Pilot Study of Immunization of High Risk Breast Cancer Patients With a Sialyl Lewisª -Keyhole Limpet Hemocyanin Conjugate Plus the Immunological Adjuvant QS-21
Phase: NA
Conditions: Breast Cancer
Sponsor: Memorial Sloan Kettering Cancer Center

Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
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Pilot Study of  Immunization of High Risk Breast Cancer Patients with a Sialyl Le wis a- 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Memorial Sloan-Kettering Cancer Center 
1275 York Ave. 
New York, NY 10021 
Approval date: 24-Sep-2018 
Keyhole Limpet Hemocyanin Conjugate plus the Immunological Adjuvant QS-21 
MSKCC THERAPEUTIC/DIAGNOSTIC PROTOCOL 
Principal Investigator: 
Teresa Gilewski M.D. (Department of Medicine) 
Co-Principal Investigator(s): Tomas Lyons, MD 
Investigator(s): 
Larry Norton M.D. 
Kathy Panageas Ph.D. (Department of Biostatistics) 
Martin Fleisher Ph.D. (Department of Clinical Chemistry) 
Consenting Professional(s): 
Teresa Gilewski M.D. 
Larry Norton M.D. 
Tomas Lyons, MD 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
Table of Contents 
MSKCC THERAPEUTIC/DIAGNOSTIC PROTOCOL ............................................................... 1 
1.0 
PROTOCOL SUMMARY AND/OR SCHEMA ................................................................ 2 
2.0 
OBJECTIVES AND SCIENTIFIC AIMS .......................................................................... 3 
3.0 
BACKGROUND AND RATIONALE ............................................................................... 3 
4.1 
OVERVIEW OF STUDY DESIGN/INTERVENTION ..................................................... 7 
4.2 
DESIGN ............................................................................................................................. 7 
4.3 
INTERVENTION ................................................................................................... 7 
5.0 
THERAPEUTIC/DIAGNOSTIC AGENTS ....................................................................... 7 
6.1 
CRITERIA FOR SUBJECT ELIGIBILITY ....................................................................... 8 
6.2 
SUBJECT INCLUSION CRITERIA ...................................................................... 8 
6.3 
SUBJECT EXCLUSION CRITERIA ..................................................................... 9 
7.0 
RECRUITMENT PLAN ..................................................................................................... 9 
8.0 
PRETREATMENT EVALUATION ................................................................................ 10 
9.0 
TREATMENT/INTERVENTION PLAN ................................................................................ 11 
10.0 
EVALUATION DURING TREATMENT/INTERVENTION ......................................... 12 
11.0 
TOXICITIES/SIDE EFFECTS ......................................................................................... 14 
12.0 
CRITERIA FOR THERAPEUTIC RESPONSE/OUTCOME ASSESSMENT ................ 15 
13.0 
CRITERIA FOR REMOVAL FROM STUDY ................................................................ 16 
14.0 
BIOSTATISTICS ................................................................................................................... 17 
15.1 
RESEARCH PARTICIPANT REGISTRATION AND RANDOMIZATION 
PROCEDURES ................................................................................................................................... 17 
15.2 RESEARCH PARTICIPANT REGISTRATION ................................................... 17 
16.1 
DATA  MANAGEMENT ISSUES .................................................................................. 18 
16.2 
QUALITY ASSURANCE ......................................................................................... 19 
16.3 
DATA AND SAFETY MONITORING ............................................................... 19 
17.1 
PROTECTION OF HUMAN SUBJECTS ............................................................................ 20 
17.2 
PRIVACY ................................................................................................................... 20 
17.3 SERIOUS ADVERSE EVENT (SAE) REPORTING .............................................. 20 
18.1 
INFORMED CONSENT PROCEDURES ........................................................................ 21 
18.2 
RESEARCH AUTHORIZATION ........................................................................ 21 
19.0 
REFERENCE(S) .................................................................................................................... 23 
20.0 
APPENDICES ........................................................................................................................ 26 
APPENDIX A: PATIENT CALENDAR ...................................................................................... 26 
APPENDIX B: PATIENT DIARY ............................................................................................... 26 
 
 
 
 
 
1.0 
PROTOCOL SUMMARY AND/OR SCHEMA 
 
This is a pilot study of injection of sialyl lewisa  (sLea) conjugated to keyhole limpet hemocyanin 
(KLH) plus the immunological adjuvant QS-21. Eligible patients include those with metastatic 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
breast cancer who have no evidence of disease or stable disease on hormone therapy. Initially 3 
patients will be entered at a lower dose (10 mcg) of sLe
a. If safe, 10 patients will be accrued at the 
full dose of 30 mcg . If 6 or more of these 10 patients have an immune response at any time point 
then an additional 10 patients will be accrued and injected with a higher dose of QS-21. 
 
Injections will be administered subcutaneously during weeks 1, 2, 3, 7 and 19. Prior to and during 
therapy, blood samples will be obtained to assess immune response. 
 
The primary endpoints of this trial are safety and immune response. Clinical response will be 
monitored but will not be a primary endpoint. A secondary endpoint is the evaluation of blood 
samples for circulating tumor cells. 
 
2.1 
OBJECTIVES AND SCIENTIFIC AIMS 
 
The primary objectives of this trial are to determine the following endpoints after injection with 
sLea-KLH plus QS-21: 
 
• 
Safety 
 
• 
The IgG and IgM antibody response to sLea  and breast cancer cells expressing 
this antigen 
 
The secondary objective of this trial is to monitor the presence of circulating tumor cells (CTC) 
prior to, during and after injection with sLea-KLH plus QS-21. 
 
Clinical response will be monitored but will not be an endpoint. As part of standard care, patients 
on this trial will be followed for progression of disease. It is unknown how many patients will 
enroll with evidence of disease. Therefore, we will report any responses that are observed for 
these patients in a purely descriptive manner (see section12). This is consistent with the pilot 
vaccine trials we have previously conducted. 
 
 
3.0 
BACKGROUND AND RATIONALE 
 
3.1 
B re as t Cance r 
 
Invasive breast cancer is now predicted to affect one in seven American women at some 
point in their lifetime.1 A significant number of patient will develop metastatic breast 
cancer, which in contrast to early stage breast cancer is generally considered incurable.2 
While patients with AJCC Stage IV breast cancer who have no evidence of disease or 
stable disease on hormone therapy can live for many years, ultimately their disease will 
likely become resistant to current treatment options. Additional therapeutic strategies for 
these patients are needed. 
 
3.2 
Tre atme nt B ackground 
 
Dr. Livingston and colleagues have shown that some patients have antibodies in their serum 
which react with highly restricted ganglioside antigens present on melanomas and a variety 
of other cancers. In some cases, they and others 3,4 noted that the presence of  these 
antibodies has been associated with an unexpectedly favorable course.  As only few patients 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
have these antibodies in their serum, we are attempting to  induce antibody formation by 
immunizing the patients with tumor vaccines containing the relevant antigens. 
 
Adjuvant immunotherapy of breast cancer with immunizations must be based on the 
identification of antigens expressed by breast cancers, which are immunogenic.  While many 
antigens have been identified on breast cancers using murine monoclonal antibodies, very 
few of these are known to be recognized by the human immune system. Several of the 
antigens commonly identified on breast cancer cells include the gangliosides GM2 and  
Globo H5,  blood group-related antigens TF, sTn, Lewis y (Ley)  6 and the protein MUC17. 
 
