Comparison of Use of Indocyanine Green and 99mTc-labeled Radiotracer for Axillary Lymphatic Mapping in Patients With Breast Cancer
Phase: NA
Conditions: Stage I Breast Cancer, Stage II Breast Cancer
Sponsor: Case Comprehensive Cancer Center

CASE 10114 Protocol v5 11-18-16                                                                                           Page 1 of 8 
CASE COMPREHENSIVE CANCER CENTER 
__________________________________________________________________ 
 
 
STUDY NUMBER:  
CASE 10114 
 
STUDY TITLE: 
 
COMPARISON OF USE OF INDOCYANINE GREEN  
 
AND 99mTc-LABELED RADIOTRACER FOR AXILLARY 
LYMPHATIC MAPPING IN PATIENTS WITH BREAST 
CANCER 
 
PRINCIPAL INVESTIGATOR:  
Stephen R. Grobmyer, MD 
 
 
Department of General Surgery 
 
 
 
 
Cleveland Clinic  
 
 
 
 
9500 Euclid Avenue 
 
 
 
 
 
 
Cleveland, OH 44195 
 
 
 
 
 
 
(216) 636-2843, grobmys@ccf.org 
 
 
CO-PI: 
Alicia Fanning, MD  
 
Department of General Surgery 
 
 
 
Cleveland Clinic  
 
 
 
9500 Euclid Avenue 
 
 
 
 
 
Cleveland, OH 44195 
 
(216) 444-3334, fannina@ccf.org  
 
 
CO- INVESTIGATORS: 
Robyn Stewart, MD 
 
Department of General Surgery 
 
 
Cleveland Clinic  
 
 
 
9500 Euclid Avenue 
 
 
 
 
 
Cleveland, OH 44195 
 
(216) 444-6293, stewarr5@ccf.org 
                     
 
Stephanie Valente, DO 
 
Department of General Surgery 
 
 
Cleveland Clinic  
 
 
 
9500 Euclid Avenue 
 
 
 
 
 
Cleveland, OH 44195 
 
(216) 444-0769, valents3@ccf.org    
 
 
Diane Radford, MD 
 
Department of General Surgery 
 
Cleveland Clinic 
 
9500 Euclid Avenue 
 
Cleveland, OH 44195 
 
(216) 444-7506, radford2@ccf.org 
 
STATISTICIAN: 
Chao Tau 
 
Quantitative Health Sciences 
 
Cleveland Clinic 
 
9500 Euclid Avenue 
 
Cleveland, OH 44195 
 
(216) 445-8350, tuc@ccf.org 

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SUPPORT: 
CCF Breast Services Division 
 
CLINICAL FACILITY: 
Cleveland Clinic  
 
 
 
 
 
 
APPROVALS: 
Protocol Review and Monitoring Committee: Approved 
 
 
Cleveland Clinic IRB: Approved 
 
 
STUDY COORDINATORS:  
 
 
Courtney Yanda 
Cleveland Clinic 
9500 Euclid Avenue 
Cleveland, OH 44195 
(216) 444-3159, yandac@ccf.org 
Barbara BeVier 
Cleveland Clinic 
9500 Euclid Avenue 
Cleveland, OH 44195 
(216) 445-9159, bevierb@ccf.org 
 
 
 
TABLE OF CONTENTS 
 
 
 
        
 
 
1. 
Introduction  
 
 
 
 
 
 
 
 
 
 
 
2.0 
Objective 
 
 
 
 
 
 
 
 
 
 
 
3.0  
Study Design  
 
 
 
 
 
 
 
4.0 
Registration   
 
 
 
 
 
 
5.0 
Adverse Events and Reporting Requirements 
 
 
 
 
 
 
 
6.0             Data Reporting/Regulatory Considerations 
 
7.0 
Statistical Considerations 
 
 
 
 
 
 
 
 
 
 
 
 
8.0 
References 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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1.0  
INTRODUCTION  
 
