Tissue Destruction and Healing in Celiac Disease
Phase: NA
Conditions: Celiac Disease
Sponsor: University of Chicago

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COVER PAGE 
 
Official Title of the Study: Tissue destruction and healing in celiac disease 
NCT Number: not yet assigned 
Document Date: 2022.08.25 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Study Investigators:   
Sonia Kupfer, MD  
 
 
 
 
Bana Jabri, MD, PhD 
 
 
 
 
Valerie Abadie, PhD 
 
 
 
 
Luis Barreiro, PhD 
 
 
 
 
University of Chicago 
 
 
 
 
 
 
Joseph Murray, MD 
 
 
 
 
Mayo Clinic 
 
 
 
 
 
Rustem Ismagilov, PhD 
 
 
 
 
California Institute of Technology 
 
 
 
1. Background  
 
Celiac disease (CeD) is an autoimmune-like small intestinal enteropathy induced by dietary 
gluten in genetically susceptible HLA-DQ2 or DQ8 individuals. There is a spectrum in tissue 
damage: potential CeD patients develop inflammatory IFN-γ anti-gluten T cell immunity in the 
absence of tissue damage, while active CeD is associated with tissue damage ranging from 
partial to total villous atrophy (VA). Gluten-free diet (GFD) is to this date the only treatment for 
CeD. However, 50% of CeD patients are unable to effectively adhere to a diet that sustainably 
excludes gluten, suffering therefore from inadvertent gluten exposure.  Moreover, more than 
30% of CeD patients have persistent high symptom burden, resulting from continued mucosal 
damage despite adhering to a GFD.  
 
Persistent mucosal damage on a GFD is associated with several severe complications, 
including lymphoproliferative malignancy and bone diseases. In addition, patients with active 
CeD display a wide range of clinical presentations, including metabolic defects (vitamin, iron, 
cholesterol) that are not correlated to the degree of VA. Although much progress has been 
made in understanding CeD, major gaps remain in biologic mechanisms underlying 
interindividual differences in clinical presentations and capacity to heal on a GFD. In CeD, there 
is no data available on gene expression variation in the targeted tissue across the disease 
spectrum. There are publications on a subset of key individual immune cell types, but we lack a 
global analysis of all immune cell subsets present in the intestinal mucosa. Furthermore, while 
there is evidence for a role of the microbiome in CeD, we lack information on the duodenal 
mucosal-associated and luminal microbiota, which are most likely to directly interact with the 
immune system and have metabolic effects. Finally, very little is known about interactions 
between intestinal epithelial cells (IECs), immune cells and the microbiota in CeD, and how they 
are linked to gluten and the different CeD clinical phenotypes.  
 
To address these gaps, we have assembled a team of internationally recognized experts in the 
field of CeD, mucosal immunology and microbiome. The RC2 proposal is anchored around 
multi-omics studies performed in the context of longitudinal interventional gluten challenge and 
de-challenge studies, on 180 clinically well-characterized CeD patients and 40 HLA-matched 
healthy controls.  
 
 
 
 

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2. Purpose  
 
The purpose of this study is to: (i) address critical gaps in our understanding of CeD 
pathogenesis and clinical presentations, (ii) test the hypothesis that interactions between IECs, 
microbiota, immune system, genetics and gluten underlie differences in clinical presentation, 
severity of tissue destruction, and ability to heal (iii) generate resources and hypotheses to 
advance patient care.  
 
Our aims are: 
Aim 1: Gluten challenge studies to gain a mechanistic understanding of tissue destruction. 
We will perform longitudinal gluten challenge studies combined with integrated multi-omics 
investigation on a well-curated cohort of 90 GFD patients. Each patient will undergo three 
endoscopies. Studies will combine epithelial, microbial, genetic, metabolic and immunological 
analyses, and yield a large diverse set of biobanked samples. We will characterize the molecular, 
epithelial metabolomic and metagenomics signature of tissue destruction and develop a predictive 
model of tissue destruction.  Extensive clinical metadata and histological data will be obtained at 
all three time points.    
 
Aim 2: Gluten de-challenge studies to gain a mechanistic understanding of tissue healing. 
Through longitudinal gluten de-challenge studies with three endoscopies that will include 90 
patients, we will characterize the molecular, epithelial, genetic, metabolomic and metagenomics 
signature of tissue healing and develop a predictive model of tissue healing. This study will also 
yield a large diverse set of biobanked samples. Extensive clinical metadata and histological data 
will be obtained at all three time points.    
 
Aim 3: Developing research resources for the scientific and medical community to 
advance patient care and hypothesis-generating science. We will collect a large array of 
biosamples (580 biopsy DNA and RNA samples, 580 PBMCs and serum samples, 580 frozen 
biopsies for bacterial cultures, 580 frozen biopsies for host-bacterial imaging, 580 urine and saliva 
samples, 400 intestinal organoids). Together, the U Chicago and Mayo Clinic biobanks will have 
∼4640 biobanked samples. In addition, a large resource of multi-omics data sets will be made 
available to the research community (∼2790) in Aims 1 and 2. To integrate such large and complex 
datasets, we will build an integrated clinical research data management and informatics platform 
that will support harmonization, curation, dissemination, and analysis of data collected at various 
levels. All these data will be made publicly available and searchable using a user-friendly web 
platform. Together, this integrated clinical research data management, biobank and informatics 
platform will constitute a powerful resource for the scientific and medical community. 
 
This study, in addition to generating large integrated omics data sets and hypotheses on 
mechanisms underlying tissue destruction and healing, will develop tools to improve the clinical 
follow-up of CeD patients, and resources for the scientific and medical community to advance 
CeD research and clinical care. 
 
3. Methodology 
a. Study overview. To understand the mechanisms underlying tissue destruction and healing we 
have designed longitudinal challenge and de-challenge studies with 90 patients in each 
interventional arm. In addition, 40 MHC-matched healthy controls will be enrolled and undergo 
one endoscopy with samples collection.  Clinical data and omics data will be obtained at each 
endoscopy, and samples will be collected for biobanking (see Fig. 1). Subjects will be recruited 

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equally at the University of Chicago and Mayo Clinic, Rochester. The fully Schedule of Events 
can be found in Appendix A. 
 
 
 
b. Longitudinal challenge study.  
 
 
 
 
 
In the gluten challenge study arm (Figure 2), 90 celiac disease patients (45 from University of 
Chicago; 45 from Mayo Clinic) will be enrolled. After informed consent, patients will have blood 
drawn for screening labs. Qualifying patients will be asked to return to undergo a research 
endoscopy with biopsies and luminal content sampling. After confirmation that their histology is 
normal, the subjects will proceed to a gluten challenge with 3g vital gluten per day. On day 6 after 
starting the challenge, a research endoscopy with be performed with biopsies and luminal content 
sampling. Subjects will continue on 3g vital gluten each day for 6 weeks. At the end of study, 
patients will undergo a research endoscopy with biopsies and luminal content sampling. In 
addition to research endoscopies, patients will have saliva, serum and urine sampling at baseline, 
2 hour after first gluten ingestion on day 6 and end of study. 
 
 
Gluten challenge will be provided to subjects in the form of a 3g vital gluten bar. Subjects will be 
instructed to ingest the gluten with the same meal every day. Phone calls will be performed weekly 
during the gluten challenge period to: 1) conduct symptom review using the ICDSQ and assign 
severity score, 2) assess compliance with gluten (must be 80% or higher), 3) assess adverse 
events and 4) remind patient of next week’s call. 
 