3.2.1 Keyhole Limpet Hemocyanin  (KLH)   Various methods have been used to increase 
the immunogenicity of antigens, in particular to induce an IgG response. The approach 
that Dr. Livingston and colleagues have found to be most successful at inducing an 
IgG response against the closely related T antigen in the mouse has been covalent 
attachment to keyhole limpet hemocyanin (KLH).  KLH is an immunogenic protein 
which is derived from the blood of the keyhole limpet. This approach also resulted in 
anti-T antigen delayed type hypersensitivity reactions and partial protection against 
cancer cells expressing T antigen. 8 Studies of immunological tolerance and of ways 
to overcome it have shown that in a variety of experimental systems, T-cell (not 
related to T antigen) unresponsiveness is more rapidly induced and more easily 
maintained than B cell unresponsiveness .
9 Levels of circulating antigens suitable for 
maintaining T-cell tolerance frequently fail to maintain B cell tolerance. Consequently 
if T-cell help is provided (as by potent irrelevant antigens such as KLH covalently 
attached to the desired immunogen), antibodies can be induced to tolerated T-cell 
dependent antigens. This approach has been successfully used to induce IgG 
antibodies against a variety of carbohydrate antigens, and the titer of antibody induced 
is greatly augmented with the use of immunological adjuvants.10 
 
3.2.1.1 Conjugationof antigens to KLH can be accomplished using a direct method 
or with a  bifunctional linker MMCCH (M2C2H): (4-(4-N-maleimidomethyl) 
cyclohexane-1-carboxyl hydrazide. This latter method of conjugation was 
developed to improve the antigen epitope:KLH ratio.11    In mice, this method 
of conjugation resulted in higher antibody titers following immunization with 
an sTn based vaccine. This method of conjugation has also been evaluated at 
MSKCC with the Globo H-KLH plus QS-21 construct in breast cancer patients 
(protocol 97-86). 
 
3.2.2 Immune Adjuvants 
 
QS-21  In addition to KLH, the immunological adjuvant QS-21 has also been used to 
increase immunogenicity of various antigens. QS-21 is a saponin fraction from the 
bark of the South American tree Quillaja saponaria Molina. 
12    Immunization of mice 
with synthetic MUC1 peptides conjugated to KLH and mixed with QS-21 resulted in 
significant antibody titers against MUC1 peptide and MUC1 positive tumor cells in 
comparison with minimal immune response to  unconjugated MUC1 peptides mixed 
with BCG or QS-21. 
13 Groups of melanoma patients have been immunized with 
various constructs containing the adjuvants DETOX, BCG and QS21 or no adjuvant at 
all. QS-21 was a significantly more effective adjuvant than the others, producing 
significantly higher titer IgM and IgG antibodies.14   A dose of 100 mcg of QS-21 was 
found to be minimally toxic and immunogenic. 15  Therefore, QS-21 was selected for 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
future trials due to its adjuvanticity and lack of toxicity.  It has also proven nontoxic 
and highly effective at augmenting the immunogenicity of an FeLV subunit vaccine in 
cats 16 and an HIV-1 recombinant vaccine in Rhesus monkeys. 
 
3.3 
M SKCC Expe rie nce with Immunization of B re as t Cance r Patie nts 
 
Immunization against antigens found on tumor cells represents a potential therapeutic 
strategy for breast cancer patients. We have conducted several clinical trials at MSKCC 
in high risk breast cancer patients to assess the safety and immunogenicity of various 
single antigens conjugated to keyhole limpet hemocyanin (KLH) and mixed with the 
immune adjuvant QS21. Antigens included the peptide MUC-1 and the carbohydrates 
Globo-H, GM2 and STn[clustered] (protocol numbers 94-130, 95-67, 97-123, 99-023). 
These immunizations were well-tolerated with the most common toxicities including 
transient local skin reactions at the injection site and mild flu-like symptoms. Although, 
there were no definite autoimmune reactions attributable to the injection, several patients 
developed hypothyroidism or hyperthyroidism. The majority of patients developed IgM 
and IgG antibody titers reactive with the immunizing antigen. There were no significant 
differences in the immune response observed among patients with no evidence of disease 
or those with stable disease. No definite T cell responses were elicited in the MUC-1 
trials.17-20 
 
We subsequently evaluated patients on protocol 01-019 with a polyvalent construct 
containing 7 antigens conjugated to KLH and mixed with either QS21 or GPI-0100 (a semi- 
synthetic saponin).21-23  The injections were well tolerated with common local skin reactions 
and mild flu-like symptoms.  Several patients receiving GPI-0100 developed a transient 
Grade 1-3 elevation in the hepatic enzymes (AST, ALT).  Following an extensive hepatic 
evaluation, it was felt that the GPI-0100 was the likely source of the hepatotoxicity. 
Therefore, the adjuvant GPI-0100 will not be incorporated into the current breast cancer 
trial. The majority of patients also developed antibody titers reactive with multiple 
immunizing antigens. 
 
 
3.4 
Rationale for Evaluation of  sialyl Lewisa (sLea)-KLH plus QS-21 
 
The carbohydrate antigen sLea  is the antigen recognized by monoclonal antibody 19.9 
and is commonly expressed on breast cancer cells. In a study of 43 breast cancer patients 
with infiltrating ductal carcinoma it was found in about 79% of samples.24 There was 
high expression in patients with greater node involvement. sLe
a  may be a ligand for 
endothelial leukocyte adhesion molecule and therefore have an impact on metastatic 
potential. 
 
sLe
a  is expressed primarily as a glycolipid on the surface of cancer cells. We are  
preparing a polyvalent vaccine for evaluation in patients with breast cancer. It contains 
four antigens expressed on mucins (MUC1, Tn, sTn and TF) and three antigens expressed 
as glycolipids (GM2, globo H and Lewis Y). There are advantages to glycolipid antigens 
including greater susceptibility to activation of complement in the presence of antibodies 
and greater induction of complement medicated cytotoxicity, as recently described.24a  We 
have recently learned that Lewis Y is poorly immunogenic, inducing antibodies in fewer 
than 30% of vaccinated patients. If sLea  proves immunogenic in this trial, we plan to 
replace Lewis Y in the polyvalent construct with sLea. 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
 
In this protocol, we propose to evaluate a construct containing the antigen sLe
a  for 
toxicity and immunogenicity. If the injections are well tolerated and immunogenic, then 
sLe
a   will be incorporated into a future polyvalent construct. This is the first time that 
sLea  is being evaluated in humans. 
 
Our prior breast cancer immunization protocols used 100 mcg of QS-21. The greatest 
toxicity primarily occurred during weeks 2 and 3. In an attempt to increase the 
immunogenicity of the construct, if six or more of the initial ten patients demonstrate an 
immune response against sLea  and the injection is well tolerated, the dose of QS-21 will 
be increased to 150 mcg during weeks 1, 7 and 19 in a second group of 10 patients. We 
do not anticipate significant increases in toxicity as prior evaluations of 200 mcg of QS- 
21 were tolerated. 
 
The potential benefits of this study include generation of an anti-tumor response by 
stimulation of the immune system.  While it is possible that this treatment will result in 
shrinkage of breast cancer tumors, delay a recurrence or decrease the risk of progression, 
it is not known whether these effects will occur. If this study demonstrates that the sLea- 
KLH conjugate is safe and immunogenic, it will be incorporated into a polyvalent 
vaccine that will be evaluated in high risk breast cancer patients for clinical benefit. 
 
3.5 
Evaluation of Circulating Tumor Cells (CTC) 
 
Circulating tumor cells (CTC) can be detected in the blood from patients with var ious 
primary or metastatic carcinomas. 
25-29  Recently the development of immunomagnetic 
separation strategies has increased the sensitivity of these assays. 28,.29  Several studies 
have reported that the presence of CTC predicted shortened disease free and overall 
survival times. These studies demonstrate that detection of CTC correlates with prognosis 
at baseline and that changes in CTC levels during and after treatment may be more 
predictive than other measures. 30-34  Monoclonal antibody based therapy (trastuzumab) 
with or without additional chemotherapy was particularly effective at eliminating CTC. 
This was associated with improved disease free and overall survival.28 
 
Use of peripheral blood immunomagnetically enriched for tumor cells using labeled 
antibodies targeting epithelial cell surface markers on breast  cancer cells and quantitated 
using a CellSpotterAnalyzer is now FDA approved for this purpose in breast cancer 
patients. Veridex, LLC. has provided MSKCC with a CellSpotterAnalyzer for this 
purpose in patients with prostate and breast cancer. To date, Dr. Martin Fleisher, Chief of 
the MSKCC Department of Clinical Laboratories, has validated the assay in prostate 
cancer patient using the same labeled antibodies which are FDA approved for use in 
breast cancer patients. 
 