Background: 
Lymphatic mapping and axillary sentinel node biopsy has become a standard procedure for the 
surgical staging of patients with early stage breast cancer. It has been shown to be safe and accu-
rate and limits significantly the need for traditional axillary dissection in many patients with ear-
ly stage breast cancer. Traditionally, lymphatic mapping has involved the use of 99mTc-labeled 
radiotracer, or visible blue dye (methylene blue or isosulfan blue dye), or a combination of 
99mTc-labeled radiotracer and visible blue dye. Sentinel nodes are defined as axillary lymph 
nodes in which there is the accumulation of one or more of these tracers or dyes. 
The use of 99mTc-labeled radiotracer has a long history and is one of the more commonly used 
techniques of identifying sentinel nodes. However, the use of radiotracers has significant limita-
tions including the requirement for nuclear licensing; the need to prepare, handle and dispose of 
nuclear material, patient inconvenience with painful injections typically occurring before sur-
gery.  
There is a growing body of literature supporting the role of indocyanine green (ICG) dye for 
lymphatic mapping in Europe and Japan.  ICG is a fluorescent dye which can be detected using 
near infrared cameras.  The use of ICG in conjunction with a near infrared camera offers many 
potential advantages over the use of other techniques including the ability to inject the material 
while the patient is under anesthesia in the operating room, the ability to use near infrared imag-
ing to directly visualize lymphatic flow, and no special material handling is required.  ICG is 
FDA approved for vascular fluorescence and liver function studies in humans.   
ICG is a di-sulfonated heptamethine indocyanine with moderate optical properties, which emits 
≈ 800 nm. There are many uses for ICG during image-guided surgery. After intravenous injec-
tion, it can be used for NIR angiography of blood vessels, identification of the extrahepatic bile 
ducts, and identification of liver metastases. After subcutaneous injection, ICG has been reported 
by numerous authors to be safely used for sentinel lymph node (SLN) mapping(1-5) and as-
sessing lymphatic function.”(6)  
ICG has been used for the last 50+ years as a visible dye (green for ICG). Thus, there is an un-
precedented body of clinical data regarding their safety when used at millimolar concentrations.” 
(6) The use of ICG for lymphatic mapping in breast cancer patients has been demonstrated to be 
safe in numerous human studies performed in Asia and Europe (7-11). There is a recent retro-
spective case series report of using ICG for lymph node mapping in patients with melanoma 
from the Cleveland Clinic documenting its safety for in human use and lymphatic mapping (12).  
A recent study from Europe has demonstrated high concordance between the number nodes 
identified with ICG and 99mTc-labeled radiotracer (11).  There has been very limited work on 
the use of ICG for axillary lymph node mapping in humans in the United States, and no com-
parison studies of ICG with 99mTc-labeled radiotracer. The goal of the project is to confirm 
that axillary lymphatic mapping with ICG leads to a similar number of nodes being labeled 
as sentinel as lymphatic mapping with 99mTc-labeled radiotracer. Successful completion of 
this project would support the use of 99mTc-labeled radiotracer with ICG for LN mapping in 
early stage breast cancer patients. 
 

CASE 10114 Protocol v5 11-18-16                                                                                           Page 4 of 8 
 
2.0  
OBJECTIVE 
 
Hypothesis:  
ICG is equivalent to 99mTc-labeled radiotracer for axillary lymphatic mapping in patients 
with early stage breast cancer in terms of the numbers of nodes identified as sentinel by each 
method being similar.  
 
 
3.0 
STUDY DESIGN 
 
 
Methods: 
Patients with a confirmed diagnosis of clinical stage 1 or 2 breast cancer who are undergoing 
breast cancer surgery with lumpectomy or mastectomy and planned axillary sentinel node 
biopsy procedure will be eligible for the study.  Patients with cancer > 3cm, clinically posi-
tive nodes, prior surgery for breast cancer in the index breast, bilateral sentinel lymph node 
mapping, thyroid dysfunction, hypersensitivity to iodine, and hepatic or renal insufficiency 
will be excluded from the study.  
• 
We plan to enroll approximately 130 adult female patients who will be identified in the 
breast clinic at Cleveland Clinic Main Campus.   
• 
Informed consent will be obtained. 
• 
Patient information will be collected from the medical record and entered in Redcap. 
Patients will undergo lymphatic mapping with  99mTc sulfur colloid in accordance with routine 
clinical practice.  Injections of 99mTc sulfur colloid will take place the afternoon prior to planned 
next morning surgery or on the morning of surgery.  Patients will undergo lymphoscintigraphy in 
accordance with standard clinical practice.    
Immediately prior to operation, after the induction of anesthesia in the operating room, up to 1cc 
of 0.5% ICG solution will be injected subdermally or intradermally close to, above the tu-
mor,into the subareolar region or into the periareolar region after disinfection of the breast skin.  
ICG movement will be facilitated by manual massage and monitored with fluorescence imaging. 
ICG fluorescence will be elicited and detected by PDE camera (Hamamatsu Photonics, Hama-
matsu, Japan) which has FDA 510(k) clearance to visualize ICG fluorescence in the humans for 
vascular imaging.  The lymphatic drainage, made evident by the fluorescent dye, will be moni-
tored in real time on a monitor. The fluorescence will be followed towards the axilla and time for 
the fluorescence to reach the axilla will be recorded. In accordance with standard practice, an 
incision will be made in the axilla. Fluorescent lymph nodes (ICG positive) will be localized and 
removed and sent for pathologic analysis as per standard surgical practice. Node removal will 
continue until no residual fluorescence is visible in the axilla.  Excised nodes will be tested for 
radioactivity using a standard gamma-detecting probe and the counts per minute will be record-
ed. Finally, the axillary region will be inspected with the gamma probe to determine if there are 
any residual radioactive nodes. Residual sentinel nodes will be removed. With regard to radioac-
tivity, sentinel nodes are defined as any node with at least 10% of the hottest nodal counts.   
The following information will be collected for each removed node: 

