Phase 1 Safety Study of a Carbohydrate Mimotope Based Vaccine With MONTANIDE ISA 51 VG Adjuvant
Phase: PHASE1
Conditions: Stage IV Breast Cancer
Sponsor: University of Arkansas

1 
 
 
 
 
 
 
 
“Vaccination of High Risk Breast Cancer Patients” 
 
Phase 1 Safety Study 
 
Of 
 
A Carbohydrate Mimotope Based Vaccine with MONTANIDETM ISA 51 
VG Adjuvant 
 
 
 
 
 
 
 
 
 
 
Laura Hutchins, M.D., Principal Investigator 
Thomas Kieber-Emmons, Ph.D., Subinvestigator 
Issam Makhoul, M.D., Subinvestigator 
Rangaswamy Govindarajan, M.D., Subinvestigator 
Konstantinos Arnaoutakis, M.D., Subinvestigator 
Peter Emanuel, M.D., Subinvestigator 
Liudmila Schafer, M.D., Subinvestigator 
Ronald Haggett, A.P.N. 
Donna Dunn, A.P.N. 
Eric Siegel, M.S., Biostatistician 
Joseph M. Beck II, M.D., Medical Monitor 
Jeannette Lee, M.D. Biostatistician 
 
 
 
 
Version 19.120 
3/28/11/1/2013 

 
Version: 19.120 
Date: 11/13/28/2013 
      
2
 
TABLE OF CONTENTS 
 
 
 
 
 
 
 
 
 
 
PAGES 
1.  Abbreviations……………………………………………………………..  
4-5 
2. Protocol Summary………………………………………………………… 
6 
3. Background……………………………………………………………….. . 
6-9 
4. Trial Objectives………………………………………………................... 
9-10 
a. Primary Objective…………………………………………………….. 
9 
b. Secondary Objectives………………………………………………..  
9-10 
c. Exploratory Objective………………………………………………… 
10 
5. Patient Population………………………………………………………… 
10-11 
6. Investigational new Drug – P10s-PADRE- MONTANIDETM ISA 51 VG 
11-12 
a. General Description…………………………………………………... 
11 
b. Vaccine Manufacturing and Formulation.………………………….. 
11-12 
c. Vaccine Preparation………………………………………………….. 
12 
d. DTH Preparation……………………………………………………… 
12 
e. Dosing Administration………………………………………………  
12 
f.    Label Information ..…………………………………………………..  
12 
g. Agent Ordering ………………………………………………………  
12 
h. Agent Accountability ……………………………………………….  
12 
7. Treatment Plan……………………………………………………………. 
13-22 
a. On-Study Evaluation…………………………………………………. 
13 
b. Dose Assignment ……………………………………………………. 
13 
c. Specimen Handling.………………………………………………….  
13 
d. Visit Breakdown ….…………………………………………………..  
13-20 
e. Prohibited Medications ……………………………………………… 
20 
f. Dose Limiting Criteria………………………………………………… 
20 
g. Dose Assignment ……………………………………………………  
20-22 
8. Risks and Toxicities to be Monitored…………………………………… 
22-25 
a. Potential Toxicities, Risks and Precautions……………………….. 
22-25 
b. Benefits………………………………………………………………… 
25 
c. Injury …………………………………………………………………… 
25-26 
9. Specimen Handling……………………………………………………….  
26 
10. Study Calendar …………………………………………………………… 
27-28 
11. Criteria for Evaluation……………………………………………………... 
29 
12. Statistical Considerations…………………………………………………. 
30-32 
a. Sample Size, Duration and General Considerations………………. 
30 
b. Data Analysis Plan…………………………………………………….. 
30-31 
c. Missing, Unused and Spurious Data………………………………… 
32 
13. Registration Guidelines……………………………………………………. 
32 
14. Data Submission Schedule……………………………………………….. 
32 
15. Ethical and Regulatory Considerations………………………………….. 
32-33 
a. Recruitment and Informed Consent…………………………………. 
32 
b. Institutional Review……………………………………………………. 
33 
c. Investigational Agent Accountability………………………………… 
33 

 
Version: 19.120 
Date: 11/13/28/2013 
      
3
16. Adverse Events (AEs)…………………………………………………….. 
33-34 
a. Adverse Events……………………………………………………….. 
33-34 
b. Serious Adverse Event (SAE)……………………………………….. 
34 
17. Monitoring………………………………………………………………….. 
34-36 
a. Medical Monitor……………………………………………………….. 
34 
b. Data Monitor…………………………………………………………..  
34-36 
1. Pre-Investigation Site Visit…………………….. 
35 
2. Periodic Site Visits……………………………… 
35  
3. Investigational Products………………………..  
35 
4. Monitoring Report………………………………. 
35 
5. Research Subject Safety and Stopping Rules . 35-36 
6. Audits…………………………………………….. 
36 
18. Bibliography……………………………………………………………….. 
37-39 
 APPENDIX 
 
APPENDIX A – NCI Common Terminology Criteria for Adverse Events (CTCAE) Version 
4.0 
 
USEFUL FIGURES AND TABLES 
Toxicity Decision Rules: Tables 1- 3……………………………………… 
21 
Potential Toxicities, Risks and Precautions……………………………… 
22-25 
Study Calendar……………………………………………………………… 
27-28 
 

