Serum Markers in Gluten Challenge

• Sponsor: Beth Israel Deaconess Medical Center (Other)
• Overall Status: Completed
• CT.gov ID: NCT00931892
• Collaborator: National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) (NIH)
• Enrollment: 21
• Location: 1
• Arms: 2
• Actual Duration (Months): 28
• Patients Per Site Per Month: 0.8

Study Details

Study Description

Brief Summary

1. The purpose of this research study is to evaluate non-invasive markers of celiac disease activity in subjects that are on a gluten-free diet, in remission from celiac disease who undergo gluten challenge.

2. The secondary aims of this protocol are to identify novel mediators important in the pathophysiology of celiac disease and to evaluate changes in metabolism with gluten exposure.

Condition or Disease	Intervention/Treatment	Phase
Celiac Disease	Dietary Supplement: Gluten; Dietary Supplement: Gluten	N/A

Detailed Description

The diagnosis of celiac disease carries with it important ramifications. Celiac disease is a systemic immunologic disorder in which the sentinel lesion is an enteropathy triggered by polypeptides derived primarily from the gliadin proteins found in wheat, rye and barley. Ingestion of the offending proteins leads to inflammation and intestinal mucosal damage, which results in a spectrum of abdominal symptoms, increased intestinal permeability, malabsorption, occult gastrointestinal bleeding and diarrhea. Systemic manifestations of celiac disease include a myriad of conditions including malignancy and autoimmune disease.

The only accepted treatment for celiac disease is lifelong adherence to a gluten free diet. Adherence to this diet, simply put avoiding all foods containing even small amounts of wheat, rye and barley, has been shown to lead to improvement in the majority of related problems and normalization of all standard diagnostic tests. Because of this many individuals who present for evaluation of possible celiac disease but who are already on a gluten free diet cannot be tested accurately as there is currently no way of differentiating between healthy individuals and individuals with well treated celiac disease. The standard practice in such cases is to perform a 'Gluten Challenge' whereby the patient eats the equivalent of 2 slices of bread per day for six to eight weeks before returning for evaluation with serologic testing and endoscopy with duodenal biopsy. The use of the gluten challenge in clinical practice is limited by patient symptoms and resistance to such a long test period, after which it may take a number of weeks for the intestine to heal and the symptoms to resolve. Autoantibodies to tissue transglutaminase or antibodies to deamidated gliadin, while being excellent tools to predict celiac disease in patients who have been on a long-term gluten containing diet, display low sensitivities to detect short-term and/or recent gluten exposure. For this reason, it would be very useful if novel circulating markers could be identified that indicate the presence of celiac disease and in particular would provide an early and less invasive marker of a positive response to gluten challenge.

Study Design

• Study Type: Interventional
• Actual Enrollment: 21 participants
• Allocation: Randomized
• Intervention Model: Parallel Assignment
• Masking: None (Open Label)
• Primary Purpose: Diagnostic
• Official Title: Circulating Markers of Celiac Disease Activity During Gluten Challenge - a Pilot Study.
• Actual Study Start Date: Apr 1, 2009
• Actual Primary Completion Date: Aug 1, 2011
• Actual Study Completion Date: Aug 1, 2011

Arms and Interventions

Arm	Intervention/Treatment
Experimental: Low gluten group Subjects will eat 3g of gluten per day	Dietary Supplement: Gluten; 3g
Experimental: High gluten group Subjects will eat 10g of gluten per day	Dietary Supplement: Gluten; 10g

Outcome Measures

Primary Outcome Measures

1. Crypt Depth to Villous Height Ratio [Screening (Day -7 to -14), Day 3, Day 14]

   Histological evaluation of duodenal biopsy samples to evaluate crypt depth to villous height ratio. On the best oriented section of each biopsy fragment, villous height to crypt depth (Vh:Cd) ratio was determined by measuring the mean height /mean depth of adjacent villi/proliferative crypt zones at magnification 100x. Evaluations were considered discrepant Vh:Cd differed by more than 0.5. Overall, there was excellent concordance with the evaluations of the 165 biopsy fragments determined to be optimal for evaluation. The Vh:Cd ratios on individual biopsies from a single averaged to produce a representative Vh:Cd ratio for each endoscopy. Normal Vh:Cd ratio was regarded as 3:1 or greater. Adelman DC, Murray J, Wu TT, Mäki M, Green PH, Kelly CP. Measuring Change In Small Intestinal Histology In Patients With Celiac Disease. Am J Gastroenterol. 2018 Mar;113(3):339-347. doi: 10.1038/ajg.2017.480. Epub 2018 Feb 20. Review. PubMed PMID: 29460921.

Secondary Outcome Measures

1. Count of Intraepithelial Lymphocytes Per 100 Enterocytes in Duodenal Biopsy Samples [Screening (Day -7 to -14), Day 3, Day 14]

   On the best oriented section of each biopsy fragment , the number of intraepithelial lymphocytes (IEL count) per 100 enterocytes was recorded at magnification 400x. IEL counts were considered discrepant if a difference of >10 IELs existed between counts. Overall, there was excellent concordance with the evaluations of the 165 biopsy fragments determined to be optimal for evaluation. The IEL counts on individual biopsies from a single endoscopy were averaged to produce a representative IEL count for each endoscopy. Adelman DC, Murray J, Wu TT, Mäki M, Green PH, Kelly CP. Measuring Change In Small Intestinal Histology In Patients With Celiac Disease. Am J Gastroenterol. 2018 Mar;113(3):339-347. doi: 10.1038/ajg.2017.480. Epub 2018 Feb 20. Review. PubMed PMID: 29460921.

