139: 182-93 (2010); Blanchard, C. et al. J. Clin. /rcvest.116:536-47 (2006)).
  • iNKT cells have been shown to be involved in an experimental model of food allergen- and aeroallergen-induced EoE (Rajavelu, P. et al. J. Allergy Clin. Immunol. In Press (2012); Rajavelu, P. et al. AJP-Gasteroenterology Online published on Dec 26, In Press (2010)). Accordingly, very high numbers of accumulated T cells in the esophageal mucosa of human EoE were found to be iNKT cells (Figure 2).
  • IL-15 is an allergen-induced cytokine that mediates diverse biological responses, ranging from proliferation and differentiation to protection from apoptosis.
  • IL-15 binds to a trimeric receptor complex consisting of IL2RP, IL2Ry, and IL15Ra chains.
  • This cytokine shares the IL2RP chain with IL2 and shares the IL2Ry chain with other cytokines (such as IL2, IL4, IL9, IL13, and IL21);
  • IL15Ra is a specific receptor subunit for IL15 (Vasilopoulos, S. and Shaker, R. Curr. Gastroenterol. Rep. 3:225-30 (2001)).
  • IL- 15 is produced by macrophages, epithelial cells, and dendritic cells. IL- 15 activates peripheral T cells and recruits additional proinflammatory cells to the site of inflammation. IL-15 activates esophageal epithelial cells and T cell subsets to produce eosinophil active chemokines (such as eotaxins) (Ohteki, T. Curr. Mol. Med. 2:371-80 (2002); Zhu, X. et al. Gastroenterology 139:182-93 (2010)) and T H 2 cytokines (such as IL-5 and IL-13) (Wilson, S. and Delovitch, T. Nat. Rev. Immunol.
  • eotaxins eotaxins
  • T H 2 cytokines such as IL-5 and IL-13
  • Dendritic cell-derived IL-15 stimulates the proliferation of naive CD8+ T cells and enhances the differentiation of naive T cells toward a 3 ⁇ 42 phenotype in the presence of IL4 (Armitage, R. et al. J. Immunol. 154:483-90 (1995); Tagaya, Y. et al. Immunity 4:329-36 (1996); Waldmann, T. and Tagaya, Y. Annu. Rev. Immunol. 17: 19-49 (1999)).
  • IL-15 is therefore important in EoE compared to the other eosinophil active T H 2 cytokines and chemokines, such as IL-13 and eotaxin-3, which are shown to be induced in response to IL-15 in EoE (Zhu, X. et al. Gastroenterology 139: 182-93 (2010); Vicario, M. et al. Gut 59: 12-20 (2010)).
  • IL-15 is found to be increased in experimental EoE and in blood and esophageal biopsies of pediatric and adult EoE patients and is thus correlated with esophageal eosinophilia in human EoE (Zhu, X. et al. Gastroenterology 139: 182-93 (2010); Figures 1A- D).
  • IL-15 and IL-15Ra gene expression are induced in human and experimental EoE and have a significant role in the pathogenesis of allergen-induced experimental EoE (Zhu, X. et al. Gastroenterology 139:82-193 (2010)), IL-15 can have a key role in the initiation and progression of EoE.
  • IL-15 is a growth, survival, and differentiation factor for iNKT cells (Armitage, R. et al. J. Immunol. 154:483-90 (1995); Tagaya, Y. et al. Immunity 4:329-36 (1996); Waldmann, T. and Tagaya, Y. Annu. Rev. Immunol. 17: 19-49 (1999)), and activated iNKT cells rapidly produce eosinophil active cytokines, such as IL-5 and IL-13 (Wilson, S. and Delovitch, T. Nat. Rev. Immunol. 3:211-22 (2003); Brigl, M. and Brenner, M. Annu. Rev. Immunol.
  • iNKT cells The role of iNKT cells in the induction of experimental EoE has been described in several ways. Firstly, the mRNA/protein of IL-15 and IL-15 responsive iNKT and ⁇ T cell surface molecules and receptors cells have been studied in the esophageal biopsies of humans with EoE induced by food (peanut and corn) allergens and aeroallergens and have been demonstrated to promote iNKT cell-dependent EoE in an allergen-sensitized murine model of EoE (Rajavelu, P. et al. J. Allergy Clin. Immunol. In Press (2012); Rajavelu, P. et al.
  • iNKT cells are selected and restricted by CD Id, a non-classical MHC I-like prototypical iNKT cell ligand and specific Va24Jal8 receptor (Kawano, T. et al. Science 278: 1626-9 (1997)).
  • IL-15 -responsive iNKT cells have been shown to activate B cells to produce IgE and IgG in both innate and acquired immunity (Kim, H. et al. J. Exp. Med. 201 :41-7 (2005); Taniguchi, M. et al. Annu. Rev. Immunol. 21 :483-513 (2003)). Both IgE and B cells have been demonstrated to be induced in human EoE (Vicario, M. et al. Gut 59: 12-20 (2010)). IgE binds and activates eosinophils, MCs, and basophils via its receptor (IgER) (Denburg, J. et al. Int. Arch.
  • IL-15 is a growth and survival factor for iNKT and ⁇ cells.
  • IL-15 also promotes proliferation and activation of B cells and MCs ( Figures 6A-B), and chronic IL-15 in vivo expression induces B cells and IgE in mice ( Figures 5A-B).