 
 
 
 
Fig. 1: RC2 data collection and biobanking 
Fig. 2: Vital gluten challenge studies. 90 GFD patients will be challenged for a total of 6-weeks. 3 endoscopies at the 
indicated time points will be performed. For analyses and biobanking see Fig.1 

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c. Longitudinal de-challenge study 
 
In the gluten de-challenge study arm (Figure 3), 90 newly diagnosed celiac disease patients (45 
from University of Chicago; 45 from Mayo Clinic) will be enrolled. Interested patients will be 
asked to sign the informed consent. During a routine endoscopic procedure, 14 additional 
biopsies will be taken and luminal content will be sampled for study purposes. Subjects will 
receive education about the gluten free diet from a registered dietician at University of Chicago 
and Mayo Clinic as part of routine care. A research endoscopy will be performed at 3 months 
after starting the gluten free diet and 12-13 months after starting the gluten free diet. In addition 
to research endoscopies, patients will have serum and urine sampling at baseline, 3 months 
and 12-13 months after starting the gluten free diet. Patients will receive monthly phone calls 
during the study period to check on celiac symptoms and diet adherence.  Additionally, subjects 
will be reminded to fill out the study questionnaires measuring symptoms, quality of life and diet 
adherence.  
 
d. Control. Forty healthy controls (20 from University of Chicago; 20 from Mayo Clinic 
Rochester) will be recruited from the local population by advertisement. After informed consent, 
control subjects will have blood drawn for screening labs.  Qualifying subjects will be asked to 
return to undergo a research endoscopy with biopsies and luminal content sampling, as well as 
saliva, serum and urine sampling. A phone call 1 day after the endoscopy will be performed to 
assess for any adverse events related to the study endoscopy. The CDSD will also be 
administered to ensure to baseline symptoms in this control population. 
 
Controls will be broadly matched to the cases by sex and age (+/- 20 years).  Controls must be 
matched by HLA class II tissue typing.   Tissue typing will be done at the first visit for controls 
only.  If HLA typing does not match celiac patients, the matched control will not continue in the 
study past the first visit. The selection criteria are open to all races and both genders, so long as 
females are not pregnant.  Only subjects with conditions that may have an impact on the aims of 
the study are excluded.  No other exclusions will be applied.  Due to the demographics of celiac 
disease, we do not expect the minority population to exceed 2% black, 2% Asian, 4.5% 
Hispanic, and 4.5% 'other' for an ideal target of 13% minority participation. Due to non-Hispanic 
Whites having the highest prevalence of celiac disease, discrepancies may occur.    
 
e. Gluten Challenge Bar 
The Gluten challenge food article will be made in a commercial kitchen (Mayo Clinic Food 
Services, Gluten Free Kitchen; address: Professional Building Campus, 41st Street NW 
Rochester MN 55904).  Ingredients will be included like common food ingredients. Vital wheat 
gluten will be incorporated into a mix that is otherwise free of gluten.  
Fig. 3: Gluten de-challenge studies. De-challenge studies will be performed in 90 active CeD patients. 3 endoscopies at the 
indicated time points will be performed. For analyses and biobanking see Fig.1 
. 

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This will be a commercial grade food article designed to be palatable (especially important for 
study adherence given daily ingestion for 6 weeks) and to contain 3 grams of gluten.  This 
gluten containing food will have the following characteristics:  
• 
Low in FODMAPs (Fermentable oligosaccharides, disaccharides, monosaccharides and 
polyols) to reduce any IBS like symptoms that FODMAPs might induce symptoms in 
susceptible individuals.  
• 
Prepared in Gluten free kitchen from GF ingredients except for the addition of vital wheat 
Gluten.  
• 
Individually wrapped with nutritional information label 
• 
Controlled for common allergens 
• 
Able to be reduced in size with predictable gluten content in case dose reduction is 
needed. 
• 
Designed to be frozen (up to 3 months with optimal taste and 12 months to food safety) 
for long term storage 
• 
To be thawed before consuming and consume within 4 days of thawing 
 
Bars will be shipped frozen to the University of Chicago.  Bars will be provided frozen to study 
participants. 
 
f. Symptom Diary and Biospecimens  
 
Symptom & Gluten free adherence Surveys 
Symptoms will be assessed using the validated CDSD and ICDSQ. The daily CDSD is a 5-item 
survey that assesses for symptoms of diarrhea, abdominal pain, bloating, nausea and fatigue 
using a 5-point scale. The CDSD will be administered once at time of screening, once at time of 
endoscopy and then daily beginning day 0 through the end of the study period. If subjects rate 
symptoms as “severe” or “very severe”, the research coordinator will discuss with the PI 
regarding continuation in the study.  
 
The ICDSQ is a 14-item survey that is administered on a weekly basis and assesses the impact 
of CeD symptoms on subjects’ quality of life. If the subject reports significant impact of 
symptoms on quality of life and daily functioning, the research coordinator will alert the PI to 
discuss continuation in the study.  Subjects will be asked to fill out the ICDSQ at the end of the 
day.  
 
The CDAT is a 7-question survey assessing gluten free diet adherence.  The CDAT will be 
administered to both the challenge cohort (to evaluate gluten free diet outside of the challenge 
bar) and de-challenge cohort at time of screening, first endoscopy and then at each endoscopy 
visit (day 6 and week 6). 
 
The IGFDQ survey is a 20-item survey covering adherence questions and will be administered 
to the de-challenge cohort to assess adherence to the gluten free diet at each study visit 
(screening, 3 months, and 12 months). 
 
All of the surveys will be administered electronically through RedCap. 
 
Saliva, Blood and Urine samples 

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The challenge and control groups will have blood collected for initial screening labs prior to their 
first endoscopic procedure.  The dechallenge group does not undergo this initial screening lab 
because this is typically already completed as part of routine care.  
 
Saliva blood and urine samples will be obtained at every study visit after that. For the challenge 
cohort, biospecimens are collected prior to gluten challenge (screening period), at 2 hours post 
challenge, 6 days post challenge, and 6 weeks post challenge.  For the de-challenge cohort, 
biospecimens are collected at baseline, 3 months after starting gluten free diet and 12-13 
months after starting gluten free diet. For the control group, biospecimens are collected once at 
the time of endoscope. 
 
Upper Endoscopy 
Three upper endoscopies will be performed for participants in the gluten challenge and de-
challenge cohorts. One endoscopy will be performed in the healthy control cohorts. For the 
gluten challenge cohort, endoscopy is performed at screening, 6 days of gluten challenge, and 6 
weeks of gluten challenge. For the de-challenge cohort, endoscopy is performed at screening 
(routine care), 3 months and 12-13 months after starting the GFD. For the healthy control 
cohort, endoscopy is performed once at baseline.  
 
The upper endoscopies will be carried out by the standard protocol in place at each of the 
medical centers.  Typically, patients will be in the left lateral decubitus position. Two liters of 
oxygen will be given by nasal cannula. Vital signs oximetry and respiration will be monitored 
continuously throughout the procedure. Adults will be given moderate sedation most typically 
with intravenous fentanyl, midazolam with the occasional addition of topical anesthesia or 
diphrenhydramine as per endoscopist’s discretion and history of prior sedation if any. In select 
patients who have a history of difficult sedation or are otherwise deemed to require higher level 
of sedation, monitored anesthesia care (MAC) will be used. Participants will be monitored after 
the procedure for at least 60 minutes and until the following criteria are met: alert and oriented, 
able to protect airway, stable vital signs over the 30 minutes, normal cardiac rhythm, heart rate 
and pulse within 20 points of baseline or within normal limits, oxygen saturation 95% or greater, 
afebrile, ambulation at baseline. The subject must have a responsible adult to provide 
transportation upon dismissal from the unit. Precautions in place for Covid-19 pandemic 
protection will be followed in line with contemporary guidance in place at the time of endoscopy.  
 
Small bowel biopsies 
Research biopsies: Up to14 biopsies will be collected for research purposes. Luminal aspirates 
from the second portion of the duodenum will be obtained during the upper endoscopy.  These 
biopsies will be placed in the requisite media and shipped to the University of Chicago.  
 