2. Measures of Intestinal Permeability (Urinary Lactulose to Mannitol Ratio) [Screening (Day -7 to -14), Day 0, Day 3, Day 7, Day 14, Day 28]

   LAMA evaluation has been reported to be an accurate measure of small intestinal mucosal permeability though assessment of differential absorption of lactulose and mannitol. For LAMA testing, a solution containing 7.5 g lactulose and 2 g mannitol in 100 mL of water was ingested by the participants in the evening after visit 1 and in the evening before each other study visit. The participants were asked to fast for at least 4 h and to void completely before drinking the sugar solution, then fast overnight and collect all overnight and morning urine. Urine sample analyzed for lactulose and mannitol using standardized methodology by liquid chromatography-tandem mass spectrometry.

3. Measures of Immune Activation [Screening (Day -7 to -14), Day 0, Day 3, Day 7, Day 14, Day 28]

   IgA anti-human tissue transglutaminase assay (Inova Diagnostics, Inc., San Diego, USA): negative <20, borderline 20-30, positive >30. IgA/IgG anti- DGP assay : NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30 METHOD IS INOVA ANTI-DEAMIDATED GLIADIN PEPTIDE IGA/IGG.

4. Assessment of Protein Expression in Intestinal Biopsies [Screening (Day -7 to -14), Day 3, Day 14]

   Proportion of participants responding to a gluten challenge. Tissue transglutaminase (tTG) scoring: NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30. Deamidated gliadin peptide (DGP) scoring: NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30 METHOD IS INOVA ANTI-DEAMIDATED GLIADIN PEPTIDE IGA/IGG.

5. Symptomatic Response to Gluten Exposure Determined by Celiac Symptom Index Questionnaire [Screening (Day -7 to -14), Day 0, Day 3, Day 7, Day 14, Day 28]

   CSI (Celiac Symptom Index) scores range from 7-80. Higher scores indicate greater degree of symptoms CSI: Leffler DA, Dennis M, Edwards George J, Jamma S, Cook EF, Schuppan D, Kelly CP. A validated disease-specific symptom index for adults with celiac disease. Clin Gastroenterol Hepatol. 2009;7:1328-34. PubMed PMID: 19665584.

6. Assessment of Protein Expression in Intestinal Biopsies Using Marsh Scores [Screening (Day -7 to -14), Day 3, Day 14]

   Proportion of participants responding to a gluten challenge. Tissue transglutaminase (tTG) scoring: NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30. Deamidated gliadin peptide (DGP) scoring: NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30 METHOD IS INOVA ANTI-DEAMIDATED GLIADIN PEPTIDE IGA/IGG.

7. Symptomatic Response to Gluten Exposure Determined by Gastrointestinal Symptom Rating Scale [Screening (Day -7 to -14), Day 0, Day 3, Day 7, Day 14, Day 28]

   Measure Description: GSRS (Gastrointestinal Symptom Rating Scale) scores range 15-105 with higher scores indicating greater degree of symptoms GSRS: Svedlund J, Sjödin I, Dotevall G. GSRS--a clinical rating scale for gastrointestinal symptoms in patients with irritable bowel syndrome and peptic ulcer disease. Dig Dis Sci. 1988;33(2):129-134. doi:10.1007/BF01535722

Eligibility Criteria

Criteria

• Ages Eligible for Study: 17 Years to 72 Years
• Sexes Eligible for Study: All
• Accepts Healthy Volunteers: No

Inclusion Criteria:

1. Age between 17 and 72 years, inclusive.

2. Subject must have been diagnosed with celiac disease by duodenal / jejunal biopsy at least 6 months prior to entrance into the study.

3. Subject has Anti-Tissue Transglutaminase (anti-tTG) ≤ 20 EU as measured by serology.

4. Subject must be on a gluten-free diet for at least the past 6 months.

5. Female subjects should be either post-menopausal (amenorrhea for at least 24 consecutive months), surgically sterile, or women of child-bearing potential (WOCP) with a negative urine beta human chorionic gonadotropin (HCG) pregnancy test prior to entering the study and who are using or agree to use acceptable methods of contraception. Abstinence is an acceptable means of avoiding pregnancy as long as the subject agrees to use contraception if they become sexually active. Acceptable contraceptives include intrauterine devices (IUDs), hormonal contraceptives (oral, depo, patch or injectable) in use for one month prior to screening and double barrier methods such as condoms or diaphragms with spermicidal gel or foam.

6. Subject must sign an Institutional Review Board approved informed consent and agree to complete required clinic visits.

7. BMI between 18.5 and 38, inclusive.

Exclusion Criteria:

1. Subject has Anti-Tissue Transglutaminase (anti-tTG) > 20 EU as measured by serology.

2. Subject has other food intolerances or food allergies (other than celiac disease) that would interfere with the conduct of the study).