  • Esophageal IL-15 overexpression in mice was found to increase MCs in the esophagus and promote epithelial and muscle cell hyperplasia in MCs and B cell proliferation and activation, including Ig class switching in the esophagus ( Figures 5-8).
  • mice designed to overexpress IL-15 were found to develop esophageal eosinophilia, MC hyperplasia, local B cell activation and proliferation, and increased blood IgE and eosinophil levels following 8 weeks of doxycycline (DOX) exposure, indicating that IL-15 promotes IgE induction and Ig class switching ( Figures 3-7).
  • DOX doxycycline
  • IL-15Ra IL-15 responsive ⁇ and iNKT cell receptors
  • CXCL16 and CXCR6 chemokines
  • IgE mediators
  • FCeRI and FCeRII IgE receptors
  • EoE patients have induced IL-15, iNKT cells, B cells, and IgE in the blood and esophagus, and their role in the initiation and progression of EoE pathogenesis has been demonstrated (Zhu, X. et al. Gastroenterology 139: 182-93 (2010); Rajavelu, P. et al. J. Allergy Clin. Immunol. In press (2012); Vicario, M. et al. Gut 59: 12-20 (2010)); similar observations have been made in mice that over-express IL-15, resulting in induction of B cells and IgE ( Figures 3, 5A-B, and 7). These observations are in accordance with an earlier clinical report that demonstrates local B cell and IgE induction in human EoE (Vicario, M. et al. Gut 59: 12-20 (2010)).
  • IL-15 is a growth and survival factor for iNKT cells (Ohteki, T. Curr. Mol. Med. 2:371-80 (2002)), and the studies described herein have demonstrated its promotion of the proliferation and activation of B cells and MCs ( Figures 5A-B and 7). IL-15 has been shown to promote IgE induction in IL-15 overexpressed mice, and IgE levels are elevated in human EoE (Mavi, P. et al. Gastroenterology Online published on Feb. 21 (2012)).
  • IL-15 and the cell surface molecules and chemokines of IL-15 -responsive cells therefore can be studied in order to determine potential non-invasive diagnostic biomarkers for human EoE that also differentiate EoE from GERD.
  • IL-15-induced and IL- 15 -responsive candidate biomarkers namely, IL-15, IL-15 responsive ⁇ and iNKT cell receptors such as IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ 1, Jal8, chemokines such as CXCR6 and CXCL16, mediators such as IgE, and IgE receptors such as FCeRI and FCeRII
  • IL-15, IL-15 responsive ⁇ and iNKT cell receptors such as IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ 1, Jal8, chemokines such as CXCR6 and CXCL16, mediators such as IgE, and IgE receptors such as FCeRI and FCeRII
  • these compounds were investigated with the aim of identifying non-invasive biomarkers of EoE disease activity that could be used to monitor esophageal inflammation without the need for invasive serial surveillance endoscopies.
  • IL- 15 protein and mRNA of an iNKT-specific chemokine were found to be induced in the blood of EoE patients ( Figures 13A-B).
  • the mRNA and protein levels of several other potential diagnostic biomarker molecules were then evaluated in the blood or blood cells of normal individuals and patients with EoE or GERD in order to establish a panel of non-invasive biomarkers for human EoE;
  • IL-15 -responsive cell surface molecules, including receptors, mediators, and chemokines were found to be non-invasive diagnostic biomarkers for EoE capable of differentiating EoE from GERD ( Figures 14 A- J).
  • T cell receptors CXCR6, Va24, ⁇ T, and ⁇ T, and IgE receptors FcsRI and FcsRII were found to be significantly altered in EoE patients compared to normal individuals.
  • the mR A levels of most of these molecules in GERD patients were found to be comparable to those of normal individuals and were significantly altered relative to EoE patients.
  • Certain T cell receptors, including Vail and J l8, were found to be reduced in EoE patients compared to normal individuals; however, these reduced levels were similar in EoE and GERD patients.
  • Additional molecules are studied in the same fashion order to determine their potential as non-invasive biomarkers for EoE capable of differentiating EoE from GERD.
  • IL-15, IL-15Ra, IL-5, IL-13, INFy, CXCL16, and TGF- ⁇ protein and mRNA levels can be compared between EoE patients, GERD patients, and normal individuals.
  • IL-15 and IL-15Ra mRNA levels can be subsequently compared to those of other T H 1 and T H 2 cytokines, such as IL-5, IL-13, and INFy, which are growth and survival factor for eosinophils and MCs/basophils.
  • T H 1 and T H 2 cytokines such as IL-5, IL-13, and INFy, which are growth and survival factor for eosinophils and MCs/basophils.
  • Both types of inflammatory cells are the source of producing TGF- ⁇ , which has been shown to be induced in EoE patients and is a key cytokine in the development of esophageal remodeling and fibrosis (Aceves, S. et al. J. Allergy Clin. Immunol. 119:206-12 (2007); Mishra, A. et al.
  • mice and human EoE have induced MCs in the esophagus and are a rich source of TGF- ⁇ (Abonia, J. et al. J. Allergy Clin. Immunol. 126: 140-9 (2010)). Therefore, these molecules can be potential non-invasive biomarkers for EoE and can potentially differentiate EoE from GERD.
