Visualization of a VEGF-targeted Near-Infrared Fluorescent Tracer in patients with Familial Adenomatous Polyposis during Fluorescence Endoscopy.
A single center pilot intervention study

There is a need for better visualization of polyps during surveillance
endoscopy in patients with hereditary colon cancer syndromes like Familial
Adenomatous Polyposis (FAP) and Lynch Syndrome (LS), to improve the adenoma
detection rate. Optical molecular imaging of adenoma associated biomarkers is a
promising technique to accommodate this need. The biomarker Vascular
Endothelial Growth Factor (VEGF) is overexpressed in adenomatous colon tissue
versus normal tissue and has proven to be a valid target for molecular imaging.
The University Medical Center Groningen (UMCG) developed a fluorescent tracer
by labeling the VEGF-targeting humanized monoclonal antibody bevacizumab,
currently used in anti-cancer therapy, with the fluorescent dye IRDye800CW. We
hypothesize that when bevacizumab-IRDye800CW is administered to patients, it
accumulates in VEGF expressing adenomas, enabling adenoma visualization using a
newly developed flexible near-infrared (NIR) fluorescence endoscope and
optoacoustic endoscope (NL43407.042.13). We want to test this hypothesis in
this feasibility study, and we will determine the best tracer dosage (10, 25 or
50mg) for adequate visualization. We chose to start in patients with FAP since
these patients have definitely adenomas.

The primary objective of this study is to determine the sensitivity of the
fluorescent tracer bevacizumab-IRDye800CW using a NIR fluorescence endoscope
ans spectroscopy probe in identifying adenomatous polyps during surveillance
endoscopy in patients with FAP. The in vivo fluorescent signal of adenomas,
measured with the NIR fluorescence endoscope, will be analyzed. The
accumulation of bevacizumab-IRDye800CW and the VEGF expression levels will be
analyzed ex vivo in resected adenomas and normal colon tissue biopsies. To
assess the primary objective, these results will be compared.

Secondary objectives:
- Testing of different tracer dosages to establish the optimal dosage for
visualization.
- To collect safety data of Bevacizumab-IRDye800CW.
- Monitor bevacizumab-800CW plasma concentration and plasma stability in blood
(based on blood drawings one hour after injection, 3 days after injection and *
optional- 7 days after injection).
- Distinguish and quantify in vivo the NIR fluorescent signal of
Bevacizumab-800CW by means of the MDSFR/SFF spectroscopy probe.
- To investigate the correlation between fluorescence intensity of adenomas
and the grade of VEGF expression and dysplasia.
- To evaluate the ability of the optoacoustic endoscope to detect
bevacizumab-IRDye800CW distribution in deeper areas of adenomas and surrounding
tissue.
- To evaluate the fluorescence intensity of normal tissue versus adenomas.

The current study is a non-randomized, non-blinded, prospective, single center
pilot intervention study. In total 120 patients with FAP will be included.
Patients with FAP will receive an intravenous injection with the VEGF-targeting
fluorescent tracer bevacizumab-IRDye800CW. Three days after tracer
administration (depending on cohort), patients will undergo the endoscopy
procedure: first optoacoustic endoscopy, followed by near-infrared fluorescence
endoscopy and spectroscopy followed by the standard care surveillance sigmoid
endoscopy with polyp removal and biopsies.

Informing patients
When patients with genetically or clinically proven FAP receive information
about their scheduled regular recto-sigmoid surveillance endoscopy, they
receive also written study information. When patients are interested in
participating in this study, they can call, e-mail or return a letter to one of
the investigators involved in this study. Interested patients will be phoned to
give them the opportunity to have an appointment at the UMCG to discuss the
study with a doctor involved in this study, but this is not mandatory. When
patients agree to participate, an appointment for tracer administration is
planned, with on the forehand signing of the informed consent.

Tracer administration
After patient and one of the study investigators have signed informed consent,
the patient will undergo a vena punction, baseline ECG and afterwards receive
the tracer Bevacizumab-IRDye800CW intravenously (10, 25 or 50mg depending on
the cohort) with 1 hour of observation to monitor safety. The total duration of
this visit is 1.5 hours.

Study cohorts
This study consists of three different cohorts to determine the optimal tracer
dosage of bevacizumab-IRDye800CW. The first cohort will receive a single dose
of 10 mg bevacizumab-800CW, 3 days before endoscopic fluorescence imaging. This
cohort will consist of 3 patients with colorectal polyps. If the data of the
first three patients give not enough consistency regarding read-out 1 and 2,
the cohort can be extended up to 6 FAP patients with colorectal polyps. To
ensure safety, a report on safety data is send to the DSMB. Based on these
results, the study team decides if the used dose is sufficient for endoscopic
imaging of colorectal polyps. If so, the dose escalation study will not be
extended to the higher doses since we aim to use a dose as low as possible. If
the study team concludes it is not sufficient, the second cohort opens, with
FAP patients receiving 25 mg bevacizumab-800CW. The same rules are applied on
this cohort and the 50 mg cohort as described before.