Two studies have identified CTC in approximately 95% of breast cancer patients with 
Stage I/II disease prior to surgery.
35,26. After resection of all known disease, 30% of 
patients continued to have CTC that persisted for at least 2 weeks. The patients that we 
propose to evaluate are patients at a high risk for recurrence. We estimate that at least 
30% of these patients will have demonstrable CTC. 
 
In this study, CTC will be evaluated in an exploratory manner to determine whether they 
can be detected in this patient population at baseline and if so whether there is evidence 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
of a decline in those patients who have an immune response. CTC will be monitored prior 
to, during and after injection with sLe
a-KLH plus QS-21. The patient and their primary 
oncologist will be notified of all CTC results. These results will be used for research 
purposes and will not be used during the study to alter clinical care. Although some 
physicians at other centers may use CTC results to evaluate response to therapy, this is 
not considered standard care at MSKCC. 
 
4.1 
OVERVIEW OF STUDY DESIGN/INTERVENTION 
 
4.2 
Design 
This is a pilot study designed to assess the safety and immunogenicity of a sLe a-KLH plus QS-21 
construct in patients with metastatic breast cancer. Initially 3 patients will be accrued to the study 
at a 10 mcg dose of sLea. If safe, then 10 patients will be accrued at the 30 mcg dose of sLea. If 6 
or more of  these 10 patients have an immune response at any time point then an additional 10 
patients will be accrued and injected with a higher dose of  QS-21. Toxicities will be monitored 
during the study. Patients will be instructed to complete a patient diary and document any side 
effects. 
 
Patient blood samples will be evaluated for an IgG and IgM antibody response to sLea  and breast 
cancer cells expressing this antigen. Immunological response for a patient is defined as an 
antibody response of greater than 1:80 if baseline value was 0 and at least an increase of 8-fold if 
baseline value is greater than 0. Patient blood samples will also be monitored for the presence of 
circulating tumor cells pre study and then during weeks 5, 13, and 23. 
 
 
4.3 
Inte rve ntio n 
Patients will receive sLe
a-KLH plus QS-21 subcutaneously during weeks 1,2,3,7 and 19. The first 
3 patients will receive a 10 mcg dose of sLea   and subsequently all patients will receive a 30 mcg 
dose of sLea. The dose of QS-21 is 100 mcg for the initial 13 patients. If immune responses are 
observed at the 100 mcg level of QS-21 then the dose of QS-21 will be escalated to 150 mcg, in a 
subsequent group of patients during weeks 1, 7 and 19. Blood samples will be obtained pre study 
and during weeks 1,2,3,5 7, 9,13, 19 and 23 to assess for immune response. 
 
sLea  is one of multiple antigens found on breast cancer cells. Data from this trial will be used to 
determine whether the sLea antigen will be included in a future polyvalent construct. Ultimately 
we plan a multicenter randomized clinical trial to assess potential clinical benefit of 
immunization with a polyvalent construct in high risk breast cancer patients. 
 
 
5.0 
THERAPEUTIC/DIAGNOSTIC AGENTS 
 
5.1 
Clinical M ate rial 
 
5.1.1 Sialyl lewisa  (sLea)-KLH conjugate  The conjugate is prepared at Optimer 
Pharmaceutical, Inc. sLea  pentenyl glycoside was synthesized using a proprietary 
synthetic method by Optimer Pharmaceuticals, Inc. under GMP. Its identity was 
confirmed by MS and NMR The glycoside was purified by reverse phase 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
chromatography and the purity established to be greater than 99% by NMR and HPLC 
analysis. 
 
The pentenyl glycoside group on sLe
a  was bound to M2C2H heterobifunctional linker 
following ozone treatment. Keyhole limpet hemocyanin (KLH) was activated by 
introduction of thiol groups on the KLH amino groups with 2-Iminothiolane. Activated 
KLH was mixed with sLea-M2C2H and the resulting sLea-M2C2H-KLH purified and 
washed by diafiltration. The carbohydrate and protein components were quantitated and 
the conjugate was tested for toxicity and immunogenicity in mice. The sLea/KLH ratio 
was determined to be greater than 600/1. The dose of sLea-KLH  to be studied is 30 mcg. 
However, due to concerns regarding potential toxicity 3 patients will first be entered at a 
dose of 10mcg. 
 
5.1.2 QS-21 QS-21 is a saponin prepared from the bark of Quillaja saponaria Molina. It 
will be supplied by Antigenics, Inc.. All injections will be vialed with a dose of 100 mcg 
of QS-21. If there is an immune response in at least six of the ten patients, then in the 
second group of patients, an additional 50 mcg of QS-21 will be added to the preparation 
during weeks 1, 7 and 19. 
 
5.1.3 The injection will contain sLea-KLH conjugate 30 mcg (10 mcg for the first 3 
patients) and QS-21 100 mcg per 1 ml 0.9% saline. It will be stored at -80°. The total 
volume per vial is 1.1 ml. 
 
 
6.1 
CRITERIA FOR SUBJECT ELIGIBILITY 
 
Breast cancer patients with a history of metastatic disease will potentially be eligible. 
 
6.2 
Subject Inclusion Criteria 
 
• 
AJCC Stage IV bre ast cancer who have no evidence of disease 
OR 
AJCC Stage IV bre ast cancer who have stable disease on hormonal therapy. Stable disease 
must be present for at least 2 months prior to enrollment in this study.  Stable disease includes 
less than a 30% decrease or less than a 20% increase in the sum of the longest diameters of 
target lesions, taking as a reference the baseline sum of the longest diameters recorded at the 
last set of scans. Stable disease also includes no new target lesions and no unequivocal 
progression of non-target lesions. 
(These patients may or may not have an elevation in the CA153 (BR2729) or CEA values). 
 
• 
Performance status > 80 (Karnofsky). 
 
• 
Patients must have completed radiation, chemotherapy or surgery at least 4 weeks prior to 
treatment. Patients must have completed prior immunotherapy at least 6 weeks prior to 
vaccination. Ongoing hormone the rapy is pe rmitte d. 
 
• 
Pathology slides from the metastatic site must be reviewed by the Department of Pathology at 
MSKCC to confirm diagnosis. 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
 
 
6.3 
Subject Exclusion Criteria 
 
• 
Pregnant women are ineligible. Premenopausal patients must have a pregnancy test to exclude 
pregnancy within 2 weeks of starting treatment. Woman who may yet bear children and 
sexually active men must use appropriate contraception during the course of this study. 
Patients must be counseled not to become pregnant during the study. Patients must also be 
counseled that injection of sLea  may have unknown affects on the viability of a future fetus. 
 
• 
Patients who are nursing 
 
• 
Patients with other active cancers are ineligible, (excluding basal cell or squamous carcinomas 
of the skin). 
 
• 
Patients with: 
 
total lymphocyte count of < 0.5 x 106/ml. 
total white blood cell count of < 3,000 cells/cmm 
serum creatinine >1.5 x upper limit of normal 
serum aspartate aminotransferase  >1.5 x upper limit of normal 
serum alkaline phosphatase >1.5 x upper limit of normal. 
 
• 
Patients with a history of a seafood allergy. 
 
• 
Patients with clinically significant NY Heart Association Class 3 or 4 cardiac disease. 
 
• 
Patients who have previously been immunized with the sLea antigen. 
 
• 
Patients with a known history of immunodeficiency or autoimmune disease.  Current use of 
immunosuppressive medications (ie corticosteroids) is prohibited. 
 
• 
Active infection requiring antibiotic treatment. 
 
7.0 
RECRUITMENT PLAN 
 
Potential research subjects will be identified by a member of the patient‟s treatment team, the 
protocol investigator, or research team at Memorial Sloan-Kettering Cancer Center (MSKCC).  If 
the investigator is a member of the treatment team, s/he will screen their patient‟s medical records 
for suitable research study participants and discuss the study and their potential for enrolling in  
the research study.  Potential subjects contacted by their treating physician will be referred to the 
investigator/research staff of the study. 
 