  • IL-15 and iNKT cells therefore can be studied in order to determine potential target molecules for EoE therapy.
  • IL-15 and surface molecules, mediators and chemokines of IL-15 responsive cells are targeted for EoE diagnosis and therapy by combining innovative experimental approaches using EoE experimental models and human samples.
  • sIL-15Ra soluble IL-15Ra
  • anti-CD 1 d neutralizing antibody has been successfully used to neutralize iNKT cells in tumor repression (Teng, M. et al. J. Immunol. 183: 1911-20 (2009); Teng, M. et al. J. Immunol. 182:3366-71 (2009)).
  • CDld gene-deficient mice are shown to be protected from experimental EoE ( Figure 12). Therefore, neutralizing iNKT cells with monoclonal anti-CDld or anti-Va24J l8 or its chemokine anti-CXCL16 can protect mice from developing food allergen- or aeroallergen-induced EoE.
  • Esophageal IL-15 overexpression in mice induces MCs and IgE levels. Therefore, experimental EoE-induced esophageal pathological abnormalities through IL-15- induced IgE neutralization is studied in order to determine whether allergen-induced, IL-15- mediated EoE pathogenesis, such as epithelial and muscle cell hyperplasia and stricture, is dependent on IgE and thus prevented by anti-IgE treatment in mice.
  • allergen-induced, IL-15- mediated EoE pathogenesis such as epithelial and muscle cell hyperplasia and stricture
  • IL-15 and iNKT cells are involved in initiation and progression of EoE
  • IL-15 -responsive iNKT cell surface molecules such as CDld, IL-15Ra, and IL-15- induced IgE, and iNKT-specific chemokine CXCL16 are investigated as potential target molecules for EoE therapy by using their respective neutralizing monoclonal antibodies in individual experiments.
  • the anti-IL-15, anti-IgE, anti-CDld, anti-Va24J l8, anti-CXCL16, and anti-IL-15Ra neutralized and non-neutralized mice are tested for EoE pathogenesis following the food allergen- or aeroallergen-induced experimental EoE protocol (Rajavelu, P.
  • Embodiments of the invention are directed to methods of treating EoE in a patient, wherein the methods comprise analyzing the sample from a patient to determine a level of one or more biomarkers associated with EoE, determining whether the level of the one or more biomarkers associated with EoE is up-regulated or down-regulated relative to a level of the one or more biomarkers measured in a normal individual, wherein the presence of an elevated or reduced level of one or more biomarkers associated with EoE results in the patient being diagnosed with EoE, and treating the patient with an appropriate therapeutic strategy based upon the diagnosis.
  • the methods can be used to distinguish EoE from GERD in a subject.
  • Embodiments of the invention are also directed to methods of diagnosing a patient with EoE, wherein the methods comprise analyzing the sample from a patient to determine a level of one or more biomarkers associated with EoE, and determining whether the level of the one or more biomarkers are up-regulated or down-regulated relative to a level of the one or more biomarkers measured in a normal individual, wherein the presence of an elevated or reduced level of one or more biomarkers associated with EoE results in the patient being diagnosed with EoE.
  • the one or more biomarkers associated with EoE include one or more cytokines, IL-15-responsive iNKT, T, and B cell receptors, chemokines, mediators, and IgE receptors.
  • the one or more biomarkers associated with EoE include IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • At least 2 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • at least 3 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • At least 4 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • at least 5 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • At least 6 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • at least 7 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • At least 8 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • at least 9 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • At least 10 biomarkers associated with EoE are selected from IL-5, IL-13, IL- 15, F Fy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • at least 11 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, F Fy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • At least 12 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, F Fy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • at least 13 biomarkers associated with EoE are selected from IL-5, IL-13, IL- 15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • At least 14 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, F Fy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • at least 15 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, F Fy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • At least 16 biomarkers associated with EoE are selected from IL-5, IL-13, IL- 15, F Fy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • all of the biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • 1, 2, 3, 4, 5, 6, 7, 8, or 9 biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • 10, 11, 12, 13, 14, 15, 16, or 17 biomarkers associated with EoE are selected from IL-5, IL-13, IL- 15, F Fy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • biomarkers associated with EoE are selected from IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • anywhere between 1-17 biomarkers associated with EoE are selected from IL-5, IL-13, IL- 15, F Fy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • the one or more biomarkers associated with EoE are measured using one or more methods and/or tools, including for example, but not limited to, Taqman (Life Technologies, Carlsbad, CA), Light-Cycler (Roche Applied Science, Penzberg, Germany), ABI fluidic card (Life Technologies), NanoString® (NanoString Technologies, Seattle, WA), NANODROP® technology (Thermo Fisher Scientific (Wilmington, DE), fluidic card, and the like.
  • Taqman Life Technologies, Carlsbad, CA
  • Light-Cycler Roche Applied Science, Penzberg, Germany
  • ABI fluidic card Life Technologies
  • NanoString® NanoString Technologies, Seattle, WA
  • NANODROP® technology Thermo Fisher Scientific (Wilmington, DE), fluidic card, and the like.
  • the person of skill in the art will recognize such other formats and tools, which can be commercially available or which can be developed specifically for such analysis.
  • Determination of the biomarker level(s) as described herein can be combined with determination of the levels of one or more additional biomarkers associated with EoE.