3. Subject has a history of severe acute symptomatic reactions to sporadic gluten ingestion

4. Subject has any chronic active GI disease other than celiac disease (e.g. Crohn's disease, IBS).

5. Subjects with symptomatic neurological or psychiatric disease(s) that would interfere with the conduct of the study.

6. Subject has clinically significant abnormal laboratory test results at the screening visit or as determined by the Principal Investigator

7. Subject is pregnant or breast feeding.

8. Subject (premenopausal females) is sexually active without contraception.

9. Subject should not have been on steroids in the past 3 months.

10. Subject is deemed inappropriate by the Principal Investigator.

Contacts and Locations

Locations

	Site	City	State	Country	Postal Code
1	Beth Israel Deaconess Medical Center	Boston	Massachusetts	United States	02215

Sponsors and Collaborators

• Beth Israel Deaconess Medical Center
• National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)

Investigators

• Principal Investigator: Ciaran P Kelly, MD, Beth Israel Deaconess Medical Center
• Study Director: Daniel A Leffler, MD, MS, Beth Israel Deaconess Medical Center

Study Documents (Full-Text)

None provided.

More Information

Additional Information:

• BIDMC Celiac Center Research website

Publications

• Svedlund J, Sjödin I, Dotevall G. GSRS--a clinical rating scale for gastrointestinal symptoms in patients with irritable bowel syndrome and peptic ulcer disease. Dig Dis Sci. 1988 Feb;33(2):129-34.

• Responsible Party: Ciaran Kelly, Professor of Medicine, Beth Israel Deaconess Medical Center
• ClinicalTrials.gov Identifier: NCT00931892
• Other Study ID Numbers: 2007P000280; K23DK082619
• First Posted: Jul 2, 2009
• Last Update Posted: Jun 4, 2021
• Last Verified: May 1, 2021

• Keywords provided by Ciaran Kelly, Professor of Medicine, Beth Israel Deaconess Medical Center: celiac disease; gluten challenge
• Additional relevant MeSH terms: Celiac Disease

Study Results

Participant Flow

Pre-assignment Detail	21 patients were enrolled in the study. 1 patient was found to be negative for DQ2 and DQ8, therefore did not have Celiac disease. This patient was excluded from the study. Therefore, for the study we had a total of 20 participants.

Arm/Group Title	Low Gluten Group	High Gluten Group
Arm/Group Description	Subjects will eat 3g of gluten per day Gluten: 3g	Subjects will eat 10g of gluten per day Gluten: 10g
Period Title: Completed Gluten Challenge
STARTED	10	10
COMPLETED	10	10
NOT COMPLETED	0	0
Period Title: Completed Gluten Challenge
STARTED	10	10
First Endoscopy	10	10
Second Endoscopy	10	10
Third Endoscopy	10	9
COMPLETED	10	9
NOT COMPLETED	0	1

Baseline Characteristics

Arm/Group Title	Low Gluten Group	High Gluten Group	Total
Arm/Group Description	Participants meeting inclusion criteria were randomized into the low gluten group. These participants ate two slices of wheat bread per day (3 g of gluten). Bread was procured from a standard source and gluten dose tested using the RIDASCREEN Gliadin R5 antibody ELISA. The study included a 14-day run-in period followed by a 14 day gluten challenge and a final visit 14 days post-gluten challenge. Other than the dispensed bread, subjects were instructed to remain on their gluten-free diet throughout the duration of the study. Gluten: 3g	Participants meeting inclusion criteria were randomized into the low gluten group. These participants ate five slices of wheat bread per day (10 g of gluten). Bread was procured from a standard source and gluten dose tested using the RIDASCREEN Gliadin R5 antibody ELISA. The study included a 14-day run-in period followed by a 14 day gluten challenge and a final visit 14 days post-gluten challenge. Other than the dispensed bread, subjects were instructed to remain on their gluten-free diet throughout the duration of the study. Gluten: 10g	Total of all reporting groups
Overall Participants	10	10	20
Age (Count of Participants)
<=18 years	0 0%	0 0%	0 0%
Between 18 and 65 years	10 100%	10 100%	20 100%
>=65 years	0 0%	0 0%	0 0%
Age (years) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [years]	48.8 (13.0)	38.8 (13.1)	43.3 (13.7)
Sex: Female, Male (Count of Participants)
Female	10 100%	9 90%	19 95%
Male	0 0%	1 10%	1 5%
Ethnicity (NIH/OMB) (Count of Participants)
Hispanic or Latino	0 0%	0 0%	0 0%
Not Hispanic or Latino	10 100%	10 100%	20 100%
Unknown or Not Reported	0 0%	0 0%	0 0%
Race (NIH/OMB) (Count of Participants)
American Indian or Alaska Native	0 0%	0 0%	0 0%
Asian	1 10%	0 0%	1 5%
Native Hawaiian or Other Pacific Islander	0 0%	0 0%	0 0%
Black or African American	0 0%	0 0%	0 0%
White	9 90%	10 100%	19 95%
More than one race	0 0%	0 0%	0 0%
Unknown or Not Reported	0 0%	0 0%	0 0%
Region of Enrollment (participants) [Number]
United States	10 100%	10 100%	20 100%
Age at Celiac Diagnosis (years) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [years]	45.4 (13.9)	31.0 (11.7)	35.7 (14.4)
Months since Celiac Disease diagnosis (months) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [months]	42.3 (21.3)	92.5 (49.1)	62.3 (87.1)
Laboratory Characteristics (Count of Participants)
HLA DQ2	4 40%	6 60%	10 50%
HLA DQ8	3 30%	3 30%	6 30%
HLA DQ2 + DQ 8	1 10%	1 10%	2 10%
HLA A1 * 05 alone	2 20%	0 0%	2 10%
Villous height to crypt depth ratio (unitless) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [unitless]	2.05 (0.80)	4.33 (6.69)	2.21 (0.8)
Mean number of Intraepithelial lymphocytes (unitless) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [unitless]	37.10 (13.66)	28.4 (13.3)	32.59 (15.1)
Baseline tissue transglutaminase (tTG) (units) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [units]	13.76 (8.30)	8.27 (5.70)	11.00 (7.50)
Baseline Deamidated gliadin peptide (DGP) (units) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [units]	6.70 (2.59)	14.04 (26.49)	10.90 (19.60)
Baseline Celiac Dietary Adherence test (CDAT) (scores on a scale) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [scores on a scale]	11.33 (3.2)	11.40 (2.9)	11.37 (2.9)
Baseline Celiac Symptom Index (CSI) (scores on a scale) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [scores on a scale]	28.50 (5.78)	22.50 (4.99)	25.26 (5.90)
Baseline Gastrointestinal Symptom Rating Scale (GSRS) (scores on a scale) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [scores on a scale]	22.88 (9.58)	19.20 (3.26)	20.32 (5.10)
Baseline Quality of Life Visual Analog Scale (VAS) (scores on a scale) [Mean (Standard Deviation) ]
Mean (Standard Deviation) [scores on a scale]	80.00 (13.10)	84.40 (12.10)	82.20 (12.50)