The principal investigator may also screen the medical records of patients with whom they do not 
have a treatment relationship for the limited purpose of identifying patients who would be eligible 
to enroll in the study and to record appropriate contact information in order to approach these 
patients regarding the possibility of enrolling in the study. 
 
During the initial conversation between the investigator/research staff and the patient, the patient 
may be asked to provide certain health information that is necessary to the recruitment and 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
enrollment process.  The investigator/research staff may also review portions of their medical 
records at MSKCC in order to further assess eligibility.  They will use the information provided 
by the patient and/or medical record to confirm that the patient is eligible and to contact the 
patient regarding study enrollment.  If the patient turns out to be ineligible for the research study, 
the research staff will destroy all information collected on the patient during the initial 
conversation and medical records review, except for any information that must be maintained for 
screening log purposes. 
 
In most cases, the initial contact with the prospective subject will be conducted either by the 
treatment team, investigator or the research staff working in consultation with the treatment team. 
The recruitment process outlined presents no more than minimal risk to the privacy of the patients 
who are screened and minimal PHI will be maintained as part of a screening log.  For these 
reasons, we seek a (partial) limited waiver of authorization for the purposes of (1) reviewing 
medical records to identify potential research subjects and obtain information relevant to the 
enrollment process; (2) conversing with patients regarding possible enrollment; (3) handling of 
PHI contained within those records and provided by the potential subjects; and (4) maintaining 
information in a screening log of patients approached (if applicable). 
 
Patients will be recruited for this protocol primarily by their doctor at MSKCC. In addition, patients 
outside of MSKCC may be aware of this trial from other patients, their local doctors or the National 
Cancer Information Center. 
 
Once a patient has indicated an interest in participation, he/she will be referred to one of the 
consenting physicians for further evaluation and discussion of informed consent. If the patient is 
determined to be eligible, he/she will be registered according to standard MSKCC policy. 
 
Most patients with breast cancer are female, but male patients will not be excluded. Children are 
excluded from participation in this study since breast cancer is rare in this patient population and 
therefore toxicity analysis would be of questionable significance. 
 
 
8.1 
PRETREATMENT EVALUATION 
 
Within 2 weeks of the first injection: 
 
Pregnancy test for premenopausal patients 
 
 
Within 3 weeks of the first injection: 
 
• 
Complete history and physical examination 
 
• 
Complete blood cell count with differential and platelet count (CBC) 
 
• 
Comprehensive panel (COMP): aspartate aminotransferase (AST), alanine 
aminotransferase (ALT), albumin, alkaline phosphatase, bilirubin, calcium, 
creatinine, glucose, total protein, BUN, sodium, potassium, chloride, C02 
 
• 
CEA, CA153(or BR27-29 or CA27-29) 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
• 
Thyroid stimulating hormone (TSH) level 
 
• 
Urinalysis 
 
• 
Rectal examination with Hemoccult 
 
• 
Blood (3 red-top tubes) for immune response studies will be sent to Dr. Philip 
Livingston‟s laboratory in 723 Kettering building. An additional 7.5 ml of blood 
will be drawn in special tubes to go to Dr. Fleisher‟s lab in Clinical Chemistry for 
assessment of circulating tumor cells. These blood samples may be drawn on the 
day of the first injection, but prior to the injection. 
 
 
Within 4 weeks of the first injection: 
Imaging and Diagnostic Studies 
• 
CT scan of the chest/abdomen/pelvis 
• 
Radionuclide bone scan 
 
 
9.0 
TREATMENT/INTERVENTION PLAN 
The initial 3 patients will be accrued at 1 or more week intervals and will receive 10mcg of sLea   
in the sLea-KLH + 100mcg QS-21 vaccine.  Four weeks or more after accrual of the third patient, 
3 additional patients may be accrued at 1 week intervals at the full 30mcg sLea  dose . Four weeks 
after accrual of the sixth patient, 7 additional patients may be accrued at a rate no greater than two 
patients per week. 
 
The sLea-KLH plus QS-21 vaccines will be injected during weeks 1, 2, 3, 7 and 19 for a total of 
five injections.  They will be stored in a freezer at -80 degrees Centigrade. At the time of 
administration the vaccine will be thawed and 1ml drawn up into a syringe.  Each 1.1 ml vial will 
contain sLe
a - KLH conjugate 30 mcg (except for the 10 mcg dose) and QS-21 100 mcg per 1 ml 
of 0.9% saline. 
 
If a second group of 10 patients is evaluated, then the dose of QS-21 in that group will be 
increased to 150 mcg during weeks 1, 7 and 19 with the dose remaining the same for the other 
injections. The same precautions will apply to the rate of patient accrual. The initial 3 patients in 
this group will be accrued at 1 or more week intervals. Four weeks or more after accrual of the 
third patient, 7 additional patients may be accrued. 
 
All injections will be administered by a trained nurse on an outpatient basis in the Experimental 
Therapeutics Center at 53rd  Street outpatient building. All patients will be observed for 30 
minutes after each injection to monitor for acute allergic reactions. 
 
The sites of immunization (arm or thigh usually- buttocks may be used if necessary) will be 
rotated for each of the immunizations. No injection will be given at a location in which the 
draining lymph nodes have been removed. 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
 
10.0 EVALUATION DURING TREATMENT/INTERVENTION 
 
The study evaluation will include parameters to assess the safety of the injections as well as 
assessments of immunity to sLea  (Appendix A). Interval safety assessments will include the „Patient 
Diary‟ (Appendix B). Patients will be provided with Appendix A and B. Long term followup will 
also be maintained to assess toxicity. Although not a primary endpoint, patients will be monitored 
for any evidence of an anti-tumor effect. Imaging to assess extent of disease will be performed at 
baseline, then as appropriate for clinical circumstances. Patients with metastatic disease are often 
evaluated with imaging every 3 months as standard of care. Clinical benefit will also be assessed 
by physical examination and biochemical studies as outlined below. 
SUMMARY TABLE OF EVALUATION 
 
STUDY WEEK 
Pre 
1 
2 
3 
5 
7 
9 
13 
19 
23 
q3 monthsg 
VACCINATION 
1 
2 
3 
4 
5 
Clinical: 
History 
X 
X 
X 
X 
X 
X 
X 
Physical Exam 
X 
X 
X 
X 
X 
X 
X 
CBC w/ Diff 
COMP b 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
CEA, CA153 
X 
Xf 
TSH 
X 
X 
X 
Urinalysis 
X 
Pathology Re vie w X 
Pregnancy 
Test 
Xa 
Rectal/Hemoccult 
X  
Imaging: 
Bone scan 
X 
X
f
 
CT Scans 
X 
X
f
 
Response 
Assessmentsd 
Anti-sLea  
Immu nity: 
Xf 
Xf 
Serology (3 RTT)   X 
Xe 
Xe 
Xe 
X 
Xe 
X 
X 
Xe 
X 
X 
CTC Assay 
X 
X 
X 
X 
a 
Within 2 weeks of the first injection for a premenopausal patient.. 
b 
Includes total bilirubin, AST, ALT, Sodium, Potassium, Chloride, CO2, Calcium, Glucose, 
Total Protein, Albumin, Alkaline Phosphatase, Creatinine, BUN, Bilirubin. 
c 
Patients  may  be  asked  to  obtain  tissue  blocks  from  previous  diagnostic/therapeutic 
procedures. 
d 
Overall response assessment includes the measurement of tumor by non-imaging studies 
such as physical exam. 
e 
Blood must be drawn before administration of vaccine at week 1,2, 3, 7 and 19. An extra 
100ml of heparinized blood may be drawn once (during weeks 9-23) during the course of 
immunizations depending on the serologic response in individual patients,  at the 
principal investigators discretion,  for evaluation of the immune response at the clonal 
level by hybridoma production. 
f 
These studies will be performed as standard of care if deemed appropriate by the patient‟s 
physician. 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
g 
If feasible, a history and physical examination and serology will be obtained every 3 
months until evidence of disease progression. 
 