Of the research biopsies that will be taken from patients, three to four may be used for 
histopathology, three will be used for analysis of antigen-specific T and B cells studies, one for 
single cell RNA seq, one for high dimensional flow cytometry, one in RNA-later for whole biopsy 
RNA sequencing (RNA-seq), one for preparation of intestinal organoids, and three frozen (one 
for proteomic studies, one for metagenomics, and one for bacterial cultures). The detailed 
information is in the figure below (Figure 4). Of 10 research biopsy samples, 6 biopsy samples 
will be put in the RPMI media, one biopsy in RNA later, three biopsy samples will be frozen 
instantly (snap freezing), and three-four biopsy samples will be placed in formalin. 
 

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g. Metadata will be collected on all study subjects at baseline and after intervention. To 
clinically phenotype CeD patients, we will employ a standardized and validated patient-reported 
symptom tool called the Celiac Disease Symptom Diary (CDSD) that assesses for presence, 
frequency and severity of classical and non-classical symptoms of CeD. The CDSD is the 
current gold standard tool in therapeutic drug trials in CeD based on the fact that it follows FDA 
guidance on patient-reported outcome. CeD patients will be stratified as classical, non-classical 
or asymptomatic according to their CDSD responses. We anticipate that the breakdown of 
clinical phenotypes will be: ~25% classical, ~55% non-classical and ~20% asymptomatic. To 
assess the clinical metabolic profile, we will analyze lipid, vitamin and metal profiles. The 
harmonized clinical data will be entered into a shared REDCap database linked with 
experimental data.  
 
This study will use the following information from subject self-report and medical record 
extraction (note data sharing outside the study team is detailed separately below): 
Name 
MRN 
Race and ethnicity 
Date of Birth 
Medical history  
Medication history 
Lab results (both clinical labs and research lab analysis) 
Pathology reports 
Imaging result 
Endoscopic results 
ICDSQ Data 
CDAT Data 
IGFDQ Data 
Dates of significant clinical events (dates of enrollment, consent, endoscopy procedures, 
diagnosis, lab results, etc) 
 

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h. Histology analysis: Any histology analysis required for routine care will be conducted at the 
respective site according to routine clinical practices. Histology performed for research purposes 
will be processed at the University of Chicago. Histology images will be uploaded in the 
Integrated Web-based clinical research and data management platform and villus height to crypt 
depth ratio (Vh:Cd ) will be quantified centrally in a blinded manner by Dr. Weber at the 
University of Chicago. Unstained sections will be shared with the team for validation 
approaches. 
 
i. Sample collection, analysis, and biobanking: Whole biopsy RNA sequencing (RNA-seq), 
single cell RNA-seq (scRNA-seq), high dimensional flow cytometry and metagneomics will be 
performed on all patients and all time points. Single cell ATACseq (scATAC-seq) for epigenetic 
profiling will be performed on 40 controls, 45 challenge and 45 dechallenge patients at the time 
of the first endoscopy. Analysis of antigen-specific T and B cells will be performed at the 1st and 
2nd endoscopy for the challenge study and the 1st and last endoscopy of the de-challenge study 
(Fig. 1). The workflow is organized in such a way that all experiments will be performed at U 
Chicago. Mayo clinic will be in charge of isolating cells from biopsies. Isolated cells will be fixed 
for scRNA seq, and stained with an antibody panel provided by U Chicago and fixed before 
shipment. Mayo Clinic will also put one biopsy in RNA later for transcriptional profiling and send 
one biopsy to U Chicago for the generation of intestinal organoids that will be stored for future 
studies. In addition, both U Chicago and Mayo Clinic will prepare one biopsy for metagenomics 
analysis and ship DNA to Caltech for analysis.  Core set biobanking will be performed on all 
patients (Fig. 1).  The banking of all samples is described below under each participating 
institution.  
 
Clinical coordinators, technicians and investigators from Mayo Clinic and U Chicago will be 
trained by U. Chicago investigators, and will follow SOPs for sample collection and shipment. 
Mayo Clinic will receive kits with instructions, barcodes, and labeled tubes for sampling and 
shipment. 
 
General sample management will be as follows: 
 
University of Chicago: 
The UofC team will bank blood, urine and tissue from both UofC and Mayo. Any samples 
shared from Mayo will be labelled with the subject’s study number and will not contain any other 
identifiers.  UofC will conduct genetic analysis for study purposes and will store DNA and RNA.  
These samples will be stored indefinitely for future research purposes.  These samples can be 
shared in accordance with the sharing process detailed below. 
 
Mayo Clinic 
The team at Mayo will bank the PBMCs and serum samples for this study. The UofC team will 
process blood samples from UofC subjects and send PBMCs and serum samples to Mayo. 
These samples will be labelled with the subject study code only and will not contain any 
identifiers. Additionally, Mayo will process and store PBMC and serum samples from their own 
site.  These samples will be stored indefinitely for future research purposes.  These samples 
can be shared in accordance with the sharing process detailed below. 
 
CalTec  
The team at CalTec will receive saliva samples and tissue RNA/DNA from UofC and Mayo to 
perform microbiome genetic analysis and for banking purposes. The samples will be labelled 

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with the subject study code only and will not contain any identifiers. Left over tissue and saliva 
samples will be stored indefinitely for future research purposes. These samples can be shared 
in accordance with the sharing process detailed below. 
 
4. Duration 
The study will last for 5 years. 
 
5. Locations and IRB Reliance 
 
All participating centers will reply on the University of Chicago IRB.  Participating centers are 
described below: 
 
Institution 
PI Name 
Activities being conducted 
IRB of Record 
University of Chicago 
 
Hyde Park and River 
East locations 
Sonia Kupfer. MD 
Study sponsor and lead site 
Subject enrollment and all study 
activities 
Sample analysis and banking 
Retention of study data  
 
University of Chicago 
Mayo Clinic 
Joseph Murray, 
MD 
Subject enrollment and all study 
activities 
 
Sample analysis and banking 
 
University of Chicago 
California Institute of 
Technology (CalTec) 
 
Rustem 
Ismagilov, PhD 
Sample analysis and banking only 
(no subject enrollment) 
 
University of Chicago 
 
 
 
6. Experimental Subjects  
 
a. number of subjects 
To understand the mechanisms underlying tissue destruction and healing we have designed 
longitudinal challenge and de-challenge studies with 90 patients in the challenge arm (45 from 
University of Chicago; 45 from Mayo Clinic Rochester) and 90 patients in the de-challenge arm 
(45 from University of Chicago; 45 from Mayo Clinic Rochester). In addition, 40 MHC-matched 
healthy controls (20 from University of Chicago; 20 from Mayo Clinic Rochester) will be enrolled. 
 
b. method of subject selection and recruitment 
The clinical research associate (CRA) will identify eligible subjects via IRB-approved HIPAA-
compliant methods. The CRA will review clinic and endoscopy schedules as well as the electronic 
medical record system for potentially eligible subjects.   
 
The CRA will contact potential subjects in clinic or by telephone to explain the study, to review 
eligibility criteria and obtain informed consent.  CRAs will talk with interested potential subjects to 
provide complete disclosure of the study objectives and procedures, potential risks and benefits, 
and expectations and then document the process of written consent obtained from subjects. The 
CRAs will be responsible for all aspects of protocol logistics for recruited subjects. This includes 
obtaining required demographic and questionnaire data, handling of all collected biospecimens, 
and ensuring smooth completion of the study requirements.   
 

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Each potential subject screened for eligibility will be identified only by sequential number, contact 
methods, and reason for recruitment failure or success. Recruitment numbers will be reported 
monthly to the site PIs to monitor accrual. 
 