Outcome Measures

1. Primary Outcome

Title	Crypt Depth to Villous Height Ratio
Description	Histological evaluation of duodenal biopsy samples to evaluate crypt depth to villous height ratio. On the best oriented section of each biopsy fragment, villous height to crypt depth (Vh:Cd) ratio was determined by measuring the mean height /mean depth of adjacent villi/proliferative crypt zones at magnification 100x. Evaluations were considered discrepant Vh:Cd differed by more than 0.5. Overall, there was excellent concordance with the evaluations of the 165 biopsy fragments determined to be optimal for evaluation. The Vh:Cd ratios on individual biopsies from a single averaged to produce a representative Vh:Cd ratio for each endoscopy. Normal Vh:Cd ratio was regarded as 3:1 or greater. Adelman DC, Murray J, Wu TT, Mäki M, Green PH, Kelly CP. Measuring Change In Small Intestinal Histology In Patients With Celiac Disease. Am J Gastroenterol. 2018 Mar;113(3):339-347. doi: 10.1038/ajg.2017.480. Epub 2018 Feb 20. Review. PubMed PMID: 29460921.
Time Frame	Screening (Day -7 to -14), Day 3, Day 14

Outcome Measure Data

Analysis Population Description

Subjects with biopsy proven celiac disease in remission were enrolled. For Day 3, 17 subjects had evaluable pathology biopsies. On Day 14, 19 participants had evaluable pathology biopsies.

Arm/Group Title	Baseline (Day -14) Low Gluten Group	Baseline (Day -14) High Gluten Group	Day 3 Low Gluten Group	Day 3 High Gluten Group	Day 14 Low Gluten Group	Day 14 High Gluten Group
Arm/Group Description	Villous height to crypt depth ratio in low gluten group In the low gluten group, subjects consumed 3 grams of gluten per day	Villous height to crypt depth ratio in high gluten group In the high gluten group, subjects consumed 10 grams of gluten per day.	Villous height to crypt depth ratio in low gluten group In the low gluten group, subjects consumed 3 grams of gluten per day.	Villous height to crypt depth ratio in high gluten group In the high gluten group, subjects consumed 10 grams of gluten per day.	Villous height to crypt depth ratio in low gluten group In the low gluten group, subjects consumed 3 grams of gluten per day	Villous height to crypt depth ratio in high gluten group. In the high gluten group subjects consume 10 grams of gluten per day.
Measure Participants	10	10	8	9	10	9
Mean (Standard Deviation) [ratio]	2.05 (0.80)	4.33 (6.69)	1.19 (0.67)	2.02 (0.99)	0.98 (0.85)	1.26 (1.08)

Statistical Analysis 1

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 3 Low Gluten Group, Day 3 High Gluten Group, Day 14 Low Gluten Group, Day 14 High Gluten Group
	Comments
	Type of Statistical Test	Other
	Comments
Statistical Test of Hypothesis	p-Value	0.670
	Comments
	Method	t-test, 2 sided
	Comments