10.1 
Clinical Course 
 
Patients will be examined by oncologists at Memorial Hospital and have laboratory and 
radiographic studies performed as outlined in the above table. As this is a pilot study 
primarily concerned with effect of injection of sLea  on immune response, intensive clinical 
follow-up at Memorial Hospital will not be required. However, if feasible, patients who 
remain clinically stable will be asked to return to Memorial Hospital every 3 months for a 
history and physical examination and laboratory studies. 
 
The patient and their primary oncologist will be notified of all CTC results. These results 
are for research purposes and will not be used during the study to alter clinical care. 
 
 
10.2 
Immunologic Res ponse 
 
Serological response:  Three 10 ml red top tubes will be drawn before the first injection, 
before each subsequent injection and also during weeks 5, 9, 13 and 23. Sera will be 
evaluated for reactivity against sLe
a  in ELISA and/or immunoprecipitation-western blot 
assays. These assays detect antibodies against sLea  and confirm identity of the reaction. If 
serologic reactivity is detected, then additional blood (~10ml) may be drawn at later  
times after the injections are completed to determine the duration of this response (every 
3 months if feasible).  An extra 100ml of heparinized blood may be obtained once during 
the course of immunizations depending on the serologic response in individual patients at 
the principal investigators discretion, for hybridoma production.  This will be used for 
evaluation of the B-cell response to vaccination at the clonal level by establishment of 
monoclonal antibodies.  This will be performed in collaboration with MabVax 
Therapeutics, Inc., (San Diego, CA).  It will make possible more detailed analysis of the 
most immunogenic epitopes on sLea  and more detailed comparison of the avidity of the 
induced IgG and IgM antibodies than is possible with sera. 
 
T cell response will not be evaluated since this is a carbohydrate antigen and significant T 
cell response is not anticipated. The immune response analyses will be conducted in the 
laboratory of Dr. Livingston under Dr. Ragupathi‟s supervision. 
 
10.3 
Circ ulating Tumo r Ce ll M e thodo logy 
 
Epithelial cell adhesion molecule (EpCAM) is used to concentrate ciruclating 
epithelial tumor cells from a 7.5 ml peripheral blood specimen. A semi-automated 
system called CellSearch, uses EpCAM antibody-based immunomagentic capture of 
epithelial cells to identify and isolate circulating tumor cells (CTC). 
Immunomagnetically captured CTCs are stained and enumerated using specially 
prepared buffers and three immunofluorescent markers: DAPI to detect cells with 
intact nuclei; Cytokeratin-PE to detect epithelial cells; and CD-45-APC to detect 
leukocytes. CTCs are defined as DAPI(+), CK-PE(+) and CD-45(-) cells. Digitally 
archived images of candidate CTCs meeting these criteria also must meet standardized 
morphologic  inspection. 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
11.0 
TOXICITIES/SIDE EFFECTS 
 
In our earlier breast cancer immunization trials, common toxicities included local skin reactions 
at the injection sites, flu-like symptoms (ie, fever, fatigue, myalgias, arthralgias), and less 
frequent symptoms such as pruritis, rashes, diarrhea, nausea and vomiting. These symptoms  
were transient and usually of less than 5 days duration. In addition, minimal transient decreases 
in hematologic values, primarily white blood cell counts and lymphocyte counts were noted. It is 
unlikely that these fluctuations were due to the vaccine. Transient elevation in various laboratory 
values (ie glucose, liver enzymes) were observed and also thought unrelated to the vaccine. 
 
Since the antigen in this injection is present on the surface of normal tissues, the theoretical 
possibility of a serious autoimmune reaction exists. In our previous breast cancer trials, there 
was no consistent evidence of an autoimmune reaction due to the immunization. Several 
episodes of  hyperthyroidism or hypothyroidism have occurred on these trials. Patients were 
subsequently treated with radioactive I-131 or thyroid supplementation. It is unclear whether 
these events were related to the immunization. Therefore, the thyroid stimulating hormone level 
will be monitored pre therapy, during therapy and post therapy. 
 
In addition, no definite evidence of autoimmunity has been observed in immunization trials in 
prostate or ovarian cancer at MSKCC (personal communication with Dr. Slovin and Dr. 
Sabbatini). Autoimmune reactions may be controlled by cessation of the immunization and anti- 
inflammatory medications. The symptoms would be dependent on the organs involved. These 
may include but are not limited to diarrhea, dyspnea, numbness, weakness, vaginal bleeding, 
inflammation of the breast or pancreas and blood in the stool. This autoimmunity is considered 
unlikely however, since cytotoxic T-cells and antibodies against various antigens found on 
breast cancer cells have been recognized in the peripheral blood of patients with breast cancer in 
the absence of any evidence for autoimmunity. Some patients have also demonstrated baseline 
antibody titers against various antigens used in our prior trials. 
 
In MSKCC protocol 01-019 several episodes of transient asymptomatic elevation of the AST and 
ALT levels (grade 1-3) were observed. The finding was thought to be due to the GPI-0100 and we 
do not anticipate this toxicity on this trial using QS-21. 
 
The rare possibility of allergic reaction exists (ie wheezing, hives, fevers, chills, dyspnea). 
 
In prior immunization trials, moderate local discomfort at injection sites and flu-like symptoms 
were observed with QS-21 at the 200 mcg dose. The 100 mcg dose was found to be effective with 
minimal toxicity. We do anticipate significant toxicity with the higher dose of QS-21 (150 mcg). 
The only expected toxicity of KLH is local injection site reaction. 
 
Grade III or greater local or systemic toxicity will result in removal of the patient from the study. 
Grade II or greater toxicity of autoimmune etiology will result in removal from the study. Grade II 
or greater allergic toxicity, excluding a fever or flushing, will result in removal from the study. 
However, patients who develop a fever > 38 degrees Centigrade, thought to be due to the vaccine, 
will be monitored for 60 minutes in the Immunology Unit following the next vaccination. 
 
Any patient removed from the study due to toxicity will result in suspension of the study pending 
further investigation and discussion with the CBER IND review team.  In that case, if mutually 
agreed on, the trial may resume at the 10mcg sLea dose, if that was well tolerated. Patients will be 
counseled to avoid pregnancy. 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
 
 
Note: Grade 3 lymphopenia is acceptable since the eligibility criteria allows for a grade 3 baseline 
value. 
 
Adve rse events will be reported to the Institutional Review Board and to the Food and 
Drug Administration. Patients will be asked to complete a diary after each vaccine. (Appendix B) 
 
Criteria for Toxicity: Toxicity will be graded in accordance with the Common Toxicity Criteria 
Version 3.0 developed by the National Cancer Institute (NCI). (Refer to the Internet web site  
http://www.nci.nih.gov) 
 
 
12.0 
CRITERIA FOR THERAPEUTIC RESPONSE/OUTCOME ASSESSMENT 
 
The primary endpoints of this trial are toxicity and immunologic response. Patients will have 
baseline imaging studies to assess the extent of the disease. However, patients will not routinely 
undergo followup imaging studies unless felt to be appropriate by their physician as standard of 
care. 
 
12.1 
Long Te rm Followup 
 
Patients will be followed every 3 months if feasible following completion of the vaccine 
trial, until evidence of disease progression. Shorter time intervals between visits may be 
required, according to the standard of care for the patient‟s state of disease. 
 
12.2 
Evaluability 
 
Efforts shall be made to account for all patients entered into the study during the evaluation 
of results.  All patients will be considered evaluable for safety and immune response. 
 
12.3 
Clinical Response 
 
Patients will be monitored for anti-tumor effect, although this is not a primary endpoint. 
Patients who have evidence of disease will be evaluated at week 19 or at the discretion of 
their physician. Responses will be categorized using RECIST response criteria as follows: 
 
Complete Response (CR): 
Target Response Criteria: The disappearance of all known disease and no new sites or 
disease related symptoms confirmed at least 4 weeks after initial documentation. All sites 
must be assessed, including non-measurable sites, such as effusions, or markers 
Non-Target Response Criteria: Disappearance of all non-target lesions and normalization of 
tumor markers level confirmed at least 4 weeks after initial documentation. 
 
