Fluorescence Molecular Endoscopy and Molecular Fluorescence-guided Surgery of locally advanced rectal cancer using cetuximab-IRDye800CW: a single-center feasibility safety study.

Treatment of patients with locally advanced rectal cancer (LARC) is
multidisciplinary and consists of neoadjuvant chemoradiotherapy (nCRT) followed
by surgical removal of the rectal tumor and potentially tumor positive lymph
nodes.(1)
1) After surgery, in 15 to 27% of patients that received nCRT no tumor cells
can be detected during histopathological examination.(2-7) In today*s clinical
practice, all of these patients with a pathological complete response (pCR) are
operated upon, with substantial morbidity and mortality. The 5-year survival is
83.3% for patients with a pCR, and 65.6% for those without pCR.(4) Response
after nCRT is currently evaluated using magnetic resonance imaging (MRI).
However, as MRI cannot differentiate between molecular characteristics of
tissue, prediction of treatment response can be inaccurate. In patients with a
potential cCR on MRI, additionally a high-definition white-light (HD-WL)
endoscopy is performed with biopsies of the previous tumor location. If both
MRI and HD-WL endoscopy confirm a potential cCR, patients can also be treated
with a watch-and-wait approach, including frequent follow-up with HD-WL
endoscopy and MRI.(8) This potentially prevents extensive surgical procedures
for patients in which this is not required. However, MRI and HD-WL endoscopy
often remain insufficient for identification of cCR. Therefore, novel imaging
methods are needed for accurate prediction of treatment response in order to
select patients. We believe fluorescence molecular endoscopy (FME) could be a
promising technique for evaluation of treatment response.
2) During surgery, tumor-negative resection margins are of great prognostic
value. Currently, surgeons rely on visual and tactile inspection for
differentiation between malignant and healthy tissue. When in doubt, a frozen
section can be obtained, which is time consuming and poses a high risk of
sampling error. However, 14.7% of patients still have tumor-positive resection
margins, increasing the risk of local recurrence and worsening outcome.(9)
Therefore, there is a need for novel imaging techniques that can be used
intraoperatively to improve margin assessment. We believe molecular
fluorescence-guided surgery (MFGS) could be a promising technique for
evaluation of resection margins.

The study consists of and endoscopic part (1) and an intraoperative part (2).
1) Endoscopic part: to determine the feasibility and safety of molecular
fluorescence endoscopy using the fluorescent tracer cetuximab-IRDye800CW
targeting the epidermal growth factor receptor (EGFR) in patients with LARC.
2) Surgical part: to determine the feasibility and safety of molecular
fluorescence-guided surgery using the fluorescent tracer cetuximab-IRDye800CW
targeting EGFR for intraoperative evaluation of resection margins.

The study design is similar to the previously approved study for molecular
fluorescence endoscopy of patients with LARC (NL43407.042.13). The current
study is a non-randomized, non-blinded, prospective, single-center feasibility,
safety study in patients with locally advanced rectal cancer that require
surgical excision.15 patients will receive an intravenous injection of first 75
mg unlabeled cetuximab followed by 15 mg cetuximab-IRDye800CW. Furthermore, 3
patients will be included that will not receive the tracer.
1) For the endoscopy part patients will undergo a fluorescence endoscopy
procedure that is scheduled after nCRT. This fluorescence endoscopy procedure
will be performed in the operation theater previously to surgery after the
patient is anesthetized or will be performed during restaging. Fluorescence
signals of tumor and normal rectal tissue will be compared. Confocal laser
endomicroscopy will optionally be used to obtain in vivo histological
information. Multi-diameter single fiber reflectance, single fiber fluorescence
(MDSFR-SFF) spectroscopy will be performed to quantify fluorescence signals on
biopsies taken from tumor and normal tissue.
2) The surgical part of the study will be conducted during surgery, directly
after the endoscopy. Surgery will be performed according to standard clinical
care, decisions will not be based on fluorescence imaging. There will be at
least two imaging moments: previously to resection of the tumor (tumor and
lymph node detection) and after resection of the tumor (evaluation of the
resection plane). MDSFR-SFF spectroscopy will be used to quantify fluorescence
signals from the fresh surgical specimen.
Thereafter, extensive ex vivo analyses will be performed in order to correlate
fluorescence signals with histology, and to gain more insight in tissue
distribution of the tracer.