2. Secondary Outcome

Title	Count of Intraepithelial Lymphocytes Per 100 Enterocytes in Duodenal Biopsy Samples
Description	On the best oriented section of each biopsy fragment , the number of intraepithelial lymphocytes (IEL count) per 100 enterocytes was recorded at magnification 400x. IEL counts were considered discrepant if a difference of >10 IELs existed between counts. Overall, there was excellent concordance with the evaluations of the 165 biopsy fragments determined to be optimal for evaluation. The IEL counts on individual biopsies from a single endoscopy were averaged to produce a representative IEL count for each endoscopy. Adelman DC, Murray J, Wu TT, Mäki M, Green PH, Kelly CP. Measuring Change In Small Intestinal Histology In Patients With Celiac Disease. Am J Gastroenterol. 2018 Mar;113(3):339-347. doi: 10.1038/ajg.2017.480. Epub 2018 Feb 20. Review. PubMed PMID: 29460921.
Time Frame	Screening (Day -7 to -14), Day 3, Day 14

Outcome Measure Data

Analysis Population Description

Subjects with biopsy proven celiac disease in remission were enrolled. For Day 3, 17 subjects had evaluable pathology biopsies. On Day 14, 19 participants had evaluable pathology biopsies.

Arm/Group Title	Baseline (Day -14) Low Gluten Dose Group	Baseline (Day -14) High Gluten Dose Group	Day 3 Low Gluten Dose Group	Day 3 High Gluten Dose Group	Day 14 Low Gluten Dose Group	Day 14 High Gluten Dose Group
Arm/Group Description	Intraepithelial lymphocyte count(per 100 enterocytes) in low gluten dose groups. Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Intraepithelial lymphocyte count(per 100 enterocytes) in high gluten dose groups. Subjects in the high gluten dose group consumed 10 grams of gluten per day	Intraepithelial lymphocyte count(per 100 enterocytes) in low gluten dose groups. Subjects in the low gluten dose group consumed 3 grams of gluten per day	Intraepithelial lymphocyte count(per 100 enterocytes) in high gluten dose groups. Subjects in the low gluten dose group consumed 10 grams of gluten per day	Intraepithelial lymphocyte count(per 100 enterocytes) in low gluten dose groups. Subjects in the low gluten dose group consumed 3 grams of gluten per day	Intraepithelial lymphocyte count(per 100 enterocytes) in high gluten dose groups. Subjects in the high gluten dose group consumed 10 grams of gluten per day
Measure Participants	10	10	8	9	10	9
Mean (Standard Deviation) [Count of Lymphocytes Per 100 Enterocytes]	37.08 (13.66)	28.91 (15.81)	44.89 (20.49)	30.68 (18.09)	47.95 (14.30)	52.36 (28.52)

Statistical Analysis 1

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 3 Low Gluten Group, Day 3 High Gluten Group, Day 14 Low Gluten Group, Day 14 High Gluten Group
	Comments
	Type of Statistical Test	Other
	Comments
Statistical Test of Hypothesis	p-Value	0.409
	Comments
	Method	t-test, 2 sided
	Comments

3. Secondary Outcome

Title	Measures of Intestinal Permeability (Urinary Lactulose to Mannitol Ratio)
Description	LAMA evaluation has been reported to be an accurate measure of small intestinal mucosal permeability though assessment of differential absorption of lactulose and mannitol. For LAMA testing, a solution containing 7.5 g lactulose and 2 g mannitol in 100 mL of water was ingested by the participants in the evening after visit 1 and in the evening before each other study visit. The participants were asked to fast for at least 4 h and to void completely before drinking the sugar solution, then fast overnight and collect all overnight and morning urine. Urine sample analyzed for lactulose and mannitol using standardized methodology by liquid chromatography-tandem mass spectrometry.
Time Frame	Screening (Day -7 to -14), Day 0, Day 3, Day 7, Day 14, Day 28

Outcome Measure Data

Analysis Population Description

Subjects with biopsy proven celiac disease in remission were enrolled. Measures of intestinal permeability (urinary lactulose to mannitol ratio) (LAMA) . Four participants were unable to tolerate LAMA testing due to gastrointestinal symptoms.

Arm/Group Title	Day -14 Low Gluten Dose Group	Day -14 High Gluten Dose Group	Day 0 Low Gluten Dose Group	Day 0 High Gluten Dose Group	Day 3 Low Gluten Dose Group	Day 3 High Gluten Dose Group	Day 7 Low Gluten Dose Group	Day 7 High Gluten Dose Group	Day 14 Low Gluten Dose Group	Day 14 High Gluten Dose Group	Day 28 Low Gluten Dose Group	Day 28 High Gluten Dose Group
Arm/Group Description	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.
Measure Participants	8	8	8	8	8	7	8	7	8	6	8	7
Mean (Standard Deviation) [ratio]	0.23 (0.17)	0.24 (0.25)	0.65 (1.15)	0.17 (0.15)	0.25 (0.15)	0.44 (0.60)	0.45 (0.71)	0.29 (0.28)	0.89 (1.53)	0.59 (0.52)	0.29 (0.24)	0.57 (0.82)

Statistical Analysis 1

Statistical Analysis Overview	Comparison Group Selection	Day 14 Low Gluten Dose Group, Day 14 High Gluten Dose Group, Day 28 Low Gluten Dose Group, Day 28 High Gluten Dose Group
	Comments
	Type of Statistical Test	Other
	Comments
Statistical Test of Hypothesis	p-Value	0.387
	Comments
	Method	t-test, 2 sided
	Comments