There is no randomization in this study. 
 
c. Inclusion and exclusion criteria: 
 
c.1. Challenge study:  
Inclusion Criteria:  
Subjects must fulfill all of the following criteria at screening to be eligible for participation: 
1. Males or females 18 to 70 years of age, inclusive. 
2. Subjects demonstrate willingness to participate in the study and to perform all required 
procedures as documented by signed informed consent. 
3. Subjects must have a diagnosis of CeD by intestinal biopsy at least 12 months and positive 
celiac serology at some point in their evaluation, prior to screening as confirmed by medical 
records or written physician statement. 
4. Subjects must have reported following a strict GFD for at least the 12 consecutive months 
and must be willing to maintain their current diet for the duration of study participation while 
being in gluten challenge.  
5. Patients who don’t show mucosal healing with a VH:CD ratio <2 at the baseline endoscopy 
will enter the GFD non-healed cohort (see below) and will not undergo gluten challenge.   
Ninety percent of patients who are asymptomatic, adhere to GFD and are seronegative have 
achieved histologic healing, 
 
Exclusion Criteria: 
Subjects are excluded from the study if any of the following criteria applies: 
1. Current diagnosis of any severe complication of CeD, such as refractory CeD (RCD) type I or 
II, enteropathy-associated T-cell lymphoma (EATL), ulcerative jejunitis, or GI perforation. 
2. Diagnosis of any chronic, active GI disease other than CeD, such as active, untreated peptic 
ulcer, eosinophilic esophagitis, erosive esophagitis, ulcerative colitis or Crohn’s disease, 
microscopic colitis, irritable bowel syndrome, small intestinal bacterial overgrowth, tropical 
sprue, or other GI and non-GI disorder or prior GI surgery that may, in the Investigator’s 
opinion, interfere with the assessment of symptoms of abdominal pain, diarrhea, or other 
components of CeD. 
3. Selective IgA deficiency, defined as having undetectable levels of serum IgA. 
4. History of severe reaction to gluten exposure that is considered incapacitating or life-
threatening. 
5. Known or suspected exposure to COVID-19 infection in the 4 weeks before the screening. 
6. History or presence of any clinically significant disease that, in the opinion of the Investigator 
may confound the subject’s participation and follow-up in the clinical trial or put the subject at 
unnecessary risk, including but not limited to: uncontrolled hypertension (BP ≥180/110 
mm/Hg), unstable angina, Class II congestive heart failure or major fluid overload, coronary 
angioplasty or myocardial infarction within the past 6 months, clinically significant arrhythmias 
or electrocardiogram (ECG) abnormalities, severe chronic pulmonary disease, or any renal, 
hematologic, GI, immunologic, dermatologic, neurologic, or psychiatric disease. 
7. History of significant substance or alcohol abuse during the 12 months prior to screening as 
obtained by medical record and/or subject report. 
8. Females who are pregnant or planning to become pregnant during the study period, or who 
are currently breastfeeding. 

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9. Participation in another investigational drug or device study or treatment with an 
investigational drug within 30 days or 5 half-lives, whichever is longer, prior to screening. 
10. Individuals with uncontrolled clotting disorders or on anti-coagulants that they can’t safely 
hold prior to endoscopy.  
11. Participation in clinical trial within 30 days prior to consenting, and 12 months for a clinical 
trial including a biologic agent.  
12. Participation in clinical trial with tolerogenic agents  
 
c.2. De-challenge study:  
Inclusion criteria: 
Subjects must fulfill all of the following criteria at screening to be eligible for participation: 
1. Males or females, 18 to 70 years of age, inclusive. 
2. Subjects demonstrate willingness to participate in the study and to perform all required 
procedures as documented by signed informed consent. 
3. Newly suspected subjects of CeD, either typical symptoms of CeD or referred to celiac clinic 
due to positive CeD serology undergoing upper endoscopy. Patients with weak positive 
serology will not be included in order to maximize the likelihood of the patients qualifying for 
the dechallenge study.      
4. Subjects who are currently not on GFD. 
 
Exclusion Criteria: 
1. Has a history of chronic inflammatory gastrointestinal disease (example, inflammatory bowel 
disease, extensive colitis, ulcerative jejunitis, drug induced enteropathy) (Microscopic colitis 
is not an exclusion criteria). 
2. Has chronic infectious gastrointestinal illness, or acute infectious gastrointestinal illness within 
the 4-week period prior to screening. 
3. Known history of lymphoproliferative disease, including monoclonal gammopathy of unknown 
significance, lymphoma, or signs and symptoms suggestive of possible lymphoproliferative 
disease, such as lymphadenopathy and/or splenomegaly. 
4. Individuals with uncontrolled clotting disorders or on anti-coagulants that they can’t safely 
hold prior to endoscopy. 
5. Known or suspected exposure to COVID-19 infection in the 4 weeks before the screening. 
6. History or presence of any clinically significant disease that, in the opinion of the Investigator 
may confound the subject’s participation and follow-up in the clinical trial or put the subject at 
unnecessary risk, including but not limited to: uncontrolled hypertension, unstable angina, 
Class II congestive heart failure or major fluid overload, coronary angioplasty or myocardial 
infarction within the past 6 months, clinically significant arrhythmias or ECG abnormalities, 
severe chronic pulmonary disease, or any renal, hematologic, GI, immunologic, dermatologic, 
neurologic, or psychiatric disease. 
7. History of significant substance or alcohol abuse during the 12 months prior to screening as 
obtained by medical record and/or subject report. 
9. Participation in another investigational drug or device study or treatment with an 
investigational drug within 30 days or 5 half-lives, whichever is longer, prior to screening. 
11. Participation in clinical trial within 30 days prior to consenting, and 12 months for a clinical 
trial including a biologic agent.  
12. Participation in clinical trial with tolerogenic agents  
13.  Participants whose initial biopsies do not reveal villous atrophy will not be followed in the 
de-challenge group but may be recruited into the potential celiac disease cross sectional 
group so long as they meet the criteria for inclusion into that group. 
 
 

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c.3. Controls: 
Inclusion criteria: 
• 
Subjects (both cases and controls) will include males and non-pregnant females, aged 18-
70 years.  
• 
Females of reproductive age or capacity (i.e. not having had tubal ligation or sterilization) 
will have a pregnancy test performed. 
 
Exclusion criteria: 
• 
Subjects on antibiotics, proton pump inhibitors, aspirin, non-steroidal anti-inflammatory 
drugs, alcohol intake within 48 hours, a bowel preparation within 4 weeks of the studies, 
and smokers 
• 
Subjects will be asked not to take any probiotics in the week before testing.  
• 
Any known intestinal inflammation such as GERD, eosinophilic esophagitis, and 
inflammatory bowel disease. 
• 
Prior gastrointestinal surgery (other than appendectomy) 
• 
Ongoing use of antiplatelet agents or anticoagulants. 
• 
Controls should not have a prior history of or family history of CD. 
• 
Subjects unable to provide informed consent 
• 
The presence of any medical or psychological condition could interfere with the safe 
performance of the upper endoscopy. 
 
7. Statistical analysis  
 
Challenge studies 
a. Developing predictive models of tissue destruction. We will perform challenge studies 
(Fig. 2), which will provide unique insights into the dynamic of the host response and microbial 
components associated with tissue destruction. Under this experimental setting, patients will 
serve as their own control as they will undergo 3 endoscopies over the course of the 
interventional study. The flow-cytometry and scRNAseq analysis preceding gluten challenge will 
determine the frequency of gluten-specific T cells and the state of IECs at baseline. The 6-day 
analysis will give insights into the early antigen-specific T and B cells responses (quantitative 
and qualitative by combining scRNA-seq on fresh biopsies, flow cytometry and scRNA-seq on 
antigen-specific T and B cell responses).  Secondly, we will investigate how early response 
impacts IECs, e.g. by promoting expression of non-classical MHC class I molecules MICA and 
HLA-E that activate NKG2D and CD94/NKG2C NK receptors expressed on cytotoxic IELs (IE-
CTLs), respectively, and consequently affect the cytolytic potential and activation status of IE-
CTLs that in turn mediate IECs destruction. We will link this initial response to the development 
of VA (we expect ∼60% of patients to develop VA). This will determine whether the frequency 
and/or intensity of the anti-gluten T cell response drives tissue destruction. We will also 
determine effects that occur early in IECs after gluten challenge (6 days) and if these lead to 
persistent changes in stem cells that then manifest in specified conditions or lineages. We will 
compare single cell transcriptomes with the single cell atlas from biopsies from patients that 
respond to gluten with damage (60-70%) versus those that do not. Thus, we can determine the 
role of IECs in driving disease (not just as a passive target) prior to their damage. Furthermore, 
we can identify which alterations in other immune subsets (such as γδ T cells and 
macrophages) promote or prevent changes in IECs, IELs activation and tissue destruction. 
Finally, we will assess whether microbial composition and function (metagenomic and 
metabolomic analysis) preceding gluten challenge influences activation of gluten specific T 
cells, and if early microbial changes are linked to IE-CTLs activation and ultimately VA. To 
determine the potential impact of the microbiota on tissue destruction and healing (see de-