 
Partial Response (PR): 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
Target Response Criteria: At least a 30% decrease in the sum of the longest diameters of 
target lesions, taking as a reference the baseline sum of the longest diameters confirmed at 
least 4 weeks after initial documentation. PR is also recorded when all measurable disease 
has completely disappeared, but a non-measurable component (i.e., ascites) is still present 
but not progressing. 
 
Stable Disease (SD): 
Target Response Criteria: Neither sufficient shrinkage to qualify for partial response nor 
sufficient increase to qualify for progressive disease. 
Non-Target Response Criteria: Persistence of one or more non-target lesions and/or the 
maintenance of tumor marker level above the normal limits. 
 
 
Progressive Disease (PD): 
Target Response Criteria: At least a 20% increase in the sum of the longest diameters of 
target lesions, taking as reference the smallest sum of the longest diameters recorded since 
the treatment started; or the appearance of one or more new lesions; or the unequivocal 
progression of a non-target lesion. 
Non-Target Response Criteria: Appearance of one or more non-target lesions and/or 
unequivocal progression of existing non-target lesions. 
 
13.1 
CRITERIA FOR REMOVAL FROM STUDY 
 
Cessation of tre atment 
 
Therapy may be discontinued at any time due to severe unacceptable side effects, dose- 
limiting toxicities as defined, life-threatening events, patient‟s non-compliance with the 
defined treatment plan, the patient becomes pregnant or the patient‟s request to withdraw 
consent. 
 
With evidence of recurrent disease or progression of disease, the patient‟s primary 
physician will be notified immediately by the protocol investigators.  The decision 
regarding optimal therapy will be made by the patient and the primary physician. The 
patient may continue on study if the patient‟s primary physician feels it is acceptable and 
the only change in therapy is hormone therapy. 
 
The following dose limiting toxicities as evaluated during weeks 1-21 will result in 
cessation of treatment: 
• 
Grade 2 or greater autoimmune reactions 
• 
Grade 2 or greater allergic reactions (excluding isolated fever and/or flushing) 
• 
Grade 3 or greater toxicity of any kind (excluding grade 3 lymphopenia) 
 
If any of these toxicities occur, no additional patients will be treated except 
following mutual agreement after discussion with the CBER IND review team. 

- 17 - 
Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
14.0 
BIOSTATISTICS 
This is a pilot study designed to assess the safety and immunogenicity of immunization with sLea- 
KLH plus QS-21 in patients with high risk breast cancer.  If immune responses are observed at  
the 100 mcg level of QS-21 then QS-21 will be escalated to the 150mcg level. 
 
The immunization schedule comprises five injections administered during weeks 1,2,3,7 and 19. 
The primary endpoint of this trial is to evaluate immunological response to this single antigen and 
evaluate toxicity. Immunological response for a patient is defined as an antibody response of 
greater than 1:80 if baseline value was 0 and at least an increase of 8-fold if baseline value is 
greater than 0.  Initially, three patients will be accrued to the study at a dose of 10 mcg of sLea. If 
safe, then ten patients will be accrued to the study at the full 30mcg sLea  dose.  If 6 or more of 
these 10 patients have an immune response at any time point then an additional 10 patients may  
be accrued and receive sLea-KLH plus QS-21 (at 150mcg) during weeks 1, 7 and 19. This sample 
size is chosen to minimize the total number of patients who need to be immunized to identify a 
promising agent with a probability of an immunological response greater than 50% in a series of 
immunization trials. The error rates of missing a promising agent or selecting a nonpromising 
agent are set to be at most 10%.36 
 
At the end of the trial a qualitative assessment of the number of responses and frequency of toxicities 
will be made to determine which dose of QS-21 will be carried forward in future studies. 
 
Severe toxicities are not anticipated. Toxicities will be tabulated according to the NCI CTC 
version 3.0. Any grade 3 or greater toxicity (excluding grade 3 lymphopenia) or grade 2 or greater 
allergic/immunologic toxicity (except isolated fever and/or flushing) will result in removal of the 
patient from the study. Since the above listed toxicities are unexpected, any event of this sort will 
result in suspension of the study pending further investigation.   Descriptive statistics summarizing 
the antibody responses will be provided for each patient. The length of the study will be 
approximately 1-2 years. 
 
Circulating tumor cells (CTC)  will be evaluated in an exploratory manner to determine whether 
CTC can be detected in this patient population at baseline and if so whether there is evidence of a 
decline in those patients who have an immune response. 
 
Severe toxicity is not anticipated. Frequencies of all toxicities will be recorded. We plan to accrue 
approximately 1-2 patients per month and therefore accrual should be completed in 10 months. 
 
15.1 
RESEARCH PARTICIPANT REGISTRATION AND RANDOMIZATION 
PROCEDURES 
 
15.2 Research Participant Registration 
 
The following persons can obtain informed consent: 
 
Teresa Gilewski M.D. 
Maura Dickler M.D. 
Clifford Hudis M.D. 
Larry Norton M.D. 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
Confirm in the electronic medical record that the patient has received the Notice of Privacy 
Practice.  This must be obtained before the eligibility confirmation and obtaining of the research 
informed consent. 
 
Confirm eligibility as defined in the section entitled Criteria for Patient/Subject Eligibility. 
 
Obtain written informed consent, by following procedures defined in section entitled Informed 
Consent Procedures. 
 
All particpants must be registered through the Protocol Participant Registration (PPR) Office at 
Memorial Sloan-Kettering Cancer Center.  PPR is available Monday through Friday from 8:30am 
- 5:30pm at (646) 735-8000. The PPR fax numbers are (646) 735-0008 and  (646) 735-0003. 
Registrations can be phoned in or faxed.  The completed signature page of the informed consent 
form, the completed signature page of the Research Authorization and a completed Eligibility 
Checklist must be faxed to  PPR. 
 
During the registration process registering individuals will be required to answer specific 
eligibility questions and provide the following information: 
 
Registering Individual 
[Last, First Name] 
Notice of Privacy Status 
[Yes, No, N/A] 
Research Authorization 
[Date] 
MSKCC IRB Protocol# 
Attending of Record  (if applicable) 
[Last, First Name] 
Consenting Professional 
[Last, First Name] 
Informed Consent Date 
Participant's Full Name 
[Last, First Name] 
Participant MRN 
 
 
 
16.1 
DATA MANAGEMENT ISSUES 
 
A data profile in Appendix A will be completed for all patients. 
 
A Research Study Assistant (RSA) will be assigned to the study. The responsibilities of the RSA 
include project compliance, data collection, abstraction and entry, data reporting, regulatory 
monitoring, problem resolution and prioritization and coordination of activities of the protocol 
study team. 
 
The data collected for this study will be entered into a secure database at MKSCC. Data from this 
tria l will be entered in the Clinical Research Data Base (CRDB). Source documentation will be 
available to support the computerized patient record. 

- 19 - 
Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
16.2 
Quality Assurance 
 
Weekly registration reports will be generated to monitor patient accruals and completeness of 
registration data.  Routine data quality reports will be generated to assess missing data and 
inconsistencies.  Accrual rates and extent and accuracy of evaluations and follow-up will be 
monitored periodically throughout the study period and potential problems will be brought to the 
attention of the study team for discussion and action. 
 
Random-sample data quality and protocol compliance audits will be conducted by the study team, 
at a minimum of  two times per year, more frequently if indicated. 
 