  • determination of the biomarker level(s) as described herein can be combined with determination of the levels of one or more genes of the EoE transcriptome.
  • Such a determination can include measurement of the gene DNA or RNA, or the gene product.
  • genes can include, for example, eotaxin-3, and the like.
  • Certain embodiments of the invention involve administering EoE therapies, including allergen removal, steroid treatment, dietary management, proton pump inhibitor (PPI) therapy, administration of one or more topical glucocorticoid, administration of one or more humanized antibody against one or more relevant cytokines and/or mediators, administration of one or more small molecule inhibitors of an eosinophil and/or allergic disease activation pathway, administration of one or more small molecule inhibitors capable of modulating levels of one or more biomarkers associated with EoE, and/or any combination thereof.
  • Topical glucocorticoids that can be used as EoE therapies include, for example, fluticasone, budesonide, ciclesonide, and the like.
  • Humanized antibodies against one or more relevant cytokines and/or mediators can include, for example, antibodies targeting one or more biomarkers associated with EoE.
  • the one or more biomarkers associated with EoE can include, for example, IL-5, IL-13, IL-15, INFy, TGF- ⁇ , IL-15Ra, ⁇ , ⁇ , CDld, Va24, ⁇ ⁇ , Jal8, CXCR6, CXCL16, IgE, FCeRI, and/or FCeRII.
  • the humanized antibody against one or more relevant cytokines and/or mediators can include, for example, anti-IL-15, anti-IgE, anti-CDld, anti-Va24J l8, anti-CXCL16, and anti-IL-15Ra, and the like.
  • EoE include for example, but are not limited to, libraries of known compounds, including natural products, such as plant or animal extracts, synthetic chemicals, biologically active materials including proteins, peptides such as soluble peptides, including but not limited to members of random peptide libraries and combinatorial chemistry derived molecular libraries made of D- or L-configuration amino acids, or both, phosphopeptides (including, but not limited to, members of random or partially degenerate, directed phosphopeptide libraries), antibodies (including, but not limited to, polyclonal, monoclonal, chimeric, human, anti- idiotypic or single chain antibodies, and Fab, F(ab') 2 and Fab expression library fragments, and epitope-binding fragments thereof), organic and inorganic molecules, and the like.
  • libraries of known compounds including natural products, such as plant or animal extracts, synthetic chemicals, biologically active materials including proteins, peptides such as soluble peptides, including but not limited to members of random peptide libraries and combinatori
  • a model can also be generated by building models of the hydrophobic helices. Mutational data that point towards residue-residue contacts can also be used to position the helices relative to each other so that these contacts are achieved. During this process, docking of the known ligands into the binding site cavity within the helices can also be used to help position the helices by developing interactions that would stabilize the binding of the ligand.
  • the model can be completed by refinement using molecular mechanics and loop building using standard homology modeling techniques. (General information regarding modeling can be found in Schoneberg, T. et. al. Molecular and Cellular Endocrinology 151 : 181-93 (1999); Flower, D. Biochimica et Biophysica Acta 1422:207-34 (1999); and Sexton, P. Current Opinion in Drug Discovery and Development 2:440-8 (1999).)
  • the model can be used in conjunction with one of several existing computer programs to narrow the number of compounds to be screened by the screening methods of the present invention, like the DOCK program (UCSF Molecular Design Institute, San Francisco, CA). In several of its variants it can screen databases of commercial and/or proprietary compounds for steric fit and rough electrostatic complementarity to the binding site.
  • Another program that can be used is FLEXX (Tripos Inc., St. Louis, MO).
  • the compounds and antibodies used as therapeutic targets or agents as described above can be administered via oral or parenteral delivery routes (subcutaneous or intravenous).
  • Such therapeutics can be administered by any pharmaceutically acceptable carrier, including, for example, any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration.
  • any pharmaceutically acceptable carrier including, for example, any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration.
  • the use of such media and agents for pharmaceutically active substances is known in the art. Except insofar as any conventional medium or agent is incompatible with the active compound, such media can be used in the compositions of the invention.
  • Supplementary active compounds can also be incorporated into the compositions.
  • a pharmaceutical composition of the invention is formulated to be compatible with its intended route of administration.
  • Routes of administration include for example, but are not limited to, intravenous, intramuscular, and oral, and the like. Additional routes of administration include, for example, sublingual, buccal, parenteral (including, for example, subcutaneous, intramuscular, intraarterial, intradermal, intraperitoneal, intracisternal, intravesical, intrathecal, or intravenous), transdermal, oral, transmucosal, and rectal administration, and the like.
  • Solutions or suspensions used for appropriate routes of administration can include, for example, the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfate; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates, or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose, and the like.
  • a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents
  • antibacterial agents such as benzyl alcohol or methyl parabens
  • antioxidants such as ascorbic acid or sodium bisulfate
  • chelating agents such as
  • the pH can be adjusted with acids or bases, such as, for example, hydrochloric acid or sodium hydroxide, and the like.
  • the parenteral preparation can be enclosed in, for example, ampules, disposable syringes, or multiple dose vials made of glass or plastic, and the like.
  • compositions suitable for injectable use include, for example, sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion, and the like.
  • suitable carriers include, for example, physiological saline, bacteriostatic water, Cremophor ELTM (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS), and the like.