 
Version: 19.120 
Date: 11/13/28/2013 
      
4
1. Abbreviations 
 
ACLS – advanced cardiac life support 
AE – adverse event  
AJCC – American Joint Commission on Cancer 
ANA – anti-nuclear antibody 
AST – aspartate aminotransferase test 
BUN – blood urea nitrogen 
CBC – complete blood count 
CDC – complement dependent cytotoxicity 
CMPs – carbohydrate mimetic peptides 
Co-PI – co-principal investigator/co-investigator 
CCTO- Cancer Clinical Trials Office 
CRA – Clinical Research Associate 
CRF – case report form 
CTC – circulating tumor cell 
CTCAE – Common Terminology Criteria for Adverse Events 
CTEP – Cancer Therapy Evaluation Program 
CTL – cytotoxic T lymphocyte 
dl – deciliter 
DLT – dose limiting toxicity 
DOD – Department of Defense 
DTH – delayed type hypersensitivity 
ELISA – enzyme-linked immunosorbent assay 
FACS – fluorescence activated cell sorting 
FDA – food and drug administration 
GCP – good clinical practice 
GGT – gamma-glutamyl transferase test 
GMP – good manufacturing practice 
id – intradermally 
IgG – immunoglobulin g 
IgM – immunoglobulin m 
IND - investigational new drug 

 
Version: 19.120 
Date: 11/13/28/2013 
      
5
IRB - institutional review board 
IUL – institutional upper limit 
kD – kiloDalton 
KLH – keyhole limpet hemocyanin 
LDH – lactate dehydrogenase 
LeY – Lewis Y antigen 
MFI – mean fluorescence intensity 
mg – milligram 
mL – milliliter 
mm – millimeter 
MTD – maximum tolerated dose 
µg – microgram 
NCI – National Cancer Institute 
NK – natural killer 
PBS – phosphate buffered saline 
PI – principal investigator 
PRMC – protocol review and monitoring committee 
PT – prothrombin time 
PTT – partial thromboplastin time 
RARE™ - RosetteSep-Applied imaging Rare Event (™StemCell Technologies, Vancouver) 
RSC – UAMS Research Support Center 
SAE – serious adverse event 
SC - subcutaneous 
SGOT – serum glutamic-oxaloacetic transaminase 
SLE – systemic lupus erythematosus 
SOP – standard operating procedure 
STn antigen – sialosyl Tn antigen 
TACAs - tumor associated carbohydrate antigens 
TSH – thyroid stimulating hormone 
UAMS - University of Arkansas for Medical Sciences 
WBC – white blood cell 
 
 

 
Version: 19.120 
Date: 11/13/28/2013 
      
6
2. Protocol Summary 
 
Primary Objective – Safety:  Determine the safety and tolerability of a peptide mimotope-based 
vaccine upon immunization of breast cancer subjects. 
Secondary Objectives – Immune Response: 
1) 
Determine whether immunization with the vaccine generates a humoral response 
against Tumor Associated Carbohydrate Antigens.  
2) 
Determine the delayed-type hypersensitivity (DTH) response to the immunizing 
mimotope 
3) 
Determine the effect of a late booster immunization on the humoral response against 
TACAs 
Exploratory Objective: Determine the effect of immunizing mimotope on circulating tumor cells 
(CTCs) 
 
Study Population:  We plan to consent approximately 24 subjects with hope of reaching our 
enrollment goal of 18 women, ages 18 and older from the breast cancer clinics (Medical 
Oncology and Ladies Oncology Clinics) at the Winthrop P. Rockefeller Cancer Institute at the 
University of Arkansas for Medical Sciences (UAMS) campus. 
Inclusion Criteria:  Females with histologically or cytologically confirmed stage IV breast cancer 
(newly diagnosed metastatic or relapsed after primary or adjunctive therapy, which has not 
required a treatment change for 2 months) will be invited to participate. 
Exclusion Criteria:  Women who are pregnant, breast-feeding, have autoimmune disease or 
are immunosuppressed or receiving systemic corticosteroids will be excluded from the study. 
Investigational product: P10s-PADRE administered with the MONTANIDETM ISA 51 VG 
Study Design: After signing IRB approved consent, cohorts of 3-6 stage IV breast cancer 
subjects will be enrolled into the study. The vaccine doses will be prepared and dispensed by the 
UAMS Pharmacy following the manufacturer’s instructions.  Subjects will receive up to 2.0 mL 
subcutaneous (SC) injections of the vaccine on 5 separate occasions during Weeks 1, 2, 3, 7, 
and 19. The first cohort will begin with the 300 μg dose, and then the subsequent cohorts will 
escalate to 500 µg or de-escalate to 100 µg as determined by the toxicity criteria shown in Table 
1 of Section 7. The immunization at week 19 is considered a booster immunization. The vaccine 
will be administered at rotating sites on the limbs or abdomen by nurses in the Infusion Center at 
the Cancer Institute, using a dose volume of up to 2.0 mL per injection.  Based on a series of 
criteria measuring tolerance and immune response in the first cohort on Week 9 (Section 7, 
Table 1), the P10s-PADRE dose will either be increased to 500 µg/mL or decreased to 100 
µg/mL for the second cohort of subjects. The study will last for approximately 12 – 24 months.  
3. Background 
 