CASE 10114 Protocol v5 11-18-16                                                                                           Page 5 of 8 
1. Fluorescence (yes/no)  
2. Counts per minute  
 
3. Pathologic status  of node (no metastasis, isolated tumor cells, micrometastasis, macro-
metastasis) 
Also collected will be procedure related data including transit time to axilla from time of injec-
tion, size of tumor, location of tumor, tumor histology, breast size, patient age, BMI. 
Consent: 
All subjects for this study will be provided a consent form describing this study and providing 
sufficient information for subjects to make an informed decision about their participation in this 
study.  The formal consent of a subject, using the IRB-approved consent form, will be obtained 
before that subject is submitted to any study procedure.  Consent will be obtained by the study 
coordinator or investigator of the study in the breast clinic on Main Campus at Cleveland Clinic.  
Consent will be documented in the medical record. 
Finance: 
There are no additional costs to the patients.  Mitaka USA will provide the PDE neo camera and 
ICG kit. 
Funding Source: 
CCF Breast Services Division 
 
 
4.0       REGISTRATION   
 
All patients enrolled on study will be entered into the Oncore database.  
 
 
5.0 
ADVERSE EVENTS AND REPORTING REQUIREMENTS  
 
 
Adverse Events and Data Monitoring Committee (DMC): 
Although we do not anticipate any adverse events, should any unexpected event occur, we will 
report the event in a timely manner per IRB policy.   A DMC will not be used during this study.  
 
 
6.0 
DATA REPORTING/REGULATORY CONSIDERATIONS 
  
Quality Control and Quality Assurance: 
The study coordinator will be responsible to keeping data confidentially documented and report-
ed in compliance with HIPAA and any other applicable regulatory requirements. The data will 
be kept for approximately 2 years after study completion, and will be accessible only to study 
staff.   

CASE 10114 Protocol v5 11-18-16                                                                                           Page 6 of 8 
Ethical Considerations: 
This study will be conducted according to US and international standards of Good Clinical Prac-
tice for all studies.  Applicable government regulations and Cleveland Clinic research policies 
and procedures will also be followed. 
This protocol and any amendments will be submitted to the Cleveland Clinic Institutional Re-
view Board (IRB) for formal approval to conduct the study.  The decision of the IRB concerning 
the conduct of the study will be made in writing to the investigator.  
  
7.0        STATISTICAL CONSIDERATIONS 
 
Sample size: 
The study will be designed to determine whether ICG based axillary sentinel lymph node biopsy 
yields a similar number of nodes as 99mTc sulfur colloid lymph node biopsy.  While SLN identi-
fication using 99mTc sulfur colloid is a common technique, neither of the two methods used in 
this study can be assumed to identify the “correct” set of nodes as sentinel. Therefore, for the 
purposes of this study, the sentinel status of a node will be defined as being flagged as sentinel 
by either one or both of the ICG or 99mTc methods.  By this definition, sentinel nodes identified 
by 99mTc but not ICG are an error by omission for the ICG method and nodes flagged by ICG but 
not 99mTc are an error by the 99mTc method. 
Let A be the number of Tc-positive and ICG-positive SNs detected, B be the number of Tc-
positive and ICG-negative SNs detected, and C be the number of Tc-negative and ICG-positive 
SNs detected. The total number (N) of SNs detected is therefore N = (A + B + C); the proportion 
of SNs (PTc) detected by the Tc method is (A + B)/N; and the proportion of SNs (PICG) detected 
by the ICG method is (A + C)/N. 
As significant differences in the numbers of nodes identified by each method would have impli-
cations regarding  the safety of ICG as a radiotracer, the null hypothesis is that the two methods 
identify an equivalent nodes of nodes as being sentinel nodes.  Here equivalence is defined as the 
difference in the proportions of nodes flagged between the 99mTc and ICG methods falling within 
an interval (−δ, +δ), where δ is taken to be 5%. Formally the null hypothesis for equivalence is:  
PICG − PTc < −δ and PICG − PTc ≥ +δ 
If the null hypothesis is rejected we may concluded for the hypothesis of equivalence that:  
δ < (PICG − PTc) < δ 
The equivalence hypothesis can be transformed into two one-sided hypotheses:  
(A) 
that the difference between the proportions is beyond the lower equivalence margin: null 
hypothesis A: PICG − PTc < −δ, which, if rejected, permits the conclusion PICG − PTc ≥ −δ 
(B) 
that the difference between the proportions is greater than the upper equivalence margin: 
null hypothesis B: PICG − PTc ≥ +δ which, if rejected, permits the conclusion 
PICG − PTc ≤ +δ. 