  • the composition should be fluid to the extent that easy syringability exists.
  • the carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), and suitable mixtures thereof, and the like.
  • the proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants.
  • Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, such as, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like.
  • isotonic agents such as, for example, sugars, polyalcohols such as mannitol, sorbitol, and sodium chloride, and the like, in the composition.
  • Prolonged absorption of the injectable compositions can be brought about by including in the composition an agent which delays absorption such as, for example, aluminum monostearate and gelatin, and the like.
  • Sterile injectable solutions can be prepared by incorporating the active compound in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by filtered sterilization.
  • dispersions are prepared by incorporating the active compound into a sterile vehicle that contains a basic dispersion medium and the required other ingredients from those enumerated above.
  • the preferred methods of preparation are vacuum drying and freeze-drying which yields a powder of the active ingredient plus any additional desired ingredient from a previously sterile- filtered solution thereof.
  • Oral compositions generally include an inert diluent or an edible carrier. They can be enclosed in gelatin capsules or compressed into tablets, for example.
  • the agent can be contained in enteric forms to survive the stomach or further coated or mixed to be released in a particular region of the gastrointestinal (GI) tract by known methods.
  • the active compound can be incorporated with excipients and used in the form of tablets, troches, or capsules, or the like.
  • Oral compositions can also be prepared using a fluid carrier for use as a mouthwash, wherein the compound in the fluid carrier is applied orally and swished and expectorated or swallowed.
  • Pharmaceutically compatible binding agents, and/or adjuvant materials can be included as part of the composition.
  • the tablets, pills, capsules, troches, and the like can contain any of the following exemplary ingredients, or compounds of a similar nature: a binder such as microcrystalline cellulose, gum tragacanth or gelatin; an excipient such as starch or lactose, a disintegrating agent such as alginic acid, Primogel®, or corn starch; a lubricant such as magnesium stearate; a glidant such as colloidal silicon dioxide; a sweetening agent such as sucrose or saccharin; or a flavoring agent such as peppermint, methyl salicylate, or orange flavoring, or the like.
  • a binder such as microcrystalline cellulose, gum tragacanth or gelatin
  • an excipient such as starch or lactose, a disintegrating agent such as alginic acid, Primogel®, or corn starch
  • a lubricant such as magnesium stearate
  • a glidant such as colloidal silicon dioxide
  • the compounds can be delivered in the form of an aerosol spray from pressured container or dispenser, which contains a suitable propellant, e.g., a gas such as carbon dioxide, or a nebulizer, or the like.
  • a suitable propellant e.g., a gas such as carbon dioxide, or a nebulizer, or the like.
  • Systemic administration can also be by transmucosal or transdermal means.
  • penetrants appropriate to the barrier to be permeated are used in the formulation.
  • penetrants are generally known in the art, and include, for example, for transmucosal administration, detergents, bile salts, and fusidic acid derivatives, and the like.
  • Transmucosal administration can be accomplished through the use of nasal sprays or suppositories.
  • the active compounds are formulated into ointments, salves, gels, or creams as generally known in the art.
  • the compounds can also be prepared in the form of suppositories ⁇ e.g., with conventional suppository bases such as cocoa butter and other glycerides) or retention enemas for rectal delivery.
  • suppositories ⁇ e.g., with conventional suppository bases such as cocoa butter and other glycerides
  • retention enemas for rectal delivery.
  • the active compounds are prepared with carriers that will protect the compound against rapid elimination from the body, such as a controlled release formulation, including implants and microencapsulated delivery systems, and the like.
  • a controlled release formulation including implants and microencapsulated delivery systems, and the like.
  • Biodegradable, biocompatible polymers can be used, such as, for example, ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid, and the like. Methods for preparation of such formulations will be apparent to those skilled in the art.
  • the materials can also be obtained commercially from Alza Corporation and Nova Pharmaceuticals, Inc.
  • Liposomal suspensions (including liposomes targeted to infected cells with monoclonal antibodies to viral antigens) can also be used as pharmaceutically acceptable carriers.
  • dosage unit form refers to physically discrete units suited as unitary dosages for the subject to be treated; each unit containing a predetermined quantity of active compound calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier.
  • the details for the dosage unit forms of the invention are dictated by and directly dependent on the unique characteristics of the active compound and the particular therapeutic effect to be achieved, and the limitations inherent in the art of compounding such an active compound for the treatment of individuals. Such details are known to those of skill in the art.
  • Certain embodiments of the invention include using quantification data from a gene-expression analysis and/or from a mRNA analysis, from a sample of blood, urine, saliva, esophageal tissue, or the like.
  • Embodiments of the invention include not only methods of conducting and interpreting such tests but also include reagents, kits, assays, and the like, for conducting the tests.
  • the correlations disclosed herein, between EoE biomarker levels and/or mRNA levels and/or gene expression levels, provide a basis for conducting a diagnosis of EoE, or for conducting a stratification of patients with EoE, or for enhancing the reliability of a diagnosis of EoE by combining the results of a quantification of an EoE biomarker with results from other tests or indicia of EoE.
  • the results of a quantification of one biomarker could be combined with the results of a quantification of one or more additional biomarker, cytokine, mRNA, or the like.