Anticipated anti-cancer impact of carbohydrate-targeted vaccines: The potential impact of 
vaccines that induce responses to tumor-associated carbohydrate antigens (TACAs) is 
demonstrated by clinical trials where patient survival significantly correlates with carbohydrate-
reactive IgM levels (2).  Such results suggest that TACA-targeting vaccines might have a 
beneficial effect on the course of malignant disease.  TACA-induced responses could augment 
naturally occurring carbohydrate-reactive IgM antibodies that trigger apoptosis of tumor cells (3).  
TACAs are attractive targets because the majority of cell-surface proteins and lipids are 
glycosylated, and the glycosyl moiety is fundamental to the biological functions of these 
molecules in cancer cells (4,5). A unique advantage in targeting TACAs is that multiple proteins 
and lipids on the cancer cell can be modified with the same carbohydrate structure.  Thus, 

 
Version: 19.120 
Date: 11/13/28/2013 
      
7
targeting the carbohydrate antigen broadens the spectrum of antigens recognized by the 
immune response, thereby lowering the risk of developing resistant tumors due to the loss of any 
one antigen (6). In addition, antibodies that recognize glycolipids are more apt to mediate 
complement-dependent cytotoxicity (CDC) and may, therefore, be more cytotoxic to tumor cells 
than antibodies that recognize protein antigens (7).  Furthermore, preclinical studies support the 
hypothesis that vaccine-induced responses against TACAs might have their greatest impact in 
the adjuvant setting, as such responses inhibit tumor outgrowth in metastatic models (8,9). 
 
Approaches to augment immune responses to TACA: A variety of approaches are being 
taken to generate responses to TACAs.  Because TACAs are T-cell-independent antigens and 
self-antigens, conjugation to immunologic carrier proteins is perceived to be essential to recruit 
T-cell help in antibody generation. Conjugation does not, however, assure an increase in 
immunogenicity because conjugation strategies do not uniformly enhance carbohydrate 
immunogenicity (10,11).  Furthermore, even with conjugation, the lack of induction of cellular 
immune responses that would amplify TACA-reactive humoral responses necessitates constant 
boosting with vaccine.  Representative examples of carbohydrate-based conjugate vaccines in 
clinical development include those directed toward gangliosides (12-14), polysialic acid (15), 
Globo-H (16), Lewis Y (LeY) (1), and the sialosyl-TN (STn) antigen (17). 
 
An approach predicted to facilitate cellular responses exploits the molecular mimicry of TACAs 
by protein surrogates, as they are T-cell-dependent antigens.  Clinical characterizations of anti-
idiotypic antibodies that mimic the GD3 ganglioside antigen (18) and GD2 (19) have been 
described.  Carbohydrate mimetic peptides (CMPs) are alternatives to anti-idiotypic antibodies.  
The characterization of CMPs is at present limited to preclinical studies.  CMPs that induce 
immune responses cross-reactive with TACA are also referred to as peptide mimotopes. Peptide 
mimotopes have been described for the GD2 (20-22), GD3 (23), sialylated Lewis a/x (24) and 
Lewis Y (LeY) antigens (20, 25).  Importantly, in preclinical prophylactic and therapeutic 
vaccination studies, peptide mimotopes were efficacious in eliciting immune responses that 
reduced tumor burden and inhibited metastatic outgrowth (8, 25, 26).  Thus, peptide mimotopes 
of TACAs represent a new and very promising tool to overcome T-cell independence and to 
increase the efficiency of the immune response to glycan antigens.  
 
Target carbohydrate antigens expressed on breast cancer cells: Tumors expressing high 
levels of certain types of TACAs exhibit greater metastasis than those expressing low levels of 
these antigens, and this negatively impacts prognosis (27-29).  In breast cancer, the LeY, STn, 
and KH-1 antigens, as well as selected gangliosides, glycosphingolipids and Globo-H 
carbohydrate antigens, are considered prime vaccine candidates because of their tissue 
distribution (30, 31).  In particular, LeY has long been recognized as a potential target for 
immunotherapy because it is expressed in 70–90% of tumors of epithelial origin (32).  The 
abundant gangliosides include GM3, GM2, GM1, and GD2, GD3 and GT3 (33). Antibodies to 
TACAs mediate a variety of effector functions and might lend to cross-presentation of tumor 
antigens to stimulate anti-tumor cellular responses. At present, LeY-conjugate vaccines appear 
to have only a limited ability to induce anti-LeY immune responses in humans (1).  Our in vitro 
studies demonstrate that peptide mimotopes of LeY and gangliosides induce serum antibodies in 
mice that recognize the appropriate carbohydrate antigens on human or murine breast cancer 
cell lines (25, 34).  Our in vivo studies demonstrate that the peptide mimotopes induce sustained 
immunity to these antigens (8, 25, 26). Collectively, these data provide the experimental 
foundation for evaluating peptide mimotopes as potential cancer vaccines in subjects with breast 
cancer.  
 

 
Version: 19.120 
Date: 11/13/28/2013 
      
8
Preclinical studies supporting the P10s-PADRE vaccine as a viable candidate for 
preventing breast cancer recurrence: The desired effect of a cancer vaccine is to modify the 
clinical outcome of the patient population of interest. Genetic studies have resurrected the 
concept that the adaptive and innate immune systems play roles in tumor surveillance. Cellular 
immunity, in which cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells are main effector 
cells, plays an important role in the antitumor defense mechanism. Tumors over express TACAs, 
which are reactive with B cells, but the use of TACAs as immunogens is restricted by a lack of B 
cell cross-talk with T cells. Consequently, this inherent limitation of plain polysaccharide vaccines 
includes limited duration of immunity, the potential for hyporesponsiveness with repeated 
vaccinations, and ineffectiveness in stimulating a cellular response. To circumvent this drawback 
we have developed CMPs with overlapping B- and T-cell epitopes to link TACAs’ reactive 
humoral responses with anti-tumor cellular responses. Among the CMPs we have developed are 
a series that contain the amino acids Trp-Arg-Tyr as a centralized motif.  CMPs with this motif 
display an ability to induce antibodies cross-reactive with tumor cells, induce cellular responses 
to tumor cells and induce or activate NK cells with anti-tumor activity.  
 