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challenge study below), we will analyze mucosal-associated and luminal bacteria as described 
in section e. The role of the microbiota in tissue destruction and healing is complex and needs 
to be integrated with the host response. These microbes could impact tissue destruction (and 
healing, see below) via three mechanisms: (i) Microbes could compete with or interfere with 
host nutrient uptake (e.g. by deconjugating and modifying bile acids), driving various metabolic 
deficiencies and therefore further weakening host antimicrobial defenses and contributing to 
tissue damage or interfering with tissue healing. In addition, they could produce metabolites 
(e.g. butyrate or modified bile acids) that promote or prevent tissue destruction  by modulating 
immune responses; (ii) Pro-inflammatory  or immunomodulatory  molecules  (e.g. LPS or 
TLR2/1 ligands or polysaccharide A, respectively), especially from mucosa-associated bacteria, 
can drive inflammatory or tolerogenic immune responses, that in turn promote or hamper tissue 
destruction. Persistence of adherent microbes after gluten removal could impact IECs function 
and continue to drive inflammation and prevent healing or lead to ongoing gastrointestinal 
symptoms. (iii) Alternatively, some microbes could be beneficial—either by outcompeting the 
inflammatory microbes or by changing how gluten is processed.   
 
b. Genetic determinants of susceptibility to CeD and variation in tissue destruction. 
Genetics can be a powerful tool to identify new mechanisms associated with the development of 
VA. More than 200 loci are associated with CeD susceptibility, but mechanisms associated with 
these loci remain largely undetermined. We hypothesize that the functional impact of many of 
these GWAS variants may only be revealed in the context of active CeD. As a first step towards 
testing that hypothesis, we performed eQTL mapping on duodenal biopsies derived from 41 
active CeD, 61 controls and 39 GFD patients for which we also had genotype data available for 
over 3 million single-nucleotide polymorphisms genome-wide. Across all three groups, >20% of 
genes expressed in duodenum samples were associated with at least 1 significant eQTL 
(referred to as eGenes), stressing the pervasive role of genetics to interindividual variation in 
gene expression levels.  Interestingly, and consistent with our hypothesis, we found 388 
eGenes  that are only identifed among active CeD patients and not in the control and GFD 
group. These genetic associations would have been missed when only looking at healthy 
individuals, underlining the importance of studying regulatory processes in disease contexts to 
improve functional interpretation of as-yet unexplained GWAS loci and the mechanism of 
disease. Interestingly, we also found a large number of eGenes (n=580) only detectable in the 
GFD condition, supporting the notion that healing is associated with significant tissue 
remodeling that leads to sustained changes to the immune and regulatory landscape of the gut. 
In this RC2, we will significantly expand on these findings by characterizing eQTLs in the 
different cellular subsets identified by scRNA-seq. Specifically, the scRNA-seq data collected in 
the interventional cohorts will be paired with genotype data from the same individuals to 
generate the most comprehensive catalog to date of genetic variants that impact gene 
expression (eQTL) in IECs and the different subsets. This unique resource will directly inform on 
the mechanisms associated with known GWAS variants, and enable development of polygenic 
risk scores to stratify patients along a susceptibility continuum for developing severe VA. 
 
De-challenge studies 
a. Developing predictive models of tissue healing. We will perform de-challenge studies 
(Fig. 3), which will identify the interaction between the host response and microbial components 
and the pathways associated with variation in mucosal healing. As for the challenge studies, 
patients will serve as their own control as they will undergo 3 endoscopies over the course of 
the interventional study. The host and microbial analysis performed on samples obtained at the 
time of the initial endoscopy will allow to assess alterations present at the time of active CeD. 
The nature and intensity of these alterations can then be linked to the presence or absence of 
healing at 3 and 12 months after gluten exclusion.  The 3-month time point will allow assessing 

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the early changes with gluten withdrawal. We expect that only 20% of patients will have healed 
by that time point. However, at this time, we will be able to detect changes in IECs that are 
secondary to loss of immune activation. We hypothesize that early changes in IECs may predict 
presence or absence of healing at 12 months (time of the third endoscopy). scRNA seq analysis 
of isolated IECs will allow to determine whether the transcriptional program of adaptive IECs 
(aVECs) and/or intestinal stem cells (ISCs) at the time of active CeD can be linked to the 
capacity or lack of capacity of the intestinal mucosa to heal. Intestinal organoids generated from 
IECs isolated from the 1st and last endoscopy will be biobanked and serve for future mechanistic 
studies. Furthermore, hypotheses on the mechanistic basis for IEC-immune-microbiome cross-
talks preventing healing on a GFD will be generated as changes in microbiota and some 
transcriptional signatures will be associated and could predict the capacity of the tissue to heal. 
 
b. Genetic determinants of susceptibility to CeD and variation in tissue healing. Genetic 
analyses and associations will be performed as in aim 1.2. to characterize genetic variants that 
impact gene expression of subsets of IECs modulating their lineage specification and 
regenerative capacity.   
 
Research resources 
This will be led by Dr. J. Johnson, co-investigator, and her Clinical Research Informatics (CRI) 
team, in collaboration with the RC2 PIs and co-PIs (Drs. Barreiro and Matthew), and Mine Cicek 
(biobank Mayo Clinic). 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Description: The proposed RC2 will include coordinated collection of a diverse set of 
biosamples and datasets from Mayo Clinic, Caltech and U Chicago. It includes metadata from 
biobanks, clinical data, sequence data, and analysis results for a wide range of omic datatypes. 
To integrate such large and complex datasets and enable multi-omics analysis, we will build an 
integrated clinical research data management and informatics platform that will support 
harmonization, curation, dissemination, and analysis of the data collected at various levels (Fig. 
4). This integrated platform will be developed and hosted at the data coordinating center at 
UChicago. It will include the following components: 1) Local Biobanks (Mayo Clinic & U 
Chicago) and Virtual Biobank: While each of the two sites biobanks will have components of 
the RC2 samples, we will implement a virtual biobank in close collaboration with the site 
biobanks. This virtual biobank will centralize detailed metadata characterizing the biosamples 
Fig. 4  An integrated, Web-based clinical research data and biobank 
management and informatics platform with data integrated from 
biobanks, clinical sites, sequencing cores, and omics analyses. 

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via an automated sync from each biobank. We will implement project specific identifiers that will 
be used as a cross-reference between various components and datatypes within the platform 
including the virtual biobank and the clinical study database described later. The virtual biobank 
will be a key component of the platform and will provide a powerful interface for an enhanced 
access to the metadata and sample tracking information; 2) Clinical Data: Research 
coordinators from both clinical sites will upload all the relevant clinical data in a central study 
database that will be designed and implemented using REDCap; 3) Sequence & Analysis 
Results: For all the various classes of sequencing performed during the RC2, we will develop a 
central repository of raw sequence data. We will implement analytical pipelines for all sequence 
data types and the results of analyses will be modeled and stored in a searchable multi-omics 
database; 4) WebUI: The proposed platform will include two Web applications – a) Public Web 
application that will allow users to search and access all de-identified data, b) A robust Internal 
Web application with multi-faceted search capabilities, cohort creation capabilities, detailed 
sample tracking, and access to raw sequence data and analyses results; 5) Multi-omics 
analysis: An innovative Web-based multi-omics module will support correlations between multi-
omics datasets and clinical parameters of interest. It will include robust visualization of results 
for the results of the various omics data analyses. 
 