NIR fluorescence endoscopy procedure
Thirty minutes prior to the endoscopy a laxative enema is given. The procedure
starts with optoacoustic endoscopy, when available. The optoacoustic endoscope
will be introduced to detect the distribution of bevacizumab-IRDye800CW in
deeper areas of the adenomas and lymph nodes. Subsequently, NIR fluorescence
endoscopy with subsequent spectroscopy will be performed where the fluorescence
intensity of bevacizumab-IRDye800CW in the adenomas present in the
recto-sigmoid will be determined. Study related procedures will take place:
polypectomy of maximal six small adenomas (< 5mm, if available three polyps
with fluorescent signal and three polyps without signal). Finally, 4 biopsies
will be taken from normal rectal tissue to get insight in
bevacizumab-IRDye800CW distribution in normal colon crypts. The fluorescence
endoscopy will be followed by standard white-light surveillance endoscopy where
standard care will take place: all polyps of * 5mm are removed and send to the
department of Pathology. The optoacoustic and fluorescence endoscopy procedures
will be digitally documented. Polypectomy specimens and biopsies will be
analyzed ex vivo. The visit for the endoscopy procedure will take up to 1.5
hours, this is approximately 0.5 hour longer than patients who undergo only the
standard surveillance endoscopy.

Patients with FAP will receive an intravenous injection with the VEGF-targeting
fluorescent tracer bevacizumab-IRDye800CW. Three days after tracer
administration, patients will undergo the endoscopy procedure: first
optoacoustic endoscopy, followed by near-infrared fluorescence endoscopy and
spectroscopy, followed by the standard care surveillance sigmoid endoscopy with
polyp removal and biopsies. Nex to this there will be several blooddrawings (1
baseline and 2-3 for tracer plasma stabilization and concentration analysis)
and a baseline ECG.

Time investment
The study consists of a total of two study procedure related visits: one visit
for the intravenous administration of bevacizumab-IRDye800CW and one visit for
consecutive fluorescence, optoacoustic and white-light surveillance endoscopy.
The time investment of the participating subjects is considered reasonable. The
tracer administration visit will take 1.5 hours. The visit for the endoscopy
procedure will take up to 1.5 hours, this is approximately 0.5 hour longer than
patients who undergo only the standard surveillance endoscopy. Next to this
there will be a baseline blooddrawing and baslinen ECG measurement.

Risks
The additional risks of participating in this study are mainly related to the
administration of Bevacizumab-IRDye800CW .. Animal toxicological studies on
bevacizumab-IRDye800CW and preclinical tracer evaluation data showed no adverse
effects.Bevacizumab-800CW has administered intravenously to 38 patients in a
tracer dose (4,5 mg) in several clinical trials (NCT01508572, NCT01972373,
NCT02113202, NCT02129933). No toxicity of the tracer bevacizumab-800CW was
observed in any of the patients. The phase 1 dose-escalation study with
cetuximab-800CW was performed in human subjects with head and neck squamous
cell carcinoma (NCT01987375). Events may be expected after administration,
based on our experience with administrating a higher doses of unlabeled
bevacizumab. The overall safety profile of bevacizumab is based on data of over
4,500 patients with various malignancies, predominantly treated with
bevacizumab 5-15 mg/kg body weight every 2-3 weeks in combination with
chemotherapy in clinical trials. The most serious adverse reactions were:
gastrointestinal perforations, hemorrhage (predominantly tumor-associated
hemorrhage and minor mucocutaneous hemorrhage), arterial thromboembolism and
necrotizing fasciitis, usually secondary to wound healing complications. The
most frequently observed adverse reactions across clinical trials in patients
receiving therapeutic dose of bevacizumab were hypertension, fatigue or
asthenia, diarrhea and abdominal pain. Hypersensitivity reactions to
bevacizumab can occur within a short term after administration (up until 1
hour). Also, hypertension can occur after bevacizumab administration.

The proposed investigational endoscopy procedure is a similar procedure as
standard surveillance sigmoid endoscopy. In this procedure 4 biopsies will be
taken and maximal 6 extra small polyps will be cold snare resected. This has in
general a minimal risk of bleeding, especially since routinely all polyps > 5mm
will also be resected. Though, if this complication occurs, which is thus very
uncommon, the gastroenterologist has several tools to handle this problem
adequately. The endoscopic systems used in this study, the NIR fiber bundle,
the spectroscopy probe and the optoacoustic endoscope, will be certified by the
technical department of the UMCG before use.

Benefit
The investigational endoscopy procedure does not have direct benefits for the
participating patients. Also, it won*t interfere with standard clinical care
since the fluorescence and optoacoustic endoscopy procedure will be followed by
standard white-light surveillance endoscopy of the rectum.