Development of Fluorescent Lectin Tracers With Dedicated Technology for in Vivo Detection of Esophageal Dysplasia in Barrett Patients (GRAIN)

SYNOPSIS Glycan Near-Infrared Imaging Using Fluorescently Labeled Wheat Germ Agglutinin (WGA): Evaluation of Safety and Feasibility in a Prospective Pilot Study

Rationale Early detection of dysplastic and early carcinomas in the esophagus is critical for improving long term survival rates. Patients with Barrett's Esophagus (BE), a precancerous condition, undergo surveillance endoscopies with random four-quadrant biopsies to detect disease progression. However, this approach carries a high risk of sampling error, resulting in a detection miss-rate of up to 37%.

Improved imaging modalities may help facilitate accurate disease detection. For example, several studies have shown that near-infrared quantified fluorescence molecular endoscopy (qFME) could serve as a red flag detection technique to detect invisible tumor lesions and dysplastic tissue during endoscopy.

Cell membrane glycosylation patterns are affected in dysplastic and cancerous tissue formation. Lectins are glycan-binding proteins present in most living organisms, animals, plants and microorganisms. The University Medical Center Groningen (UMCG) developed a fluorescent tracer by labeling the common wheat lectin WGA with the fluorescent dye 800CW to detect early carcinomas in the esophagus. WGA targets the glycans N-acetylneuraminic acid - Neu5Ac (sialic acid - NANA) and N-acetylglucosamine - GlcNAc (NAG), which are dysregulated in the progression of esophageal adenocarcinoma.

WGA-800CW is a low-cost, reliable, plant-derived fluorescent lectin tracer that binds specifically to glycan residues overexpressed on dysplastic epithelial cells. Topical administration has a favorable safety profile and minimizes systemic side effects, as the tracer is topically sprayed on the esophageal mucosa, enabling localized visualization of aberrant glycosylation patterns by near-infrared fluorescence imaging to detect dysplasia and endoscopically invisible tumors.

Objective The primary objective is to evaluate the feasibility of WGA-800CW with dedicated imaging systems for detection of invisible esophageal dysplasia

Main trial endpoints A) What is the optimal dose of WGA-800CW that maximizes the tumor-to-background ratio and enables clear visualization of the tumor? B) Can fluorescence endoscopy with WGA-800CW in combination with qFME detect dysplastic esophageal lesions? Secondary trial endpoints A) Collect safety data on topical administration of WGA-800CW through evaluation of possible SAEs/AEs and vital parameters (blood pressure, heart rate and saturation).

B) Quantify and evaluate the in vivo NIR fluorescent signal of WGA-800CW by using the spectroscopy probe.

C) Correlate and validate fluorescent signals detected in vivo with ex vivo histopathology grade of dysplasia and in the resected mucosal lesions and/or biopsies taken.

D) Compare fluorescence imaging with qFME to NIRF-OCT capsule and explore whether tracer detection is feasible without conventional fluorescence endoscopy

Exploratory trial endpoints Detect target cells and tissue distribution of WGA-800CW with ex vivo analysis on extracted lesions and biopsies using fluorescence microscopy and GlcNAc and Neu5Ac levels.

Trial design The current study is a non-randomized, non-blinded, prospective, feasibility intervention study. In total a maximum of 49 patients with BE and therefore appropriate candidate for diagnostic or therapeutic gastroscopy will be included. WGA-800CW syringe of 15 mL will be administered topically via a spray catheter during diagnostic or therapeutic gastroscopy procedures prior to SEATTLE protocol biopsies or EMR/ESD resection.

Study part A - Dose finding experiments:

In study part A, a maximum of 9 patients scheduled for endoscopic treatment (EMR/ESD) will receive intra-procedurally a single dose, figure 2, of WGA-800CW topically sprayed on the esophageal mucosa. Dose escalation or de-escalation will be performed to determine the optimal dose. The vital parameters will be monitored. qFME will be performed following administration and rinsing off the excess tracer with water. All fluorescence areas will be inspected by HD-WLE and biopsied. A maximum of six biopsies of healthy, non-BE tissue will be taken to evaluate tracer distribution and specificity in esophageal tissue. The total endoscopic procedure-time will increase by ~10-15 minutes compared to standard clinical care for all steps with the study design. Based on previous NIR-FME studies, this experimental time is sufficient for all steps of the study design.

Study part B - Collecting study data using the optimal dose:

In study part B, a maximum of 40 patients scheduled for endoscopic evaluation/surveillance or endoscopic treatment will be included. Intra-procedurally, the optimal dose of WGA-800CW will be topically sprayed on the esophageal mucosa. The vital parameters will be monitored. qFME and/or NIRF-OCT will be performed following administration and rinsing off excess tracer with water. The HD-WLE suspected (pre)malignant lesion, normal esophageal epithelium, gastric mucosa and, when present, non-dysplastic BE segment will be evaluated according to their fluorescence intensity. For patients scheduled for evaluation of BE (i.e. diagnostic endoscopy), biopsies will be taken according to the Seattle protocol (standard of care). For patients scheduled for endoscopic treatment of BE (i.e. therapeutic endoscopy), dysplastic lesions will be resected with EMR or ESD (standard of care). When present, maxixmun of eight biopsies will be taken, up to six of non-fluorescent tissue and two of fluorescent tissue. The total endoscopic procedure-time will increase by approximately 10-15 minutes compared to standard clinical care for all steps with the study design.

Trial population Patients eligible for inclusion have confirmed Barrett's esophagus, esophageal dysplasia, or superficial esophageal adenocarcinoma and are scheduled for endoscopic evaluation or endoscopic treatment within the UMCG.

Interventions During the endoscopy procedure, WGA-800CW will be topically administered via a spray catheter. The near-infrared fluorescence signal will be quantified with spectroscopy and measured and analysed with fluorescence and/or tomography measuring techniques in vivo and ex vivo. A maximum of four healthy biopsies will be collected.

Ethical considerations relating to the clinical trial including the expected benefit to the individual subject or group of patients represented by the trial subjects as well as the nature and extent of burden and risks For the participating patients, there is no diagnostic or treatment benefit related to the study. Participation may possibly lead to useful data for future research. The risk of participating in this study is the administration of WGA-800CW, however, this risk is deemed negligible and acceptable given the protein fluorophore conjugate consists of a naturally occurring protein from common wheat (WGA lectin) and a fluorophore (IRDye 800CW) that has been used extensively in many clinical studies without adverse events. Clinical decisions will not be affected or influenced by the study results.