Statistical Analysis 2

Statistical Analysis Overview	Comparison Group Selection	Day 3 Low Gluten Group, Day 3 High Gluten Group, Day 28 Low Gluten Dose Group, Day 28 High Gluten Dose Group
	Comments
	Type of Statistical Test	Other
	Comments
Statistical Test of Hypothesis	p-Value	0.208
	Comments
	Method	t-test, 2 sided
	Comments

4. Secondary Outcome

Title	Measures of Immune Activation
Description	IgA anti-human tissue transglutaminase assay (Inova Diagnostics, Inc., San Diego, USA): negative <20, borderline 20-30, positive >30. IgA/IgG anti- DGP assay : NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30 METHOD IS INOVA ANTI-DEAMIDATED GLIADIN PEPTIDE IGA/IGG.
Time Frame	Screening (Day -7 to -14), Day 0, Day 3, Day 7, Day 14, Day 28

Outcome Measure Data

Analysis Population Description

Subjects with biopsy proven celiac disease in remission were enrolled.

Arm/Group Title	Day -14 Low Gluten Dose Group	Day -14 High Gluten Dose Group	Day 0 Low Gluten Dose Group	Day 0 High Gluten Dose Group	Day 3 Low Gluten Dose Group	Day 3 High Gluten Dose Group	Day 7 Low Gluten Dose Group	Day 7 High Gluten Dose Group	Day 14 Low Gluten Dose Group	Day 14 High Gluten Dose Group	Day 28 Low Gluten Dose Group	Day 28 High Gluten Dose Group
Arm/Group Description	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day
Measure Participants	10	10	10	10	10	10	10	10	10	10	10	10
IgA anti-tTG	6.70 (2.60)	10.34 (5.75)	13.76 (10.88)	11.50 (6.63)	11.84 (9.50)	10.34 (5.75)	13.77 (10.90)	11.50 (6.63)	12.63 (8.73)	15.16 (10.30)	37.81 (45.07)	43.20 (53.77)
IgA/IgG anti- DGP	6.70 (2.60)	14.04 (26.49)	7.83 (2.54)	14.69 (27.01)	12.95 (8.26)	14.04 (26.49)	7.83 (2.54)	14.69 (27.01)	17.81 (22.97)	22.74 (29.31)	45.42 (62.56)	41.95 (33.91)

Statistical Analysis 1

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 14 Low Gluten Dose Group, Day 14 High Gluten Dose Group
	Comments	P Value for IgA anti-tTG
	Type of Statistical Test	Superiority
	Comments
Statistical Test of Hypothesis	p-Value	0.010
	Comments
	Method	ANOVA
	Comments

Statistical Analysis 2

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 14 Low Gluten Dose Group, Day 14 High Gluten Dose Group
	Comments	P Value for IgA/IgG anti-DGP
	Type of Statistical Test	Superiority
	Comments
Statistical Test of Hypothesis	p-Value	0.036
	Comments
	Method	ANOVA
	Comments

Statistical Analysis 3

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 28 Low Gluten Dose Group, Day 28 High Gluten Dose Group
	Comments	P Value for IgA anti-tTG
	Type of Statistical Test	Superiority
	Comments
Statistical Test of Hypothesis	p-Value	0.013
	Comments
	Method	ANOVA
	Comments

Statistical Analysis 4

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 28 Low Gluten Dose Group, Day 28 High Gluten Dose Group
	Comments	P Value for IgA/IgG anti-DGP
	Type of Statistical Test	Superiority
	Comments
Statistical Test of Hypothesis	p-Value	0.006
	Comments
	Method	ANOVA
	Comments

5. Secondary Outcome

Title	Assessment of Protein Expression in Intestinal Biopsies
Description	Proportion of participants responding to a gluten challenge. Tissue transglutaminase (tTG) scoring: NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30. Deamidated gliadin peptide (DGP) scoring: NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30 METHOD IS INOVA ANTI-DEAMIDATED GLIADIN PEPTIDE IGA/IGG.
Time Frame	Screening (Day -7 to -14), Day 3, Day 14

Outcome Measure Data

Analysis Population Description

All 20 subjects were included for the tTG and DGP analysis. For Day 28 High Gluten Challenge Dose Group, 1 participant had both a tTG and DGP >20 which is why the count is greater than the number analyzed.

Arm/Group Title	Day 14 Low Gluten Dose Group	Day 14 High Gluten Challenge Group	Day 28 Low Gluten Dose Group	Day 28 High Gluten Dose Group	Day 14 or Day 28 Low Gluten Dose Group	Day 14 or Day 28 High Gluten Dose Group
Arm/Group Description	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day
Measure Participants	10	10	10	10	20	20
tTG > 20	3 30%	2 20%	5 25%	6 NaN	8 NaN	8 NaN
DGP > 20	2 20%	4 40%	4 20%	5 NaN	6 NaN	9 NaN
tTG > 20 or DGP > 20	5 50%	6 60%	9 45%	10 NaN	14 NaN	17 NaN

6. Secondary Outcome

Title	Symptomatic Response to Gluten Exposure Determined by Celiac Symptom Index Questionnaire
Description	CSI (Celiac Symptom Index) scores range from 7-80. Higher scores indicate greater degree of symptoms CSI: Leffler DA, Dennis M, Edwards George J, Jamma S, Cook EF, Schuppan D, Kelly CP. A validated disease-specific symptom index for adults with celiac disease. Clin Gastroenterol Hepatol. 2009;7:1328-34. PubMed PMID: 19665584.
Time Frame	Screening (Day -7 to -14), Day 0, Day 3, Day 7, Day 14, Day 28

Outcome Measure Data

Analysis Population Description

Subjects with biopsy proven celiac disease in remission were enrolled. On Day -14, one subject in the high gluten dose group missed the CSI. On Day 7 one subject in the high gluten dose group missed the CSI. On Day 14 one subject in the high gluten dose group missed the CSI. On Day 28, one subject in the high gluten dose group and one subject in the low gluten dose missed the CSI.