  • the correlation can be one indicium, combinable with one or more others that, in combination, provide an enhanced clarity and certainty of diagnosis. Accordingly, the methods and materials of the invention are expressly contemplated to be used both alone and in combination with other tests and indicia, whether quantitative or qualitative in nature.
  • p values below 0.05 are considered to be statistically significant, it is within the scope of embodiments of the present invention to make use of correlations having a reported p value above 0.05 as well as below 0.05.
  • a p value can be above 0.05, such as, for example, 0.06, 0.07, 0.08, 0.09, 0.10, 0.15, or more.
  • p value is affected by sample size, two studies can have the same proportion of outcomes, and a study with a smaller sample size can have a p value above 0.05, while the study with the larger sample size can have a p value below 0.05, even though the correlation is proportionally the same.
  • a p value below 0.05 for any sample size, is a strong indication of a statistically significant correlation, a genuine correlation can exist, that is tested with a small sample size, and the p value of such a test can be above 0.05.
  • IL-15 is increased in the blood and esophageal biopsies of EoE patients (Zhu, X. et al. Gastroenterology 139: 182-93 (2010)). Therefore, IL-15 immunoreactivity was tested on non-EoE and EoE patient esophageal biopsy samples.
  • EoE patient esophageal biopsies were shown to contain a number of IL- 15-positive cells. There were no IL-15 -positive cells in the esophageal biopsies of non-EoE patients ( Figures 1A-B), whereas a number of IL-15 -positive cells were detected in EoE patient biopsies ( Figures 1C-D). These data indicate that infiltrating cells are the source of IL-15 and that IL-15 blood levels can be used as a non-invasive diagnostic biomarker for human EoE.
  • IL-15 is a growth and survival cytokine for iNKT cells. Therefore, a study was conducted to determine whether esophageal induction of IL-15 causes accumulation of iNKT cells in the esophageal mucosa of EoE patients.
  • Esophageal biopsies of normal individuals and EoE patients were stained with various media ( Figures 2A-D) to show the cell nuclei of a representative EoE patient esophageal biopsy.
  • Figure 2A shows the cell nuclei through 4',6-diamidino-2-phenylindole (DAPI) staining.
  • Figure 2B shows T cells through anti-CD3 staining.
  • Figure 2C shows NK cells through anti-CD56 staining.
  • Figure 2D shows iNKT cells through co-localization of anti-CD3/anti-CD56 staining.
  • IL-15 has a role in EoE pathogenesis and the induced expression of IL-15 mRNA and protein in human EoE (Zhu, X. et al. Gastroenterology 139:182-93 (2010)). Therefore, the mechanism of IL-15-induced EoE pathogenesis can be examined by generating DOX-regulated rtTA-CClO-IL-15 bitransgenic mice.
  • Esophageal IL-15 overexpressing bitransgenic mice were developed using rtTA (activator) and CClO-IL-15 (responder) constructs.
  • the rtTA-CClO-IL-15 transgenic mice generated to chronically over-express IL- 15 in the esophagus were either exposed to a doxycycline (DOX) diet or a DOX- free diet at 8 weeks old and were then kinetically examined for esophageal B cell activation and proliferation, the induction of Ig class switching genes, and serum levels of IgE and IgG.
  • DOX doxycycline
  • DOX-induced IL-15 was detectable within 4 weeks of continued DOX treatment in the lungs and esophagi of the mice.
  • Initial examination after 4, 8, and 12-weeks of DOX or no-DOX treatment showed a time dependent increase of esophageal eosinophilia in DOX-regulated IL-15 bitransgenic mice, along with the kinetics ( Figure 3). These mice can be used to examine EoE pathogenesis induced by chronic, postnatal IL-15.
  • IL-15 IS A GROWTH AND SURVIVAL FACTOR FOR MCS/BASOPHILS
  • IL-15 can be a growth factor for MCs/basophils.
  • BMCs bone marrow cells
  • SCF stem cell growth factor
  • IL-15 Blood and esophageal IL-15, IgE, and B cell levels are increased in EoE patients. Therefore, IL-15 can directly activate and cause proliferation of B cells.
  • Purified B cells were exposed to different concentrations of IL-15 in vitro in order to determine the dose-dependent activation and proliferation of B cells in response to
  • IL-15 Flow cytometry was used to determine IL-15 (20 ng/ml)-induced activation of B cells, analyzed using anti-CD69 antibody ( Figure 6A), and dose-dependent B cell proliferation, analyzed by thymidine incorporation ( Figure 6B).
  • IL-15 is shown to have B cell stimulatory activity for B cell activation and proliferation. These results indicate that EoE pathogenesis is B cell-induced and IgE- mediated.
  • MCs are localized in the esophageal lamina limbal growth factor (IL1), and muscularis mucosa following the induction of EoE in mice. Therefore, induced MCs can promote esophageal muscle cell hyperplasia and hypertrophy in EoE.
  • Bromodeoxyuridine (BrdU) incorporation analysis was performed in saline- and allergen-challenged wild type (WT) and MC-deficient (WW V ) mice.
  • BrdlT cells were detected in the epithelium of saline-challenged mice ( Figure 8A), whereas allergen- challenged mice showed BrdU + cells in both the epithelium and muscularis mucosa of WT mice ( Figure 8B).
