Diagnosis and Monitoring of Eosinophilic Esophagitis Using the Cytosponge

Study design overview (all Aims) This will be a prospective cohort study, with patient
enrollment conducted at UNC and Mayo Clinic with sample analysis performed by the University
of Cambridge. In Aim 1, patients with EoE will be enrolled, tissue will be obtained from both
the Cytosponge and endoscopy, and the methods will be compared for a single time point to
determine accuracy of Cytosponge for quantifying esophageal eosinophil counts. In Aim 2, the
patients who are being treated will be followed and tissue will be assessed over time with
both Cytosponge and endoscopy to determine the utility of Cytosponge for monitoring treatment
response in patients with EoE. For all patients, safety will be monitored and subjects will
complete a survey about the acceptability of Cytosponge (Aim 3).

Cytosponge protocol:

After the study has been explained and a patient provides informed consent, the Cytosponge
will be administered prior to endoscopy by trained research staff under physician
supervision. If subjects opt to receive a local anesthetic, then they will be provided with a
2% lidocaine gargle prior to administration of the Cytosponge. The Cytosponge will be
administered according to it's instructions for use. After retrieval, the string is cut and
the sponge (which contains the tissue specimen) is placed in a container, immersed in
fixative, and stored in a refrigerator at 4°C. The fixative is then spun in a centrifuge, and
the pelleted cells are embedded in a paraffin block using standard techniques.

Upper endoscopy and biopsy:

After the Cytosponge has been removed, the patient will undergo standard of care (routine
care) upper endoscopy and biopsy, as clinically indicated. During this exam, research staff
will record all endoscopic features of EoE, including rings, furrows, white plaques,
decreased vascularity, and strictures. The severity of the endoscopy findings will be
measured using the recently validated EREFS scoring system.23 Four esophageal biopsies will
be taken both from the distal (5 cm above the gastro-esophageal junction) and proximal (15 cm
above the gastro-esophageal junction) esophagus. This number of biopsies has been shown to
maximize the diagnostic sensitivity for EoE.24

Histology and eosinophil counts:

All tissue samples from the Cytosponge and endoscopy will be coded with a subject's ID
number, but will otherwise be masked for all clinical data, including EoE activity, symptoms,
patient characteristics, and treatments prescribed. Using the paraffin blocks, pathology
slides will be cut and the tissue processed with routine H&E staining. The slides will then
be digitized, and using the Aperio ImageScope (Aperio Technologies, Vista, CA), the maximum
eosinophil density (eosinophils/mm2 [eos/mm2]) will be determined using our previously
validated protocol.25 For purposes of comparison to previous studies, eosinophil density will
then be converted to eosinophil counts (eos/hpf) for an assumed hpf size of 0.24 mm2, the
size of an average field as reported in the literature.26 The study pathologists from UNC and
Mayo Clinic will review the specimens from their sites, and the study pathology from
Cambridge will provide a second review of all specimens to ensure the most accurate
quantification of eosinophil counts possible.

In addition, investigators plan to perform special staining and analysis of the existing
biopsy and sponge samples with the goal of determining if the diagnostic accuracy of this
test can be improved. In particular investigators will examine markers of eosinophil
function, activation, and inflammation, such as eosinophil peroxidase (EPX), a granule
protein that clearly identifies intact eosinophils, as well as extracellular EPX deposition
suggestive of degranulation. This can be detected with immunohistochemistry. This would be
done at Mayo clinic with our current collaborators who currently have the coded specimens.

Safety and accessibility assessments:

Patients will be assessed at multiple points to determine the safety of the Cytosponge in
EoE. Investigators will assess for any symptoms or events as soon as the sponge capsule is
swallowed, as well as immediately after the expanded sponge is removed. Participants will be
contacted 1 and 7 days after the endoscopy to assess for adverse events. For Aim 3,
participants will be administered the acceptability survey at the 7 day follow-up point, so
patients have adequate time to reflect on their experiences with both tissue collection
approaches. In particular this survey will record the patient's experience with swallowing
the Cytosponge, whether they would do it again, and whether they prefer the Cytosponge or
endoscopy for diagnosis and monitoring of EoE.

Inclusion Criteria:

- Able to read, comprehend, and complete the informed consent form

- Male or female subjects, age 18-80 years,

- Suspected EoE or has a diagnoses of EoE with current active disease,

Exclusion Criteria:

- History of esophageal stricture precluding passage of the endoscope or sponge,

- Pregnancy, or planned pregnancy during the course of the study,

- Any history of esophageal varices, liver impairment of moderate or worse severity
(Child's- Pugh class B & C) or evidence of varices noted on any past endoscopy,

- Any history of esophageal surgery, except for uncomplicated fundoplication

- History of coagulopathy, with INR>1.3 and/or platelet count of <75,000.

- Current use of blood thinners such as coumadin, warfarin, clopidogrel, heparin and/or
low molecular weight heparin (requires discontinuation of medication 7 days prior to
and 7 days after esophagogastroduodenoscopy [EGD] and Cytosponge administration,
aspirin use is OK).

- Are allergic to local anesthetics such as lidocaine (these subjects may opt not to
receive the optional lidocaine gargle prior to the Cytosponge administration and still
be eligible).

- Have not fasted the night before administration of the Cytosponge.

- History of perforation