8. Potential risks and benefits to subjects 
a. Potential Risks:  
The potential risks to study participants include those related to blood draws, upper endoscopy 
and/or those related to gluten challenge (for the “challenge” cohort).  
 
Blood Draw Risks 
Blood drawing could result in temporary slight discoloration of the skin after blood draws.  
 
Endoscopy Risks 
For participants undergoing endoscopy, the primary risks are related to unexpected sedation 
medication reactions (e.g., cardiopulmonary instability), aspiration, infection, perforation and 
bleeding. Based on ASGE guidelines, upper GI endoscopy carries low risk of adverse events. 
Large series report adverse event rates of 1 in 200 to 1 in 10,000 and mortality rates from none 
to 1 in 2000. Higher risk patients and higher risk procedures will be excluded in this study thereby 
minimizing risk.  
 
Gluten Challenge Risks 
Participants in the gluten challenge cohort could experience a number of GI or extra-intestinal 
symptoms with gluten ingestion. These symptoms could include abdominal pain, bloating, 
nausea, vomiting, diarrhea, constipation, headache, and fatigue. Based on our personal 
experience and a recent trial, CeD patients generally have mild to moderate symptoms on 
gluten with only a small number (~1%) requiring dose reduction or stopping the challenge. If 
subjects report symptoms as “severe” or “very severe” on the CDSD on any day, the research 
coordinator will alert the PI to review ongoing participation in the study vs. possible dose 
reduction.  
 
Loss of Confidentiality: 
As this study involves the use of subject identifiers and health information, there is a potential risk 
of loss of confidentiality. 
 
 
 
 

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b. Protection Against Risks:  
 
Recruitment and Informed Consent  
Prior to initiating the study, all subjects will provide written informed consent using forms approved 
by the Institutional Review Boards. The consent form becomes a permanent part of the medical 
record at each site participated. The consent form includes information relative to genetic studies 
and storage of genetic material for future research, as well as conforms with the high standards 
required for such studies. Since this genetic information is not yet pertinent to clinical practice, 
the information will not be included in the medical record; however, this information will be 
maintained in a coded fashion that excludes patient identifiers.  
 
Loss of Confidentiality 
There is the potential for loss of confidentiality. In order to minimize this risk, each subjects’ 
intestinal samples will be assigned a random unique research ID number and barcode so that 
anonymity is preserved according to HIPPA regulations. Only the PI and the research team will 
have access to the file that links PHI to the study ID number and barcode. Any documents 
containing patient identifiers will be kept in locked files in the PI’s office. Computer documents 
with PHI will be protected by passwords known only by the PI and research team 
 
Blood Draw Risks 
Blood will be drawn by study personnel trained in phlebotomy and using standard practices. 
 
Endoscopy Risks 
Endoscopy and tissue sampling will be performed by GI physicians experienced in endoscopic 
procedures, using standard endoscopic and tissue sampling methods.  Inclusion and exclusion 
criteria insures that higher risk patients are excluded from this study. 
 
Gluten Challenge Risks 
The symptom severity tool will help guide whether a subject should be considered for study 
discontinuation. The CDSD is collected electronically using a secure RedCAP form that the 
study team will monitor regularly. Also subject symptoms will be monitored during all follow up 
phone calls. The study team will notify the PI regarding any subject who experiences severe 
symptoms. This will be assessed using the CDSD and any symptoms noted as “severe” or “very 
severe” will be discussed with the PI to determine ongoing participation in the study vs. possible 
dose reduction. 
 
DSMP 
A data and safety-monitoring plan (DSMP) has been established for this trial. Reporting 
mechanisms will be put in place for adverse events to the IRB and the NIH. An independent 
Data Safety and Monitoring Board will be appointed to oversee the trial as described in the 
application. This is described in section 9 below. 
 
c. Potential Benefit to the Subjects and Others 
The potential benefits to all participants include close assessment of diet and nutrition during the 
course of study. Participants in the de-challenge cohort will also have endoscopic assessments 
of mucosal healing and provide useful information for assessment of response to the gluten free 
diet. 
 
 
 

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d. Importance of the Knowledge to Be Gained 
The indirect benefits are increased insight into the pathogenesis and heterogeneity of CeD. 
Specifically, this knowledge will improve our understanding of epithelial and microbial factors that 
either trigger CeD or promote patient susceptibility to this condition. In addition, the knowledge 
acquired through this study may allow for the future development of new risk biomarkers and 
interventions to prevent the development and/or treatment of CeD. 
 
9. Monitoring of safety of subjects  
 
Adverse Event Monitoring in Subjects: 
Subjects will be monitored throughout the study for adverse events related to the gluten 
challenge, blood draws and endoscopy. The research coordinator will review daily CDSD for 
any subjects reports “severe” or “very severe” symptoms on any day during the gluten 
challenge. These will be discussed with the PI and determination made regarding ongoing 
participation vs. possible dose reduction in the amount of gluten. Any adverse events possibly 
related to blood draws or endoscopy will be discussed with the PI immediately as outlines 
below. 
 
Adverse Event (AE) Reporting: 
Any problematic or unfavorable medical occurrence in a human subject, including any abnormal 
sign (e.g. abnormal physical exam or laboratory finding), symptom, or disease, temporally 
associated with the subject’s involvement in the research, whether or not considered related to 
participation in the research. 
 
Serious Adverse Event (SAE): 
Any adverse event that: 
 
Results in death 
 
Is life threatening, or places the subject at immediate risk of death from the event as it 
occurred 
 
Requires or prolongs hospitalization 
 
Causes persistent or significant disability or incapacity 
 
Results in congenital anomalies or birth defects 
 
Causes cancer 
 
Is an important medical event  
Important medical events are those that may not be immediately life threatening, but are clearly 
of major clinical significance.  They may jeopardize the subject, and may require intervention to 
prevent one of the other serious outcomes noted above.  For example, drug overdose or abuse, 
a seizure that did not result in in-patient hospitalization, or intensive treatment of bronchospasm 
in an emergency department would typically be considered serious. 
 
Unanticipated Problem: 
Any incident, experience, or outcome that meets all of the following criteria: 
 
Unexpected, in nature, severity, or frequency, considering the research procedures 
described in this protocol and characteristics of the study population; 
 
Related or possibly related to participation in the research, meaning there is a 
reasonable possibility that the incident, experience, or outcome may have been caused 
by the study procedures and/or use of Montelukast; 
 
Is serious in nature or otherwise suggests that the research places subjects or others at 
a greater risk of harm (including physical, psychological, economic, or social harm) than 
was previously known or recognized. 
 

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Severity 
The severity of AEs will be assessed according to the following grading scale.  Adverse Events 
will be classified according to severity, expectedness and potential relatedness to the study 
drug. 
 
Mild: Awareness of signs or symptoms, but easily tolerated and are of minor irritant type, 
causing no loss of time from normal activities. Symptoms do not require therapy or a 
medical evaluation; signs and symptoms are transient.  
 
Moderate: Events introduce a low level of inconvenience or concern to the subject and 
may interfere with daily activities, but are usually improved by simple therapeutic 
measures; moderate experiences may cause some interference with functioning.  
 
 Severe: Events interrupt the subject’s normal daily activities and generally require 
hospitalization. 
 
Expectedness 
AEs must be assessed as to whether they were expected to occur or unexpected, meaning not 
anticipated based on current knowledge of procedures and drugs involved in the research 
study. Categories are:  
 
Unexpected - nature or severity of the event is not consistent with known information 
about Montelukast or other study procedures.  
 
Expected - event is known to be associated with Montelukast or other study procedures. 
 
Adverse Event Tracking 
Adverse events will be gathered during each study and phone visit, as well as any spontaneous 
report occurring between scheduled study calls/visits.  All adverse events will be evaluated and 
graded by the PI, and will be tracked in the study RedCap database.  
 