Arm/Group Title	Day -14 Low Gluten Dose Group	Day -14 High Gluten Dose Group	Day 0 Low Gluten Dose Group	Day 0 High Gluten Dose Group	Day 3 Low Gluten Dose Group	Day 3 High Gluten Dose Group	Day 7 Low Gluten Dose Group	Day 7 High Gluten Dose Group	Day 14 Low Gluten Dose Group	Day 14 High Gluten Dose Group	Day 28 Low Gluten Dose Group	Day 28 High Gluten Dose Group
Arm/Group Description	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.
Measure Participants	10	9	10	10	10	10	10	9	10	9	9	9
Mean (Standard Deviation) [score on a scale]	28.5 (5.78)	22.5 (4.99)	28.44 (6.44)	23.00 (5.29)	30.33 (7.81)	27.54 (10.27)	31.11 (7.90)	28.80 (13.36)	31.38 (11.40)	31.00 (12.70)	26.88 (7.70)	23.90 (6.740)

Statistical Analysis 1

Statistical Analysis Overview	Comparison Group Selection	Day 14 Low Gluten Group, Day 14 High Gluten Group
	Comments	P Value for Celiac Symptom Index (CSI) score for Day 3 of Gluten Challenge
	Type of Statistical Test	Other
	Comments
Statistical Test of Hypothesis	p-Value	0.05
	Comments
	Method	t-test, 2 sided
	Comments

Statistical Analysis 2

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 14 Low Gluten Dose Group, Day 14 High Gluten Dose Group
	Comments	P Value for Celiac Symptom Index (CSI) Score from baseline to Day 14
	Type of Statistical Test	Other
	Comments
Statistical Test of Hypothesis	p-Value	0.020
	Comments
	Method	t-test, 2 sided
	Comments

Statistical Analysis 3

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 3 Low Gluten Group, Day 3 High Gluten Group, Day 14 Low Gluten Group, Day 14 High Gluten Group, Day 7 Low Gluten Dose Group, Day 7 High Gluten Dose Group, Day 14 Low Gluten Dose Group, Day 14 High Gluten Dose Group, Day 28 Low Gluten Dose Group, Day 28 High Gluten Dose Group
	Comments	P Value for Celiac Symptom Index (CSI)score between high gluten group and low gluten group across study
	Type of Statistical Test	Superiority
	Comments
Statistical Test of Hypothesis	p-Value	0.060
	Comments
	Method	ANOVA
	Comments

7. Secondary Outcome

Title	Assessment of Protein Expression in Intestinal Biopsies Using Marsh Scores
Description	Proportion of participants responding to a gluten challenge. Tissue transglutaminase (tTG) scoring: NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30. Deamidated gliadin peptide (DGP) scoring: NEGATIVE <20, BORDERLINE 20-30, POSITIVE >30 METHOD IS INOVA ANTI-DEAMIDATED GLIADIN PEPTIDE IGA/IGG.
Time Frame	Screening (Day -7 to -14), Day 3, Day 14

Outcome Measure Data

Analysis Population Description

Only 19 subjects were evaluated for Marsh scores because one patient had a single endoscopy which did not produce evaluable samples.

Arm/Group Title	Day 14 Low Gluten Dose Group	Day 14 High Gluten Challenge Group	Day 28 Low Gluten Dose Group	Day 28 High Gluten Dose Group	Day 14 or Day 28 Low Gluten Dose Group	Day 14 or Day 28 High Gluten Dose Group
Arm/Group Description	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day	Subjects in the low gluten dose group consumed 3 grams of gluten per day	Subjects in the high gluten dose group consumed 10 grams of gluten per day
Measure Participants	10	9	10	9	20	19
Marsh III	5 50%	8 80%	0 0%	0 NaN	5 NaN	8 NaN
Marsh III or tTG >20	8 80%	9 90%	5 25%	6 NaN	13 NaN	16 NaN
Marsh III or DGP > 20	7 70%	9 90%	4 20%	5 NaN	11 NaN	17 NaN
Marsh III or DGP > 20 or tTG > 20	10 100%	9 90%	9 45%	9 NaN	19 NaN	19 NaN