P10s-PADRE immunization in prophylactic and therapeutic models of murine cancer suggest 
that the vaccine promotes tumor growth inhibition. The safety and efficacy of P10s-PADRE in 
humans remains to be determined.  In many clinical trials, study subjects receive peptide 
vaccines in combination with a certain adjuvant, such as incomplete Freunds Adjuvant, for the 
purpose of enhancing immune responses against cancer. Water-in-oil or water-in-oil-in-water 
emulsions such as MONTANIDE™ ISA 51 VG are very well characterized adjuvants and used 
for more than 50 years in pre-clinical studies and in human clinical trials.  It is known that these 
types of adjuvant can induce strong Th2-type of antibody responses because they prolong the 
lifetime of antigens in vivo, enhance phagocytoses of antigens by professional Antigen 
Presenting Cells (APC), and induce transient expansion of CD4 positive Th cells which might 
also expand Th1 responses. 
 
In our studies in mice an adjuvant was added with the P10s-PADRE vaccine; as administered 
with Stimulon™QS-21 (Antigenics Inc.) and in limited instance MONTANIDE™ ISA 51 VG.  In 
this proposed Phase I study of P10s-PADRE, study subjects will receive P10s-PADRE dissolved 
in physiological saline admixed with MONTANIDE™ ISA 51 VG.  We have chosen to use 
MONTANIDE™ ISA 51 VG over QS-21 for two reasons.  Antigenics, Inc., the provider of QS-21, 
imposed new restrictions upon the further supply of QS-21.  These new restrictions contravene 
the stated mission of the University of Arkansas for Medical Sciences.  Therefore, QS-21 is no 
longer available for use. Second, even if no serious adverse effects in an extensive non-clinical 
safety study as performed in our GLP study with QS-21was observed, it cannot be guaranteed 
that the vaccine/adjuvant formulation presents no risks to vaccinees and unexpected events can 
occur.  Unpredictability of adjuvant effects in humans results from a complex interplay between 
such factors as route of administration, antigen dose, and nature of the antigen. For this reason 
a final safety evaluation can only be conducted on the basis of a clinical trial. MONTANIDE™ 
ISA 51 VG has been used in Phase I and II clinical trials for vaccines against malaria, HIV, and 
importantly various cancers (see Investigational brochure for MONTANIDE™ ISA 51 VG.  
MONTANIDE™ ISA 51 VG, has been tested in AIDS and cancer vaccine trials which represent 
more than 10,000 patients and around 100,000 injections. A survey of ongoing clinical trials 
listed in ClinicalTrials.gov revealed 36 trials currently accruing patients that are using the olive-
derived MONTANIDE™ ISA 51 VG.  The formulation is generally well tolerated and induces 
transient local reactions. Some transient general reactions such as flu-like symptoms can also be 
noticed. The results suggest that numerous repeated vaccine doses with MONTANIDE™ ISA 51 
VG can be safely administered.  

 
Version: 19.120 
Date: 11/13/28/2013 
      
9
Dose Justification: Immunologic testing in mice indicates that there is no difference in antibody 
responses to P10s in mice between the 300 µg and 500 µg dose (GLP Report: “Determination of 
the Safety and Tolerability of Immunization with LeY Peptide Mimotope Vaccine in Mice”). Dose 
escalation is meant to test the safety of the preparation in terms of its non-immunological toxicity. 
In this case the assumption is that the putative adverse effects increase monotonically with the 
dose, and exploring the higher-than-necessary doses ensures safety of the lower ones. As for 
the immunological effects, they are characteristically non-monotonic with an optimal effect in a 
specific dose range, and weaker effects and even induction of tolerance for doses much higher 
or lower than the optimal one. Thus, the classical dose escalation trial may or may not detect the 
immunological effects, including adverse autoimmune phenomena. A dose-escalation algorithm 
better suited for determining the immunological impact should include both increasing and 
decreasing arms, and should be terminated when doses beyond the one that leads to optimal 
immunological effect are reached, provided they are tolerated otherwise. When increasing 
immunological effect is detected in one of the arms, the other one may be discontinued, if not 
necessary for the toxicology study.  
4. Trial Objectives 
 
a. 
Primary Objective – Safety: The safety and tolerability of the P10s-PADRE/ 
MONTANIDETM ISA 51 VG vaccine will be determined by toxicity assessments throughout the 
duration of the study. Subjects will be evaluated for toxicity using the National Cancer Institute 
(NCI) Common Terminology Criteria for Adverse Events (CTCAE) Version 4.0. A toxicity of 
Grade 3 or higher will be considered a dose limiting toxicity (DLT).   
 