CASE 10114 Protocol v5 11-18-16                                                                                           Page 7 of 8 
Hypotheses A and B will be tested by comparing a two-sided 90% confidence interval (CI) for 
the difference in the proportions of SNs flagged by the two methods, with the 5% equivalence 
region (−δ, +δ). Tests for equivalence will be based on the methods for analyzing clustered 
paired binary data described in Obuchowski(13). To conclude for equivalence the 90% CI needs 
to be contained entirely within the defined equivalence region, which is equivalent to both null 
hypotheses (A) and (B) being rejected. 
To estimate the number of SNs that will need to be examined, we assumed a detection rate (A + 
B)/N of 97% for the Tc method, a discordance rate between the two methods (B + C)/N of 6%, 
and set δ at 5%. We found through simulation that 255 SNs will give adequate power (>80%) to 
demonstrate equivalence between the two methods, with a type I error (α) rate of 5%.  This is 
estimated to require ~130 patients as the average SN yield is expected to be approximately 2 per 
patient. 
 
 
8.0 
REFERENCES 
 
1. Nimura H, Narimiya N, Mitsumori N, et al. Infrared ray electronic endoscopy combined 
with indocyanine green injection for detection of sentinel nodes of patients with gastric 
cancer. Br J Surg. 2004;91:575–9.  
2. Miyashiro I, Miyoshi N, Hiratsuka M, et al. Detection of sentinel node in gastric cancer 
surgery by indocyanine green fluorescence imaging: comparison with infrared imaging. 
Ann Surg Oncol. 2008;15:1640–3.  
3. Kusano M, Tajima Y, Yamazaki K, et al. Sentinel node mapping guided by indocyanine 
green fluorescence imaging: a new method for sentinel node navigation surgery in gastro-
intestinal cancer. Dig Surg. 2008;25:103–8.  
4. Fujiwara M, Mizukami T, Suzuki A, Fukamizu H. Sentinel lymph node detection in skin 
cancer patients using real-time fluorescence navigation with indocyanine green: prelimi-
nary experience. J Plast Reconstr Aesthet Surg. 2008  
5. Tsujino Y, Mizumoto K, Matsuzaka Y, et al. Fluorescence navigation with indocyanine 
green for detecting sentinel nodes in extramammary Paget’s disease and squamous cell 
carcinoma. J Dermatol. 2009;36:90–4.  
6. Gioux et al. “Image-Guided Surgery using Invisible Near Infrared Light: Fundamentals 
of Clinical Translation.” Mol. Imaging 9(5): 237, 2011. 
7. Tong et al. “Use of Fluorescence Imaging in Combination with Patent Blue Dye versus 
Patent Blue Dye Alone in Sentinel Lymph Node Biopsy in Breast Cancer.” Journal of 
Breast Cancer. 17(3):250, 2014.  
8. Chi et al. “Use of Indocyanine Green for Detecting the Sentinel Lymph Node in Breast 
Cancer Patients: From Preclinical Evaluation to Clinical Validation.” PLOS One. 8(12): 
e83927, 2013. 

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9. Wishart et al. “A Feasibility Study of ICG Fluorescence Mapping for Sentinel Node De-
tection in Early Breast Cancer.” EJSO. 38: 651, 2012.  
10.  Sugie, T et al. “Comparison of the ICG Fluroscence and Blue Dye Methods in Detection 
of Sentinel Lymph Nodes in Early Stage Breast Cancer.” Ann Surg Onc. 20: 2213, 2013.  
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Method for Identifying the Sentinel Node in Breast Cancer: A Concordance and Valida-
tion Study.” EJSO 39: 1332, 2013.  
12. Korn et al. “ING SPY Elite-Assisted Sentinel Lymph Node Biopsy in Cutaneous Mela-
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13. Obuchowski, NA.  On the Comparison of Correlated Proportions for Clustered Data. Stat 
Med 1998 Jul 15;17(13): 1495-507. 
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