Reporting Adverse Events 
Adverse events will be reported to the IRB according to the IRB’s reporting policy.  AE data will 
be provided to the DSMB for analysis prior to all formal meetings, and more urgently in case of 
serious adverse events that are deemed related or possibly related to the study. 
 
10. Data and Safety Monitoring 
 
Multi-Center Coordination  
The study teams at each site will conduct weekly research meetings to review recruitment, 
biospecimen collection, and any safety issues. A monthly meeting will be held with study 
personnel at University of Chicago and Mayo Clinic Rochester to review overall recruitment, 
biospecimen collection and any safety issues. 
 
Data and Safety Monitoring Board (DSMB): 
The Principal Investigator (PI) along with site PIs and study personnel will be responsible for 
ensuring participants’ safety on a daily basis. The Data and Safety Monitoring Board (DSMB) 
will act in an advisory capacity to monitor participant safety, evaluate the progress of the study, 
to review procedures for maintaining the confidentiality of data, the quality of data collection, 
management, and analyses. The overall study PI is responsible for distributing adverse event 
reports and coordination across multiple sites. DSMB communications will be shared on a 
quarterly basis with all sites involved in the trial. 

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Frequency of Data and Safety Monitoring  
The DSMB will meet twice annually by teleconference call to review study progress, data 
quality, and participants safety. Safety reports are sent to the site PI and PO at least twice a 
year and will include a detailed analysis of study progress, data and safety issues. 
Data Analysis and Coordination 
Data analysis and coordination will be performed by study personnel (but not the PI). All data 
from this trial will be directly available to the DSMB on request except raw data identifying 
individual participants. Since this is not an intervention trial, participants will not be notified of 
specific trial results. Any serious adverse events will be communicated with the subjects’ health 
providers with appropriate information from the trial. 
Content of Data and Safety Monitoring Report  
The content of the data and safety monitoring report will include: study status, participant 
descriptive information, safety information, study quality. 
DSMB Membership and Affiliation   
The following individual(s) are proposed members of the DSMB. Should there be any questions 
regarding the independence of the DSMB, it will be addressed and corrected if necessary at that 
time.  
 
Julian Solway, MD 
Professor of Medicine, University of Chicago 
Dr. Solway is Professor of Medicine, Dean for Translational Medicine and Director of the 
Institute for Translational Medicine. Dr. Solway is experienced in clinical and translational 
science and will provide important oversight regarding patient safety. 
 
Christina Ciaccio, MD 
Associate Professor of Medicine, University of Chicago 
Dr. Ciaccio is the Section Head of Allergy and Immunology with expertise in allergic disorders 
including those related to food antigens. Dr. Ciaccio has been involved in a number of large 
drug trials in food allergy and will provide important oversight regarding patient safety. 
 
Konstantinos A. Papadakis, MD 
Professor of Medicine, Mayo Clinic Rochester 
Dr. Papadakis is an adult gastroenterologist with expertise in intestinal disorders including celiac 
disease and inflammatory bowel diseases. Dr. Papadakis will provide important oversight 
related to gluten challenge and endoscopic safety. 
 
Vandana Nehra, MD 
Associate Professor of Medicine, Mayo Clinic Rochester 
Dr. Nehra is an adult gastroenterologist with expertise in intestinal disorders including celiac 
disease. She will provide important oversight related to gluten challenge and endoscopic safety. 
 
Conflict of Interest for DSMB 
Each DSMB member will sign a Conflict of Interest Statement which includes current 
affiliations, if any, with pharmaceutical and biotechnology companies (e.g., stockholder, 
consultant), and any other relationship that could be perceived as a conflict of interest related to 
the study and / or associated with commercial interests pertinent to study objectives.  

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Protection of Confidentiality  
Data will be presented in a blinded manner during the open sessions of the DSMB. At DSMB 
meetings, data and discussion are confidential. Participant identities will not be known to the 
DSMB members. 
 
DSMB Responsibilities 
The responsibilities of the DSMB include:  
• Review the research protocol, informed consent documents and plans for data safety and 
monitoring; 
• Recommend subject recruitment be initiated after receipt of a satisfactory protocol; 
• Evaluate the progress of the trial, including periodic assessments of data quality and 
timeliness, recruitment, accrual and retention, participant risk versus benefit, performance of 
the trial sites, and other factors that can affect study outcome; 
• Consider factors external to the study when relevant information becomes available, such as 
scientific or therapeutic developments that may have an impact on the safety of the 
participants or the ethics of the trial; 
• Review study performance, make recommendations and assist in the resolution of problems 
reported by the PI; 
• Protect the safety of the study participants; 
• Report to NIH on the safety and progress of the trial; 
• Make recommendations to the NIH and the PI concerning continuation, termination or other 
modifications of the trial based on the observed beneficial or adverse effects of the treatment 
under study; 
• Ensure the confidentiality of the study data and the results of monitoring; and, 
• Assist the NIH by commenting on any problems with study conduct, enrollment, sample size, 
and/or data collection.  
 
11. Payment to subjects:  
 
Challenge Subjects: 
Subjects will be compensated $300 after each endoscopy for a potential total of $900.  If a 
subject screen fails based on lab results before the first endoscopic procedure, they will be paid 
$50 for the blood draw visit, and then withdrawn from study. 
 
Dechallenge Subjects: 
Subjects will be compensated $300 after each endoscopy for a potential total of $900.   
 
Control Subjects: 
Subjects will be compensated $300 for a single endoscopy.  If a subject screen fails based on 
lab results before the first endoscopic procedure, they will be paid $50 for the blood draw visit, 
and then withdrawn from study. 
 
12. Informed Consent 
Subjects will undergo an initial screening research study visit. Consent will be obtained during 
this initial screening visit before any research-related procedures take place. Ample time will be 
given for consent review and questions.   
 
 
 

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13. Data and Sample Confidentiality and Sharing 
 
Study data will be entered into a password protected REDCap database maintained on a HIPAA 
compliant, secure server hosted at the University of Chicago.  All data will be entered and 
stored using the subjects unique study ID.  The file that links the study ID to the subject’s 
identity will be kept separately on password protected, HIPAA compliant, encrypted computers 
at the respective sites.  The study data set will contain the dates of significant clinical events but 
will not contain any other subject identifiers. 
 
Study samples will be labelled only with the subject’s unique study ID and will not contain any 
other identifying information.  
 
Plans for Sharing 
The data and samples collected as part of this study will be banked and stored for future 
research. These material may be shared with other researchers with approval of the advisory 
committee (Drs. Jabri, Kupfer, Murray, Ismagilov, Abadie and Barreiro) and with proof of 
necessarily ethic review board approvals. Prioritization will be made depending on the specific 
request and availability of biobanked samples as well as the scientific merit. Data and samples 
will be shared using the subject’s unique study ID only.  Data may contain dates, but will not 
contain any other identifying information. 
 
Data and samples will be also shared as required through the NIH. Data shared under this 
requirement be shared using the subject’s unique study ID only.  Data may contain dates, but 
will not contain any other identifying information. 
 
14. Recruiting methods  
 
Participants will be recruited primarily through the adult GI clinics at the University of Chicago 
and Mayo Clinic Rochester. The study team will also place an advertisement on the local 
institution’s celiac center website (UofC - http://www.cureceliacdisease.org/). Additionally, 
subjects who have agreed to be contacted for future studies may be recruited through the celiac 
database at the University of Chicago (IRB #16103A - patients would be contacted via phone or 
email about the study).  All patients will be screened for baseline eligibility by medical record 
review, and enrolled if they meet all inclusion and exclusion criteria.  
 
Recruitment could include employees at the participating institutions. To minimize any risk of 
coercion, all recruitment and consenting activities with employees will be conducted by a study 
coordinator and will not involve any faculty from the study team. 
 
15. Notification of subject's primary physician  
The subject’s primary care physician will be notified by electronic message through EPIC. If the 
primary care physician is outside UCM, a phone call and letter will be sent. 
 