b.  Secondary Objectives – Immune Response: 
1) The ability of the P10s-PADRE/ MONTANIDETM ISA 51 VG vaccine to generate 
a humoral response against TACAs will be determined in the investigator’s 
research laboratory by titering anti-TACA serum IgG and IgM antibodies 
measured at pre-study and on weeks 2, 3, 4, 7, and 9 from study participant 
blood samples. IgM and IgG titers to TACAs will be evaluated by enzyme-linked 
immunosorbent assay (ELISA) of TACAs adhered to plates and by fluorescence-
activated cell sorting (FACS) of TACAs adhered to liposomes.  Both approaches 
will be validated prior to examination of research study samples. Titer will be 
defined as the highest serum dilution yielding an OD405 ≥0.15, in accordance with 
previous studies (1) or a mean fluorescence intensity (MFI) two standard 
deviations higher than background. A positive TACA-directed immune response 
will be defined as an anti-TACA serum antibody titer of 1:40 for a baseline pre-
vaccination titer of 0 or a ≥ 4-fold increase for a baseline titer  > 0 (1).   
2) Delayed type hypersensitivity (DTH) responses to the immunizing mimotope and 
control antigens will be determined by the amount of induration surrounding the 
injection site at 48- 72hrs post-injection. An induration diameter of > 5 mm at 48-
72 hrs post injection will be considered a positive response. Control agents to be 
tested in addition to the P10s-PADRE/ MONTANIDETM ISA 51 VG vaccine 
include Trichophyton Antigen and the Candida antigen. 
3) The effect of a late booster immunization (week 19 immunization) of P10s-
PADRE/ MONTANIDETM ISA 51 VG on the humoral response against TACAs will 
be determined by titering anti-TACA serum IgG and IgM antibodies measured on 
weeks 19 and 21 with the methodology described above.  

 
Version: 19.120 
Date: 11/13/28/2013 
      
10 
4) As part of the standard of care, disease assessments will be performed by PET 
CT or CT CAP at 8 to 12 week intervals.  Enrollment will be done after one such 
assessment. 
c.   Exploratory Objective 
1) The effect of immunizing mimotope on the presence of circulating tumor cells 
(CTCs) will be determined by subjecting pre-and post-vaccination blood (drawn on 
weeks 0 and 21, respectively) to the RARE™ technique – a single-step negative 
enrichment process with a bispecific antibody directed against erythrocytes and 
CD45-postiive cells (StemCell Technologies) (35-41). Determining the presence of 
CTCs prior to and following mimotope administration could provide a potential 
indicator for clinical disease (Müller V, Hayes DF, Pantel K. Breast Cancer Res. 
2006;8(5):110).Though previous efforts to assay CTCs have had technical 
difficulties, we will use a simple protocol that requires only 7.5ml of blood, thereby 
decreasing the likelihood of technical complications.  
5. Patient Population 
 
Eligibility Criteria:  Subjects are eligible for the vaccine study if the following inclusion and 
exclusion criteria are met: 
 
1) 
Female subjects of all races with histologically or cytologically confirmed stage IV 
breast cancer are eligible. The cancer may be newly diagnosed metastatic or 
relapsed after primary or adjunctive therapy and must not have required a treatment 
change for 2 months. Treatments with anti-estrogen therapy or chemotherapy are 
allowed. The chemotherapy regimen cannot contain steroids in the pre or post 
supportive care medications. If a subject is on an investigational drug, the drug must 
be cleared from the body over a period of 4 weeks. 
2) 
 Disease staging will be done according to the American Joint Commission on Cancer 
(AJCC), sixth edition. The breast cancer staging information can be found at the 
following address:  
  
 
 
http://www.cancerstaging.org/education/tnmschema/breast.ppt 
 
3) 
Age 18 years and older of all races and ethnicity. 
4) 
ECOG Performance Status 0 or 1. 
5) 
Subjects must not have an active infection requiring treatment with antibiotics. 
6) 
Subjects must not have other significant medical, surgical or psychiatric conditions, or 
require any medication or treatment, which may interfere with compliance of the 
treatment regimen. 
7) 
Subjects must not have a diagnosis or evidence of organic brain syndrome, 
significant impairment of basal cognitive function or any psychiatric disorder that 
might preclude participation in the full protocol. 
8) 
Subjects must have no other current malignancies.  Subjects with prior history at any 
time of any in situ cancer, including lobular carcinoma of the breast in situ, cervical 
cancer in situ, atypical melanocytic hyperplasia or Clark I melanoma in situ or basal or 
squamous skin cancer are eligible, provided they are disease-free at the time of 
registration. Subjects with other malignancies are eligible if they have been 
continuously disease free for ≥ 5 years prior to the time of registration. 

 
Version: 19.120 
Date: 11/13/28/2013 
      
11 
9) 
Subjects must not have autoimmune disorders or conditions of immunosuppression. 
This includes, but is not limited to being treated with corticosteroids, including oral 
steroids (i.e. prednisone, dexamethasone), continuous use of topical steroid creams 
or ointments or any steroid-containing inhalers. Subjects who have been on systemic 
steroids will require a 6-week washout period. Subjects who discontinue the use of 
these classes of medication for at least 6 weeks prior to registration are eligible if, in 
the judgment of the treating physician, the subject is not likely to require these 
classes of drugs during the treatment period. Replacement doses of steroids for 
subjects with adrenal insufficiency are allowed. 
10) 
Women of childbearing potential must not be pregnant (negative serum pregnancy 
test must be done 48 hours prior to receiving the first dose of study drug) or breast-
feeding, due to the unknown effects of peptide/mimotope vaccines on a fetus or 
infant. 
11) 
Women of childbearing potential must be counseled to use an accepted and effective 
method of contraception (including abstinence) while on treatment and for a period of 
18 months after completing or discontinuing treatment. Accepted methods include 
oral contraceptives, barrier method, IUDs, and abstinence.  
12) 
Subjects must have obtained a white blood cell (WBC) count ≥ 3,000/mm3 and 
platelet count ≥ 100,000/mm3 within 2 weeks prior to registration. 
13) 
Subjects must have a serum glutamic-oxaloacetic transaminase (SGOT)/aspartate 
aminotransferase test (AST) and bilirubin ≤ 2 x institutional upper limit (IUL) of normal 
and serum creatinine ≤ 1.8 mg/dl, all obtained within 2 weeks prior to registration. 
14) 
Subjects must be immunocompetent as measured by responsiveness to one recall 
antigens by skin testing.   
15) 
All subjects who wish to participate in the study must sign an informed consent 
approved by the UAMS Institutional Review Board (IRB). 
16) 
Laboratory tests must be completed within 2 weeks before the first dose. 
17) 
Subjects must not have a diagnosis or evidence of Central Nervous System 
metastases. 
 