Tracer administration: the EGFR-targeted fluorescent tracer
cetuximab-IRDye800CW will be administered intravenously 3 days prior to the
endoscopy/surgery procedure at the UMCG. Afterwards patients will be monitored
for one hour by measurements of vital parameters (i.e. heartrate, blood
pressure and temperature) and ECG for potential side-effects. Additionally,
three patients will be included that won't receive the tracer as a negative
control.
1) Endoscopy procedure: three days after tracer injection patients will
undergo a sigmoidoscopy at the surgical theater after the patient is
anesthetized or during restaging. First, the gastroenterologist will use
routine high-definition white-light (HD-WL) inspection, followed by
fluorescence inspection. Confocal laser endomicroscopy (CLE; optionally) may be
performed to obtain in vivo histological information of suspicious lesions.
Additionally, biopsies will be taken from the tumor (max. 8), from normal
tissue (max. 4) and from additional fluorescence lesions when present (max 4).
Quantification of fluorescence using spectroscopy will be performed either in
vivo or ex vivo on biopsies, depending on the lesions.
2) Surgical procedure: the surgical procedure will be performed according to
standard clinical care. Fluorescence inspection will be performed before
resection of the tumor for detection of lymph nodes, and after resection for
evaluation of the resection planes. MDSFR/SFF spectroscopy can be performed in
vivo in case a positive resection margin is suspected based on fluorescence
signals. Additionally, biopsies can be taken when fluorescence is detected in
additional lesions and/or the resection plane.

Burden: Patients do not need to make extra visits to the UMCG if the tracer
administration for both the endoscopy and surgery procedure can be scheduled on
the day of admission for the surgery. If this is not possible due to logistics,
we will try to schedule the tracer administration on the same day of outpatient
clinic visits. However, if we cannot arrange this the patients will need to
make an extra visit for the tracer administration which will take about 2-3
hours. The endoscopy will take place at the surgical theater after the patient
is anesthetized or the procedures will be scheduled during restaging after nCRT
which is part of the normal clinical workflow. Hereby we ensure that the
endoscopy is not an extra burden for the patients. Prior to the endoscopy
procedures, patient will have to undergo bowel cleaning by a phosphate enema.
Risks: The intravenous injection and the use of a cannula are known to carry a
small risk of infection and hematoma. Theoretically, a possible SAE for
injection of cetuximab-IRDye800CW could be an allergic and anaphylactic
reaction. Therefore, tavegil, adrenalin and prednisone will always be present
at the site of the injection. However, this is considered a very low risk and
was not seen in previously injected patients. No preclinical or clinical study
reported higher than grade 2 adverse events.
The study procedures will prolong a standard endoscopy with
approximately 15 minutes, due to fluorescence imaging, confocal laser
endomicroscopy and biopsies. The risks of the investigational endoscopy
procedures are comparable to the minimal risks of a standard clinical
sigmoidoscopy. These superficial biopsies that will be taken pose a small risk
of bleeding. Most bleedings coagulate spontaneously. If not, which is very
uncommon, the gastroenterologist has several tools to coagulate the small
bleeding.
Next to this, the operation time may be prolonged with approximately 15
minutes, due to fluorescence imaging. Therefore, the time under general
anesthesia will be prolonged. Additional biopsies from the resection surface or
other fluorescence lesions can only be taken if judged as safe by both surgeons
operating the patient. Therefore, no additional risk will be expected for the
patient, since also the tracer is already administered to the patient because
of the endoscopy procedure.
Benefit: Patients will have not benefit from this study directly. Surgery will
be planned and performed according to standard clinical care. During surgery,
no decisions will be made based on the fluorescence imaging. The benefit of
this study will be the establishment of usefulness of cetuximab-IRDye800CW
during endoscopy for prediction of treatment response and evaluation of
resection margins for future patients.