8. Secondary Outcome

Title	Symptomatic Response to Gluten Exposure Determined by Gastrointestinal Symptom Rating Scale
Description	Measure Description: GSRS (Gastrointestinal Symptom Rating Scale) scores range 15-105 with higher scores indicating greater degree of symptoms GSRS: Svedlund J, Sjödin I, Dotevall G. GSRS--a clinical rating scale for gastrointestinal symptoms in patients with irritable bowel syndrome and peptic ulcer disease. Dig Dis Sci. 1988;33(2):129-134. doi:10.1007/BF01535722
Time Frame	Screening (Day -7 to -14), Day 0, Day 3, Day 7, Day 14, Day 28

Outcome Measure Data

Analysis Population Description

Subjects with biopsy proven celiac disease in remission were enrolled. On Day -14, one subject in the high gluten group missed completing the GSRS. On Day 0, two subjects (one in the low gluten group and one subject in the high gluten group) missed completing the GSRS. On Day 7, one subject in the high gluten group missed completing the GSRS. On Day 14, one subject in the high gluten group missed completing the GSRS. On Day 28, one subject in the high gluten group missed completing the GSRS.

Arm/Group Title	Day -14 Low Gluten Dose Group	Day -14 High Gluten Dose Group	Day 0 Low Gluten Dose Group	Day 0 High Gluten Dose Group	Day 3 Low Gluten Dose Group	Day 3 High Gluten Dose Group	Day 7 Low Gluten Dose Group	Day 7 High Gluten Dose Group	Day 14 Low Gluten Dose Group	Day 14 High Gluten Dose Group	Day 28 Low Gluten Dose Group	Day 28 High Gluten Dose Group
Arm/Group Description	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.	Subjects in the low gluten dose group consumed 3 grams of gluten per day.	Subjects in the high gluten dose group consumed 10 grams of gluten per day.
Measure Participants	10	9	9	9	10	10	10	9	10	9	10	9
Mean (Standard Deviation) [score on a scale]	21.63 (7.05)	19.20 (3.26)	22.88 (9.58)	18.40 (3.06)	30.22 (11.63)	26.45 (13.18)	28.22 (9.87)	27.30 (14.94)	31.00 (13.71)	31.00 (17.09)	23.89 (11.59)	19.40 (4.33)

Statistical Analysis 1

Statistical Analysis Overview	Comparison Group Selection	Day 14 Low Gluten Group, Day 14 High Gluten Group
	Comments	P Value for the Gastrointestinal Symptom Rating Scale (GSRS) on Day 3 of the Gluten Challenge
	Type of Statistical Test	Other
	Comments
Statistical Test of Hypothesis	p-Value	0.01
	Comments
	Method	t-test, 2 sided
	Comments

Statistical Analysis 2

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 14 Low Gluten Dose Group, Day 14 High Gluten Dose Group
	Comments	P Value for Gastrointestinal Symptom Rating Scale (GSRS) from baseline to Day 14 of Gluten Challenge
	Type of Statistical Test	Other
	Comments
Statistical Test of Hypothesis	p-Value	0.012
	Comments
	Method	t-test, 2 sided
	Comments

Statistical Analysis 3

Statistical Analysis Overview	Comparison Group Selection	Baseline (Day -14) Low Gluten Group, Baseline (Day -14) High Gluten Group, Day 3 Low Gluten Group, Day 3 High Gluten Group, Day 14 Low Gluten Group, Day 14 High Gluten Group, Day 7 Low Gluten Dose Group, Day 7 High Gluten Dose Group, Day 14 Low Gluten Dose Group, Day 14 High Gluten Dose Group, Day 28 Low Gluten Dose Group, Day 28 High Gluten Dose Group
	Comments	P Value for Gastrointestinal Symptom Rating Scale (GSRS) between high gluten group and low gluten group across study
	Type of Statistical Test	Superiority
	Comments
Statistical Test of Hypothesis	p-Value	0.090
	Comments
	Method	ANOVA
	Comments

Adverse Events

Time Frame	4 weeks after participant received first dose
Arm/Group Title	Low Gluten Group		High Gluten Group
Arm/Group Description	Subjects will eat 3g of gluten per day Gluten: 3g		Subjects will eat 10g of gluten per day Gluten: 10g
All Cause Mortality
	Low Gluten Group		High Gluten Group
	Affected / at Risk (%)	# Events	Affected / at Risk (%)	# Events
Total	0/10 (0%)		0/10 (0%)
Serious Adverse Events
	Low Gluten Group		High Gluten Group
	Affected / at Risk (%)	# Events	Affected / at Risk (%)	# Events
Total	0/10 (0%)		0/10 (0%)
Other (Not Including Serious) Adverse Events
	Low Gluten Group		High Gluten Group
	Affected / at Risk (%)	# Events	Affected / at Risk (%)	# Events
Total	1/10 (10%)		0/10 (0%)
Gastrointestinal disorders
Nausea and vomiting	1/10 (10%)	1	0/10 (0%)	0

Limitations/Caveats

This study did have few limitations. First, the study was only moderate in size and drawn from a single center. Many subjects had significant inflammation on duodenal biopsy at baseline which may have limited the effect of gluten challenge. Finally, the high and low gluten groups were significantly different in a few factors including time since diagnosis, time on a gluten-free diet and GSRS score.

More Information

Certain Agreements

Principal Investigators are NOT employed by the organization sponsoring the study.

There is NOT an agreement between Principal Investigators and the Sponsor (or its agents) that restricts the PI's rights to discuss or publish trial results after the trial is completed.