6. Investigational New Drug – P10S-PADRE/ MONTANIDETM ISA 51 VG  
 
a. 
General Description: P10s-PADRE is a short peptide (P10s) coupled to PADRE, a 
synthetic, non-natural, peptide that binds with high or intermediate affinity to 15 of 16 of the 
most common HLA-DR types tested to date.  Both components contribute to the efficacy of 
this molecule in stimulating an immune response.  Because of its binding promiscuity, 
PADRE should overcome the problems posed by the extreme polymorphism of HLA-DR 
molecules in the human population. Furthermore, the PADRE peptide was specifically 
engineered as an antigen-presenting molecule for use in humans.  Carbohydrate moieties, 
such as TACAs, typically do not induce T-cell responses.  Thus, P10s, a TACA peptide 
mimotope, was developed.  By coupling P10s to PADRE, the likelihood of generating an 
immune response increases, including T-cell "help" in the vaccine construct designed for 
human use.  
 
 
MONTANIDETM ISA 51 VG is defined as an oil adjuvant containing surfactant based 
mannide monooleate and oil, which is of white mineral oil origin. 
 

 
Version: 19.120 
Date: 11/13/28/2013 
      
12 
b. 
Vaccine Manufacturing and Formulation: AmbioPharm Inc. (North Augusta, SC 29842, 
USA) will synthesize Mimotope P10s covalently linked with PADRE as a sterile lyophilized 
powder manufactured under GMP conditions.  Mimotope P10s-PADRE will be manufactured 
in facilities registered with the FDA (FDA registration #3006446551).   Once P10s-PADRE is 
manufactured by AmbioPharm the lyophilized powder will be shipped to NextPharma 
Technologies  (FDA registration # 2027352, 5340 Eastgate Mall, San Diego, CA 92121) for 
sterile packaging in appropriate sized glass vials under GMP conditions, in quantities of 500 
µg/vial. Advantar Laboratories, Inc. (3030 Bunker Hill Dr., Suite 102, San Diego, CA 92109) 
will perform product release and stability testing on the P10s-PADRE study drug.  The study 
drug will be stored at ≤ -20º C for maximum stability. 
 
 
MONTANIDETM ISA 51 VG will be supplied by SEPPIC, Inc. (30 Two Bridges Road, Suite 
20, Fairfield, NJ 07004-1530). See investigator’s brochure for formulation and storage 
conditions. 
 
c. 
Vaccine Preparation:   
The following doses of P10s-PADRE and MONTANIDETM ISA 51 VG vaccine will be 
administered to subjects subcutaneously in rotating injection sites in the abdomen or 
extremities.  Subjects will receive up to 2.0 ml subcutaneous (SC) injections of the vaccine 
on 5 separate occasions (Weeks 1, 2, 3, 7, and 19). 
 
• 
500 µg P10s-PADRE/ MONTANIDETM ISA 51 VG Vaccine Dose preparation:  The 
UAMS pharmacist will prepare the vaccine dose following the manufacturer’s instructions. 
(See CMC section of the IND). 
 
• 
300 µg P10s-PADRE/ MONTANIDETM ISA 51 VG Vaccine Dose preparation:  The 
UAMS pharmacist will prepare the vaccine dose following the manufacturer’s instructions. 
(See CMC section of the IND). 
 
d. 
DTH Preparation: 
• 
Triychophyton:  per insert instructions. 
• 
Candida:  per insert instructions. 
• 
100 µg P10s-PADRE: The UAMS pharmacist will prepare the DTH dose following 
the manufacturer’s instructions. (See CMC section of the IND). 
 
e. 
Dosing Administration: 
 
The DTH dose will be injected SC on the back of the patient by clinic staff.  
 
f. 
Label Information: The vaccine drug supply (both P10s-PADRE and MONTANIDETM ISA 
51 VG will be labeled with the following statement: “Caution: New Drug – Limited by Federal 
Law to Investigational Use”. 
 
g. 
Agent Ordering: All drug study supplies (P10s-PADRE vials and MONTANIDETM ISA 51 
VG vials) will be sent to the attention of the UAMS Research Pharmacy staff.  Initial drug 
supply order will be placed by primary investigator and both P10s-PADRE vials and 
MONTANIDETM ISA 51 VG vials will be shipped directly to UAMS Research Pharmacy, 
4301 W. Markham Street, # 547-10, Little Rock, AR 72205. 
 