16.3 
Data and Safety Monitoring 
 
The Data and Safety Monitoring (DSM) Plans at Memorial Sloan-Kettering Cancer Center were 
approved by the National Cancer Institute in September 2001.  The plans address the new policies 
set forth by the NCI in the document entitled “Policy of the National Cancer Institute for Data 
and Safety Monitoring of Clinical Trials” which can be found at:  
http://cancertrials.nci.nih.gov/researchers/dsm/index.html.    The DSM Plans at MSKCC were 
established and are monitored by the Office of Clinical Research.  The MSKCC Data and Safety 
Monitoring Plans can be found on the MSKCC Intranet at: 
http://mskweb2.msk cc.org/irb/index.htm 
 
There are several different mechanisms by which clinical trials are monitored for data, safety and 
quality.  There are institutional processes in place for quality assurance (e.g., protocol monitoring, 
compliance and data verification audits, therapeutic response, and staff education on clinical 
research QA) and departmental procedures for quality control, plus there are two institutional 
committees that are responsible for monitoring the activities of our clinical trials programs.  The 
committees: Data and Safety Monitoring Committee (DSMC) for Phase I and II clinical trials, and 
the Data and Safety Monitoring Board (DSMB) for Phase III clinical trials, report to the Center‟s 
Research Council and Institutional Review Board. 
 
During the protocol development and review process, each protocol will be assessed for its level 
of risk and degree of monitoring required. Every type of protocol (e.g. NIH sponsored, in-house 
sponsored, industrial sponsored, NCI cooperative group, etc.) will be addressed and the 
monitoring procedures will be established at the time of protocol activation. 

- 20 - 
Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
17.1 
PROTECTION OF HUMAN SUBJECTS 
 
The study will maintain the privacy of the participant as noted in Section 7.1 The patient will be 
informed that the study is voluntary and the participant may withdraw at any time. Alternative 
options for the patient are not to participate in this study and receive no therapy or to receive 
other treatments that may include hormone therapy, chemotherapy, radiation therapy, targeted 
therapy, surgery or other investigational therapy. The patient will be informed that there is no 
known clinical benefit to the immunization under evaluation in this study. Potential benefits 
include generation of an anti-tumor response by stimulation of the immune system. 
 
The patient is responsible for all costs excluding the sLea-KLH plus QS-21 immunization, the 
evaluation of immune response and the assessment of circulating tumor cells. Potential toxicities 
are outlined in Sections 11 and 13. 
 
17.2 
Privacy 
 
It is the responsibility of the Research Staff to ensure that protocol subjects received the Center‟s 
Notice of Privacy Practices.  If the subject has not received one, MSK personnel must provide a 
Notice of Privacy Practices and obtain acknowledgment before the subject participates in the 
study. 
 
MSKCC‟s Privacy Office may allow the use and disclosure of protected health information 
pursuant to a completed and signed Research Authorization form.  The use and disclosure of 
protected health information will be limited to the individuals described in the Research 
Authorization form.  A Research Authorization form must be completed by the Principal 
Investigator and approved by the IRB and Privacy Board. 
 
17.3 Serious Adverse Event (SAE) Reporting 
 
Any SAE must be reported to the IRB as soon as possible but no later than 5 calendar days. The 
IRB requires a Clinical Research Database (CRDB) AE report to be delivered to the Institutional 
SAE Manager (307 East 63rd  Street, 1st  Floor) containing the following information: 
Fields populated from the CRDB: 
• 
Subject‟s name 
• 
Medical record number 
• 
Disease/histology (if applicable) 
• 
Protocol number 
Data needing to be entered: 
• 
The date the adverse event occurred 
• 
The adverse event 
• 
Relationship of the adverse event to the treatment (drug, device, or intervention) 
• 
If the AE was expected 
• 
The severity of the AE 
• 
The intervention 
• 
Detailed text that includes the following information: 
o A explanation of how the AE was handled 
o A description of the subject's condition 
o Indication if the subject remains on the study 

- 21 - 
Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
o If an amendment will need to be made to the protocol and/or consent form 
The PI‟s signature and the date it was signed are required on the completed report. 
For IND/IDE protocols : 
The CRDB AE report should be completed as above and the FDA assigned IND/IDE number 
written at the top of the report. The report will be forwarded to the FDA by the Institutional SAE 
Manager through the IND Office. 
 
17.2.1  IND Safety Reports 
This protocol is sponsored under an IND. Therefore, any serious adverse events will be reported 
to the Food and Drug Administration through the IND office of Memorial Sloan-Kettering 
Cancer Center, as set forth in FDA regulation 312.32. 
 
17.3 
Reporting of Othe r Adve rse Eve nts 
 
Une xpected Toxicities -   an event that is not  identified in terms of  its nature, 
severity or frequency of occurrence in the expected toxicities. 
 
The MSKCC IRB must be notified in writing within 5 working days of the event. 
 
Non-tre atme nt Re late d Toxicities 
 
If a toxicity is not a serious adverse event and unrelated to the protocol treatment, 
this must clearly be documented on the flow sheets which are submitted to the 
MSKCC clinical trials office. This does not in any way obviate the need for 
reporting the toxicities described above. 
 
 
 
18.1 
INFORMED CONSENT PROCEDURES 
 
A consenting professional will explain the protocol to the patient. In addition to the 
verbal explanation, patients will receive the informed consent document to read.  If the 
patient agrees to participate in the trial, the patient will sign three copies of the informed 
consent. One copy will be placed in the patient‟s medical record, one copy will be 
returned to the patient.  One copy will be placed in the research file. 
 
 
18.2 
Research Authorization 
 
Procedures for obtaining Research Authorization: Before any protocol-specific procedures are 
carried out, investigators and/or designated staff will fully explain the details of the protocol, 
study procedures, and the aspects of patient privacy concerning research specific information.  In 
addition to signing the IRB Informed Consent, all patients must sign the Research Authorization 
component of the informed consent form.  The Research Authorization requires a separate set of 

- 22 - 
Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
signatures from the patient.  The original signed documents will become part of the patient‟s 
medical record, and each patient will receive a copy of the signed documents. 

- 23 - 
Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
19.1 
REFERENCE(S) 
 
1 Cancer Control Continuum. [Online]. Available: 
http ://ca ncerco ntro l.c a ncer. go v/od/co ntinuum. html [accessed April 19, 2005]. 
 
2 Estimated U.S. Cancer Prevalence. [Online]. Available: 
http ://ca ncerco ntro l.c a ncer. go v/ocs/p re va le nce/pre va le nce. html [accessed May 31, 2005] 
 
3 Livingston PO, Natoli EJ, Jones Calves M, Stockert E, Oettgen HF, O ld LJ. Vaccines 
containing purified GM2 ganglioside elicit GM2 antibodies in melanoma patients. Proc Natl 
Acad Sci USA 84:2911-2915, 1987. 
 
4 Jones PC, Sze LL, Liu PY, Morton DL, Irie RF. Prolonged survival for melanoma patients 
with elevated IgM antibody to oncofetal antigen. JNCI 66:249-254, 1981. 
 
5 Zhang S, Cordon-Cardo C, Zhang HS, Reuter VE, Adluri S, Hamilton WB, Lloyd KO, 
Livingston PO. Selection of tumor antigens as targets for immune attack using 
immunohistochemistry: I. Focus on gangliosides. Int J Cancer 73:42-49, 1997. 
 
6 Zhang S, Zhang HS, Cordon-Cardo C, Reuter VE, Singhal AK, Lloyd KO, Livingston PO. 
Selection of tumor antigens as targets for immune attack using immunohistochemistry: II. 
Blood group-related antigens. Int J Cancer 73:50-56, 1997. 
 
7 Zhang S, Zhang HS, Cordon-Cardo C, Ragupathi G, Livingston PO. Selection of tumor 
antigens as targets for immune attack using immunohistochemistry: Protein antigens. Clin 
Cancer Res 4:2669-2676, 1998. 
 
8 Henningsson CM, Selvaraj S, MacLean GD, et al. T-cell recognition of a tumor-associated 
glycoprotein and its synthetic carbohydrate epitopes: stimulation of anti-cancer T-cell 
immunity in vivo. Cancer Immunol Immunother 25:231, 1987. 
 
9 Romball GC, Weigle WO. T-cell competence to heterologous and homologous 
thyroglobulins during the induction of experimental autoimmune thyroiditis. Eur J Immunol 
14:887, 1984. 
 
 
10 Longenecker BM, Willans DJ, MacLean GD, et al. Monoclonal antibodies and synthetic 
tumor-associated glycoconjugates in the study of the expression of Thomsen-Friedenereich- 
like and Tn- like antigens on human cancers. JNCI 78:489-496, 1987. 
 