16. Bibliographic references  
 
1) Khan, M. R. et al. The Risk of Autoimmune Disorders in Treated Celiac Disease Patients 
in Olmsted County, Minnesota. J Pediatr Gastroenterol Nutr 69, 438-442, 
doi:10.1097/MPG.0000000000002418 (2019). 
2) Lindfors, K. et al. Coeliac disease. Nat Rev Dis Primers 5, 3, doi:10.1038/s41572-018-0054-

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z (2019). 
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4) Sollid, L. M. & Jabri, B. Is celiac disease an autoimmune disorder? Curr Opin Immunol 
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doi:10.1016/S0140- 6736(17)31796-8 (2018). 
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doi:10.1053/j.gastro.2017.02.002 (2017). 
7) Taylor, D. K., Miller, S. S. & Moltz, K. A. Adolescent health care: an assessment of 
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8) Sulkanen, S. et al. Tissue transglutaminase autoantibody enzyme-linked immunosorbent 
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9) Walker, M. M. et al. Detection of celiac disease and lymphocytic enteropathy by parallel 
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10) Abdulkarim, A. S., Burgart, L. J., See, J. & Murray, J. A. Etiology of nonresponsive celiac 
disease: results of a systematic approach. Am J Gastroenterol 97, 2016-2021, 
doi:10.1111/j.1572-0241.2002.05917.x (2002). 
11) Rubio-Tapia, A. et al. ACG clinical guidelines: diagnosis and management of celiac 
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12) Tennyson, C. A., Simpson, S., Lebwohl, B., Lewis, S. & Green, P. H. Interest in medical 
therapy for celiac disease. Therap Adv Gastroenterol 6, 358-364, 
doi:10.1177/1756283X13492580 (2013). 
13) Tomal, J., McKiernan, D., Guandalini, S., Semrad, C. E. & Kupfer, S. Celiac patients' 
attitudes regarding novel therapies. Minerva Gastroenterol Dietol 62, 275-280 (2016). 
14) Lebwohl, B. et al. Mucosal healing and mortality in coeliac disease. Aliment Pharmacol 
Ther 37, 332- 339, doi:10.1111/apt.12164 (2013). 
15) Lebwohl, B., Michaelsson, K., Green, P. H. & Ludvigsson, J. F. Persistent mucosal 
damage and risk of fracture in celiac disease. J Clin Endocrinol Metab 99, 609-616, 
doi:10.1210/jc.2013-3164 (2014). 
16) Rubio-Tapia, A. et al. Mucosal recovery and mortality in adults with celiac disease after 
treatment with a gluten-free diet. Am J Gastroenterol 105, 1412-1420, 
doi:10.1038/ajg.2010.10 (2010). 
17) Rostom, A., Murray, J. A. & Kagnoff, M. F. American Gastroenterological Association 
(AGA) Institute technical review on the diagnosis and management of celiac disease. 
Gastroenterology 131, 1981-2002, doi:10.1053/j.gastro.2006.10.004 (2006). 
18) Institute, A. G. A. AGA Institute Medical Position Statement on the Diagnosis and 
Management of Celiac Disease. Gastroenterology 131, 1977-1980, 
doi:10.1053/j.gastro.2006.10.003 (2006). 
19) Ludvigsson, J. F. et al. Diagnosis and management of adult coeliac disease: guidelines 
from the British Society of Gastroenterology. Gut 63, 1210-1228, doi:10.1136/gutjnl-2013-
306578 (2014). 
20) Silvester, J. A. et al. Tests for Serum Transglutaminase and Endomysial Antibodies Do 
Not Detect Most Patients With Celiac Disease and Persistent Villous Atrophy on Gluten-
free Diets: a Meta-analysis. Gastroenterology 153, 689-701 e681, 
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Challenge study  
 
Screening  
Endoscopy 
visit 
(screening 
part 2) (2)  
Day 0  
Day 6  
Week 6  
Informed 
Consent  
X 
 
 
 
 
Blood 
collection  
X  
X 
X (1) 
X 
X 
Gluten 
dispense (7, 
8)   
 
 
X 
X 
 
Endoscopy 
 
X  
 
X 
X 
Biopsies  
 
X 
 
X 
X 
Luminal 
content  
 
X 
 
X 
X 
Vital signs 
X 
 
 
 
 
Physical 
Exam  
X 
 
 
 
 
PBMC  
 
X 
 
X 
X 
Urine  
 
X 
X 
X 
X 
Saliva  
 
X 
 
X 
X 
Urine 
pregnancy 
test (3) 
 
X 
 
X 
X 
CDSD  
X (4) 
X  
X 
X 
X 
ICDSQ  
X (5) 
X  
X 
X 
X 
CDAT 
X (6) 
X 
X 
X 
X 
 
1. 
Serum collected 2 hour post challenge time 
2. 
Patient moves to endoscopy portion of screening after celiac serology results are negative  
3. 
Only applicable if child bearing, to be done prior to endoscopy  
4. 
CDSD is a daily symptom survey. This will be performed once at time of screening, once at time of endoscopy and then daily 
beginning Day 0 through the end of the study period. This symptom diary will be administered electronically using RedCap. 
5. 
ICDSQ is a weekly quality of life survey. This will be performed once at time of screening, once at time of first endoscopy and then 
weekly through the end of the study period. 
6. 
CDAT is a measure of gluten free diet adherence. This will be performed at time of screening and time of first endoscopy and then at 
each endoscopy visit (day 6 and week 6). 
7. 
Histology must be normal (VH:CD> 2) to move forward to gluten challenge  
8. 
Day 0 dispense gluten for day 0 to day 6.   At Day 6 dispense 5 week gluten supply.  
 
 

Protocol Version 2022.08.25 
 
 
 
 
        Page 31 | 32 
 
De-Challenge study  
 
 
Screening/Day 0  
3 months  
12-13 months  
Informed Consent  
X 
 
 
Blood collection  
X (7) 
X 
X (7) 
Start gluten free diet  
(1)  
X (2) 
 
 
Endoscopy  
X (SOC)  
X 
X 
Biopsies  
X 
X 
X 
Luminal content  
X 
X 
X 
PBMC  
X 
X 
X 
Urine  
X 
X 
X 
Saliva  
X 
X 
X 
Urine pregnancy 
test  (3) 
X (SOC) 
X 
X 
IGFDQ (4) 
X 
X 
X 
ICDSQ (5)  
X 
X 
X 
CDSD (6) 
X 
X 
X 
CDAT (4) 
X 
X 
X 
 
1. 
Gluten free diet continues throughout study 
2. 
Gluten free diet begins after histology is reviewed confirming diagnosis 
3. 
Only applicable if child bearing, to be done prior to endoscopy  
4. 
Study questionnaires to be completed prior to specimen collection and endoscopy  
5. 
ICDSQ performed once at time of screening, once at time of first endoscopy and then weekly through the end of the study period. 
6. 
CDSD is performed once at time of screening, once at time of first endoscopy and then daily through the end of the study period. 
7. 
Baseline standard of care (or research if insurance doesn’t cover or not done previously) labs: ferritin, 25 VD, zinc, copper, B12, 
folate, CBC, CMP, cholesterol; End of study standard of care labs (or research if insurance doesn’t cover): ferritin, 25 VD, zinc, 
copper, B12, folate, CBC, CMP, cholesterol. 
 
 
 
 
 
 
 

Protocol Version 2022.08.25 
 
 
 
 
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Control  
 
Screening  
Endoscopy visit 
Informed Consent  
X 
 
Blood collection  
X (2) 
X 
Endoscopy  
 
X 
Biopsies  
 
X 
Luminal content  
 
X 
Physical Exam  
X 
 
PBMC  
 
X 
Urine  
 
X 
Saliva  
 
X 
Pregnancy test (1) 
 
X 
CDSD (3) 
 
X 
 
1. 
Only applicable if child bearing, to be done prior to endoscopy  
2. 
Patient moves to endoscopy portion of screening after HLA and celiac serology results confirmed  
3. 
Symptom diary to be completed prior to specimen collection and endoscopy