h. 
Agent Accountability: P10s-PADRE and MONTANIDETM ISA 51 VG will be stored in the 
UAMS Pharmacy under the supervision of the research pharmacist who will be responsible 

 
Version: 19.120 
Date: 11/13/28/2013 
      
13 
for maintaining the supply according to the manufacturer’s specifications, dispensing the 
drug for administration and maintaining all accountability logs. Standard NCI accountability 
logs will be used. 
7. Treatment Plan 
 
a. On-study Evaluation: After signing the IRB-approved informed consent form, research 
participants will be assigned to a cohort at the time of consent by a clinical research 
associate (CRA) in the Cancer Clinical Trials Office (CCTO) in the Cancer Institute. All 
research participants will receive the Mimotope P10s-PADRE/ MONTANIDETM ISA 51 VG 
vaccine via subcutaneous (SC) injection following the schedule on the Study Calendar in 
Section 9. 
 
b. Dose Assignment:   The decision to escalate or de-escalate the dose, expand the 
cohort or terminate the study will be based on assessment for DLT, which will require 9 
weeks per subject.  The time to assess a cohort of 3 for DLTs and immune responses is 
thus anticipated to be 13 weeks based on an accrual rate of 2 eligible Stage IV subjects 
per month. 
c. Specimen Handling:  A trained phlebotomist or registered nurse will draw the blood 
required for clinical and research purposes.  Research specimens will be delivered to Dr. 
Kieber-Emmons’ research laboratory.   
  
d. Visit Breakdown: 
 
Pre-study Visit 
• 
Complete medical history and physical examination 
• 
Complete blood count with differential 
• 
Other blood tests include: 
o Collection of serum samples used to measure the immune response to the vaccine.  
o SGOT (Serum glutamic-oxaloacetic transaminase)  
o Alkaline Phosphatase Level 
o LDH (lactate dehydrogenase) 
o GGT (gamma-glutamyl transferase) 
o Creatinine 
o Calcium 
o Albumin 
o Amylase 
o TSH (thyroid stimulating hormone) 
o T4  
o ANA (Anti-nuclear antibody) 
o Bilirubin 
o PT/PTT 
• 
Concomitant medications will be collected from registration to end of treatment.  
• 
Serum pregnancy test for women of childbearing potential. 
• 
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays.  

 
Version: 19.120 
Date: 11/13/28/2013 
      
14 
• 
DTH Skin Test to test the subjects’ immunocompetency. Trichophyton Antigen and the 
Candida antigen will be administered intradermally (id) as control antigens at separate 
locations on the subject’s back, and the resulting induration will be read 48– 72 hours 
later. 
• 
CTC (circulating tumor cells) response to the vaccine will be measured in serum samples 
drawn to determine the presences of CTCs pre- and post-mimotope administration. 
 
48 – 72 hours after Pre-Study Visit 
• 
DTH Skin Test Reading 
 
48 hours prior to dosing 
• 
Serum pregnancy test for women of childbearing potential.  
 
Week 1 
• 
Administration of the P10s-PADRE/MONTANIDETM ISA 51 VG vaccine. The vaccine will 
be administered subcutaneously (SC) by nurses in the Infusion Center at the Cancer 
Institute. 
• 
Post vaccine administration, the following vital signs will be monitored every 15 (+/- 5 
minutes) minutes for one hour:  blood pressure, temperature, and pulse. 
• 
Complete medical history and physical examination 
• 
Toxicity evaluations using the NCI CTCAE (National Cancer Institute Common 
Terminology Criteria for Adverse Events) Version 4.0. Toxicities to be assessed are the 
laboratory parameters listed in the study calendar, as well as any sign or symptom found 
during the history and physical examination not noted at prestudy or on the baseline 
evaluation. Special attention will be paid to signs and symptoms related to injection 
reactions, injection site reactions or symptoms or laboratory findings indicating 
autoimmune toxicities.  
• 
Concomitant medications will be collected from registration to end of treatment.  
 
Week 2 
• 
Administration of the P10s-PADRE/MONTANIDETM ISA 51 VG vaccine. The vaccine will 
be administered subcutaneously (SC) by nurses in the Infusion Center at the Cancer 
Institute. 
• 
Post vaccine administration, the following vital signs will be monitored every 15 (+/- 5 
minutes) minutes for one hour:  blood pressure, temperature, and pulse. 
• 
Complete medical history and physical examination 
• 
Complete blood count with differential  
• 
Toxicity evaluations using the NCI CTCAE (National Cancer Institute Common 
Terminology Criteria for Adverse Events) Version 4.0. Toxicities to be assessed are the 
laboratory parameters listed in the study calendar, as well as any sign or symptom found 
during the history and physical examination not noted at prestudy or on the baseline 
evaluation. Special attention will be paid to signs and symptoms related to injection 
reactions, injection site reactions or symptoms or laboratory findings indicating 
autoimmune toxicities.  
• 
Concomitant medications will be collected from registration to end of treatment.  
• 
Collection of serum samples used to measure the immune response to the vaccine. If 
vaccine is given on these days, the study labs will be drawn before the vaccine is 
administered.  

 
Version: 19.120 
Date: 11/13/28/2013 
      
15 
• 
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays.  
 