11 Ragupathi G, Koganty RR, Qiu D, Lloyd KO, Livingston PO. A novel and efficient 
method for synthetic carbohydrate conjugate vaccine preparation: synthesis of sialyl Tn-KLH 
conjugate using a 4-(4-N-maleimidomethyl) cyclohexane-1-carboyxl hydrazide (MMCCH) 
linker arm. Glycoconjugate Journal 15:217-221, 1998. 

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Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
12 Kensil CR, Patel U, Lennick M, and Marciani D, Separation and characterization of 
saponins with adjuvant activity from Quillaja saponaria Molina cortex. J. Immunol 146:431- 
437, 1991. 
 
13 Zhang S, Graeber LA, Helling F, Ragupathi G, Adluri S, Lloyd KO, Livingston PO. 
Augmenting the immunogenicity of synthetic MUC1 peptide vaccines in mice. Cancer Res 
56:3315-3319, 1996. 
 
14 Livingston PO, Ragupathi G. Carbohydrate vaccines that induce antibodies against cancer. 
2. Previous experience and future plans. Cancer Immunol Immunother 45;10-19, 1997. 
 
15 Livingston PO, Adluri S, Helling F, Yao TJ, Kensil CR, Newman MJ and Marciani D. 
Phase I trial of immunological adjuvant QS-21 with a GM2 ganglioside-keyhole limpet 
hemocyanin conjugate vaccine in patients with malignant melanoma. Vaccines 12;1275-1280, 
1994. 
 
16 Marciani DJ, Kensil CR, Beltz GA, Hung CH, Cronier J, Aubert A. Genetically 
engineered subunit vaccine against feline leukaemia virus: protective immune response in 
cats. Vaccine 9:89-96, 1991. 
 
17 Gilewski T, Adluri S, Ragupathi G, et al: Vaccination of high risk breast cancer patients 
with mucin-1 (MUC-1) keyhole limpet hemocyanin conjugate plus QS-21.  Clin Cancer 
Research 6:1693-1701, 2000. 
 
18 Gilewski TA, Ragupathi G, Bhuta S, et al: Immunization of metastatic breast cancer 
patients with a fully synthetic globo H conjugate: a phase I trial. Proc Natl Acad Science 
98:3270-3275, 2001. 
 
19 Gilewski T, Norton L, Houghton A, et al: Vaccination of high risk breast cancer patients 
(pts) lacking identifiable disease with GM2-kehyhole limpet hemocyanin (KLH) conjugate 
plus QS-21 (abstr) Proc Am Soc Clin Onc 18;316a, 1999. 
 
20 Dickler M, Gilewski T, Ragupathi G, et al: Vaccination of breast cancer patients (pts) with 
no evidence of disease (NED) with sialyl Tn cluster (sTn[c])-keyhole limpet hemocyanin 
(KLH) conjugate plus adjuvant QS-21: preliminary results. (abstr) Proc Am Soc Clin Onc 
16:439a, 1997. 
 
21 Armstrong JL, Ragupathi G, Powell S, et al. Preliminary data of vaccination of high risk 
breast cancer (BC) patients (pts) with a heptavalent antigen: keyhole limpet hemocyanin 
(KLH) conjugate plus the immunologic adjuvant QS-21. Proc Am Soc Clin Onc 22:675, 
2003. 
 
22 Marciani D, Press JB, Reynolds RC, Pathak AK, Pathak V, Gundy LE, Farmer JT, 
Koratich MS, May RD. Development of semisynthetic triterpenoid saponin derivatives with 
immune stimulating activity. Vaccine 18:3141-51, 2000. 

- 25 - 
Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
23 Slovin S, Ragupathi G, Fernandez C, Randall E, Diani M, Biechy C, Verbel D, Arauz G, 
Spassova M, Bornmann W, Livingston PO, Danishefsky S, Scher H. A bivalent vaccine 
containing glycosylated MUC-2-KLH and globo H-KLH conjugates using a new semi- 
synthetic saponin immunological adjuvant, GPI-0100 in biochemically relapsed prostate 
cancer (PC). Proc Am Soc Clin Oncol 11;196a, 2002 (abstr). 
 
24 Steplewska-Mazur K, Gabriel A, Zajecki W, Wylezov, Gluck M. Breast cancer 
progression and expression of blood group-related tumor-associated antigens. Hybridoma 
10:129-133, 2000. 
 
24a Ragupathi G, Liu NX, Musselli C, Powell S, Lloyd K and Livingston PO. Antibodies 
against tumor cell glycolipids and proteins, but not mucins, mediate complement-dependent 
cytotoxicity. J Immunol 174 (9):5706-5712, 2005. 
 
25 Austrup F, Uciechowski P, Eder C, Bockmann B, Suchy B, Driesel G, Jackel S, Kusiak I, 
Grill HJ and Giesing M.. Prognostic value of genomic alterations in minimal residual cancer 
cells purified from the blood of breast cancer patients. Br J Cancer 83(12):1664-1673, 2000. 
 
26 Beitsch PD and Clifford E. Detection of carcinoma cells in the blood of breast cancer 
patients. Am J Surg 180(6):446-448, 2000. 
 
27 Carey RW, Taft PD, Bennett JM and Kaufman S. Carcinocythemia (carcinoma cell 
leukemia). An acute leukemia- like picture due to metastatic carcinoma cells. Am J Med 
60(2):278-278, 1976. 
 
28 Cristofanilli M, Budd GT, Ellis MJ, Stopeck A, Matera J, Miller MC, Reuben JM, Doyle 
GV, Allard WJ, Terstappen LW, Hayes DF. Circulating tumor cells, disease progression and 
survival in metastatic breast cancer. N Engl J Med 351(8):781-791, 2004. 
 
29 Fehm T, Sagalowsky A, Clifford E, Beitsch P, Saboorian H, Euhus D, Meng S, Morrison 
L, Tucker T, Lane N, Ghadimi BM, Heselmeyer-Haddad K, Ried T, Rao C, and Uhr J. 
Cytogenetic evidence that circulating epithelial cells in patients with carcinoma are 
malignant. Clin Cancer Res 8(7):2073-2084, 2002. 
 
30 Gaforio JJ, Serrano MJ, Sanchez-Rovira P, Sirvent A, Delgado-Rodriguez M, Campos M, 
de la Torre N, Algarra I, Duenas R, and Lozano A. Detection of breast cancer cells in the 
peripheral blood is positively correlated with estrogen-receptor status and predicts for poor 
prognosis. Int J Cancer 107(6):984-990, 2003. 
 
31 Gallivan MV and Lokich JJ. Carcinocythemia (carcinoma cell leukemia). Report of two 
cases with English literature review. Cancer 53(5):1100-1102, 1984. 
 
32 Myerowitz Rl, Edwards PA and Sartiano GP. Carcinocythemia (carcinoma cell leukemia) 
due to metastatic carcinoma of the breast: report of a case. Cancer 40(6):3107-3111 1977. 

- 26 - 
Memorial Sloan Kettering Cancer Center 
IRB Number: 06-156 A(4) 
Approval date: 24-Sep-2018 
 
 
 
 
33 Terstappen LW, Rao C, Gross S and Weiss AJ. Peripheral blood tumor cell load reflects 
the clinical activity of the disease in patients with carcinoma of the breast. Int J Oncol 
17(3):573-578, 2000. 
 
34 Ashworth TR. A case of cancer in which cells similar to those in the tumors were seen in 
the blood after death. Aust Med J 14:146-147, 1869. 
 
35 Krag DN, Ashikaga T, Moss TJ, et al. Breast cancer cells in the blood: a pilot study. 
Breast J 5:354-358, 1999. 
 
36 Yao TJ, Begg CB, Livingston PO. Optimal sample size for a series of pilot trials of new 
agents. Biometrics 55:992-1001, 1996. 
 
20.0 
APPENDICES 
 
APPENDIX A: PATIENT CALENDAR 
APPENDIX B: PATIENT DIARY