  • the morphometric quantitative analysis indicated a significant increase of BrdU + cells in the muscularis mucosa of allergen-challenged WT mice compared to the allergen-challenged WW V mice ( Figure 8C).
  • IgE, B cells, MCs and IgE + cells have been shown to be induced in the blood and/or esophageal biopsies of EoE patients (Vicario, M. et al. Gut 59: 12-20 (2010)).
  • Anti-CD 19 B cells were detected in the esophageal biopsies of EoE patients compared to normal individuals.
  • Significant numbers of B cells were not detected in the esophageal biopsies of normal individuals ( Figure 9A); however, a number of B cells and lgE + cells were detected in the biopsies of EoE patients ( Figure 9B).
  • IgE + cell numbers in the esophageal biopsies of normal and EoE patients were quantified ( Figure 9C).
  • Esophageal stricture has been reported in patients in a number of esophageal diseases, including EoE (Ruigomez, A. et al. Am. J. Gastroenterol. 101 :2685-92 (2006); Vasilopoulos, S. and Shaker, R. Curr. Gastroenterol. Rep. 3:225-230 (2001)). Therefore, esophageal stricture can develop in experimental murine model of EoE.
  • Esophageal stricture following chronic eosinophilic esophageal inflammation in mice was examined in an experimental murine model of EoE. Esophageal stricture was detected by feeding the mice barium, then subsequently analyzing the gastrointestinal tract x-ray. Esophageal stricture is demonstrated in mice following stomach IL-5 transgene-induced EoE ( Figure 10). This esophageal abnormality was not observed in age- and sex-matched WT mice.
  • IL-15 mRNA and protein production are induced in the esophagus following the induction of experimental and human EoE. Therefore, IL-15 -deficiency can protect against induction of EoE in mice.
  • CD Id is an iNKT-specific cell surface molecule
  • FACS fluorescence- activated cell sorting
  • CDld null mice were shown to have significant reductions in EoE compared to WT mice ( Figure 12). These data indicate that CDld or iNKT cell-specific or cell surface molecules, such as Jal8, can be used as targets for EoE therapy.
  • IL-15 and CXCL16 are induced in the esophageal biopsies of human EoE (Zhu, X. et al. Gastroenterology 139: 182-93 (2010); Rajavelu, P. et al. Proc. Natl. Acad. Sci. Under review (2011)). Therefore, ELISA and real time PCR analysis were performed to determine the protein and mRNA levels of IL-15 and CXCL16 in the blood and blood leukocytes of EoE patients.
  • the blood from patients with EoE typically has a small population of infiltrating mononuclear cells consistent with lymphocytes (Bullock, J. et al. J. Pediatr. Gastroenterol. Nutr. 45:22-31 (2007)), but these cells have not been extensively characterized.
  • EoE patients blood leukocyte subsets of normal individuals and GERD or EoE patients were investigated following flow cytometric and qPCR analysis, in order to determine whether EoE patients have increased levels of IL-15 -responsive iNKT cells and B cells, as well as increased mRNA and protein levels of cell-specific receptors, such as IL- 15Ra, ⁇ , Va24, Jal8, and CXCR6, in the blood leukocytes compared to normal individuals or GERD patients in order to determine their potential as non-invasive diagnostic biomarkers for human EoE that also differentiate EoE from GERD.
  • Most of the molecular candidates for monitoring EoE pathogenesis were selected based on their significant roles in EoE, as reported in previous studies (Zhu, X.
  • GPDH glyceraldehyde 3 -phosphate dehydrogenase
  • nCounter Analysis SystemTM Nastring Technology, Seattle, WA
  • the nCounter Analysis System was used if necessary, using a novel digital technology that is based on direct multiplexed measurement of gene expression and offers high levels of sensitivity ( ⁇ 1 copy per cell) and precision.
  • the technology uses single-molecule imaging and high numbers of unique transcripts in a single reaction. Such technology permits using the small amount of RNA available from the blood for monitoring the patient disease status following the treatment.
  • Another advantage of the nCounter AnalysisTM is that design of the diagnosis chips can be custom designed based on the genes of interest.
  • the mRNA levels of the ⁇ T cell receptor were also studied, and both ⁇ and ⁇ were found to be significantly reduced in the blood of EoE patients compared to GERD patients and normal individuals ( Figures 14C-D).
  • the mRNA levels of the ⁇ T cell receptor were also studied, and both a and ⁇ were found to be significantly reduced in EoE patients compared to normal individuals and GERD patients ( Figures 14E-F).
  • the mRNA levels of the CXCR6 T cell receptor were also found to be significantly reduced in the blood of EoE patients compared to GERD patients and normal individuals ( Figure 14G).
  • mRNA levels of ⁇ 1 and J l8 were found to be reduced in the blood of EoE patients compared to normal individuals, but these levels did not differentiate GERD patients from EoE patients ( Figures 14H-I).
  • the mRNA levels of Va24 were found to be significantly reduced in the blood of EoE patients compared to normal individuals and EoE patients ( Figure 14 J).
  • transcript levels of FCeRI, FCeRII, ⁇ , ⁇ , CXCR6, ⁇ ⁇ , Jal8, and Va24 can be used as non-invasive biomarkers for EoE. Further, transcript levels of FCeRI, FCeRII, ⁇ , ⁇ , CXCR6, and Va24 can be used to distinguish EoE from GERD.
  • the relative levels of IL-15 and IL-15Ra mRNA are measured in the blood leukocytes and IL-15 protein in the serum of normal individuals and patients with EoE or GERD, in order to determine whether IL-15 and IL-15Ra mRNA and protein levels in the blood can be used as non-invasive diagnostic biomarkers for EoE capable of differentiating EoE from GERD.
  • the IL-15 and IL-15Ra mRNA levels are subsequently compared to those of other T H 1 and T H 2 cytokines, such as IL-5, IL-13, and INFy.
  • the relative expression of cytokine mRNA is quantified by qPCR using the LightCycler instrument (Bio-Rad, Philadelphia, PA), as previously described (Zhu, X. et al. Gastroenterology 139: 182-93 (2010); Zhu, X. et al. Am. J. Physiol. Gastrointest. Liver Physiol. 297:G550-8 (2009); Rayapudi, M. et al. J. Leukoc. Biol. 88:337-46 (2010)).
  • the results are normalized by amplified GAPDH of same cDNA and expressed as fold induction.
  • IL-5, IL-13, IL-15, and INFy protein and mRNA levels are examined in serum from normal individuals and patients with EoE or GERD using a commercially available ELISA kit (R&D Systems, Minneapolis, MN). Increased or decreased levels of IL-15 and IL-15Ra mRNA and protein (compared to other T H 2 cytokines) in the blood of EoE patients compared to GERD patients or normal individuals allow these molecules to be used as non-invasive biomarkers for EoE.
  • the mRNA and protein levels of CXCL16 are also examined in blood obtained from normal individuals and EoE or GERD patients by performing qPCR, as previously described (Zhu, X. et al.
  • the soluble CXCL16 is subsequently measured in the blood using a commercially available ELISA kit (R&D Systems).
  • Soluble TGF- ⁇ in the blood of these patients is also measured in blood obtained from normal individuals and EoE or GERD patients by qPCR, in order to determine whether EoE patients have increased mRNA and protein levels of TGF- ⁇ compared to normal individuals and GERD patients.
  • IL-15 and iNKT cells can be studied in order to determine potential target molecules for therapeutic interventions.
  • the proposed molecules are tested in the food allergen- and aeroallergen-induced murine model of experimental EoE using specific humanized neutralizing monoclonal antibodies.
  • a neutralizing antibody therapy found to be successful in a mouse model of experimental EoE provides an experimental framework to support clinical trials of these molecules in human EoE.
  • the anti-IL-15, anti-IgE, anti-CDld, anti-Va24Jal8, anti- CXCL16, and anti-IL-15Ra neutralized and non-neutralized mice are tested for EoE pathogenesis following the food allergen- or aeroallergen-induced experimental EoE protocol (Rajavelu, P. et al. AJP-Gasteroenterology Online published on Dec 26, In Press (2010); Mishra, A. et al. J. Clin. Invest. 107:83-90 (2001)).
  • Food allergen- and aeroallergen-induced experimental EoE is induced in mice neutralized with anti-IL-15 and anti-IL-15Ra neutralized or non-neutralized mice, along with isotype-matched IgGs, as previously described (Mishra, A. et al. J. Clin. Invest. 107:83-90 (2001); Blanchard, C. et al. Clin. Exp. Allergy 35: 1096-103 (2005)), in order to determine whether neutralizing IL-15 or IL-15Ra antibody treatment prior to aeroallergen challenge can protect mice from developing allergen- induced experimental EoE. [0128] Mice are sacrificed and first analyzed for neutralization by performing ELISA and FACS analysis, as previously described (Zhu, X.
  • mice are then neutralized with the relevant antibodies, all of which are commercially available (eBiosciences or R&D Systems). Studies are then conducted to measure esophageal eosinophilia in antibody-neutralized mice compared to non-neutralized mice following experimental EoE induction.
  • the numbers expressing quantities of ingredients, properties such as molecular weight, reaction conditions, and so forth, used to describe and claim certain embodiments of the application are to be understood as being modified in some instances by the term "about.” Accordingly, in some embodiments, the numerical parameters set forth in the written description and attached claims are approximations that can vary depending upon the desired properties sought to be obtained by a particular embodiment. In some embodiments, the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of some embodiments of the application are approximations, the numerical values set forth in the specific examples are reported as precisely as practicable.

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Abstract

L'invention concerne des procédés et des compositions qui sont associés en général à des procédés qui permettent de fournir ou d'améliorer un diagnostic d'œsophagite éosinophile (EoE) par l'identification, la validation et la mesure de biomarqueurs quantifiables, cliniquement pertinents, de diagnostic et de réponses thérapeutiques pour un dysfonctionnement du tractus gastro-intestinal, particulièrement lorsque ces réponses sont associées à EoE. En particulier, l'invention concerne l'identification d'un ou de plusieurs biomarqueurs associés à EoE, l'obtention d'un échantillon prélevé sur un patient ayant au moins une indication d'une EoE, puis la quantification à partir de l'échantillon d'une quantité d'un ou de plusieurs desdits biomarqueurs, le niveau dudit biomarqueur étant corrélé avec le résultat prédit. L'invention concerne en outre des nécessaires de diagnostic, des tests et/ou des réseaux qui peuvent être utilisés pour quantifier le ou les biomarqueurs associés à EoE.
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