 
Week 3 
• 
Administration of the study vaccine. The vaccine will be administered subcutaneously 
(SC) by nurses in the Infusion Center at the Cancer Institute. 
• 
Post vaccine administration, the following vital signs will be monitored every 15 (+/- 5 
minutes) minutes for one hour:  blood pressure, temperature, and pulse. 
• 
Complete medical history and physical examination 
• 
Complete blood count with differential  
• 
Other blood tests include: 
o Collection of serum samples used to measure the immune response to the vaccine. 
If vaccine is given on these days, the study labs will be drawn before the vaccine 
is administered.  
o SGOT (Serum glutamic-oxaloacetic transaminase)  
o Alkaline Phosphatase Level 
o LDH (lactate dehydrogenase) 
o GGT (gamma-glutamyl transferase) 
o Creatinine 
o Calcium 
o Albumin 
o Amylase 
o TSH (thyroid stimulating hormone) 
o T4  
o ANA (Anti-nuclear antibody) 
o PT/PTT 
o Bilirubin 
• 
Toxicity evaluations using the NCI CTCAE (National Cancer Institute Common 
Terminology Criteria for Adverse Events) Version 4.0. Toxicities to be assessed are the 
laboratory parameters listed in the study calendar, as well as any sign or symptom found 
during the history and physical examination not noted at prestudy or on the baseline 
evaluation. Special attention will be paid to signs and symptoms related to injection 
reactions, injection site reactions or symptoms or laboratory findings indicating 
autoimmune toxicities.  
• 
Concomitant medications will be collected from registration to end of treatment.  
• 
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays.  
 
Week 4 
• 
Complete blood count with differential  
• 
Collection of serum samples used to measure the immune response to the vaccine. If 
vaccine is given on these days, the study labs will be drawn before the vaccine is 
administered.  
• 
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays.  

 
Version: 19.120 
Date: 11/13/28/2013 
      
16 
 
Week 5 
• 
Complete blood count with differential  
• 
DTH Skin Test to test the subjects’ immunocompetency. Trichophyton Antigen and the 
Candida antigen will be administered intradermally (id) as control antigens at separate 
locations on the subject’s back, and the resulting induration will be read 48– 72 hours 
later; 100 µg Mimotope P10s-PADRE will be given id alongside the DTH control antigens 
to determine subjects’ DTH response to vaccine. 
 
48 – 72 hours later 
• 
DTH Skin test reading.  
 
Week 6 
• 
Complete blood count with differential  
Week 7 
• 
Administration of the P10s-PADRE/ MONTANIDETM ISA 51 VG vaccine. The vaccine will 
be administered subcutaneously (SC) by nurses in the Infusion Center at the Cancer 
Institute. 
• 
Post vaccine administration, the following vital signs will be monitored every 15 (+/- 5 
minutes) minutes for one hour:  blood pressure, temperature, and pulse. 
• 
Complete medical history and physical examination 
• 
Complete blood count with differential  
• 
Other blood tests include: 
o Collection of serum samples used to measure the immune response to the vaccine. 
If vaccine is given on these days, the study labs will be drawn before the vaccine 
is administered.  
o SGOT (Serum glutamic-oxaloacetic transaminase)  
o Alkaline Phosphatase Level 
o LDH (lactate dehydrogenase) 
o GGT (gamma-glutamyl transferase) 
o Creatinine 
o Calcium 
o Albumin 
o Amylase 
o TSH (thyroid stimulating hormone) 
o T4  
o ANA (Anti-nuclear antibody) 
o PT/PTT 
o Bilirubin 
• 
Toxicity evaluations using the NCI CTCAE (National Cancer Institute Common 
Terminology Criteria for Adverse Events) Version 4.0. Toxicities to be assessed are the 
laboratory parameters listed in the study calendar, as well as any sign or symptom found 
during the history and physical examination not noted at prestudy or on the baseline 
evaluation. Special attention will be paid to signs and symptoms related to injection 
reactions, injection site reactions or symptoms or laboratory findings indicating 
autoimmune toxicities.  
• 
Concomitant medications will be collected from registration to end of treatment.  

 
Version: 19.120 
Date: 11/13/28/2013 
      
17 
• 
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays.  
 
Week 8 
• 
Complete blood count with differential  
 
Week 9 
• 
Complete medical history and physical examination 
• 
Complete blood count with differential  
• 
Other blood tests include: 
o Collection of serum samples used to measure the immune response to the vaccine. 
If vaccine is given on these days, the study labs will be drawn before the vaccine 
is administered.  
o SGOT (Serum glutamic-oxaloacetic transaminase)  
o Alkaline Phosphatase Level 
o LDH (lactate dehydrogenase) 
o GGT (gamma-glutamyl transferase) 
o Creatinine 
o Calcium 
o Albumin 
o Amylase 
o TSH (thyroid stimulating hormone) 
o T4  
o ANA (Anti-nuclear antibody) 
o PT/PTT 
o Bilirubin 
• 
Toxicity evaluations using the NCI CTCAE (National Cancer Institute Common 
Terminology Criteria for Adverse Events) Version 4.0. Toxicities to be assessed are the 
laboratory parameters listed in the study calendar, as well as any sign or symptom found 
during the history and physical examination not noted at prestudy or on the baseline 
evaluation. Special attention will be paid to signs and symptoms related to injection 
reactions, injection site reactions or symptoms or laboratory findings indicating 
autoimmune toxicities. 
• 
Concomitant medications will be collected from registration to end of treatment.  
• 
DTH Skin Test to test the subjects’ immunocompetency. Trichophyton Antigen and the 
Candida antigen will be administered intradermally (id) as control antigens at separate 
locations on the subject’s back, and the resulting induration will be read 48– 72 hours 
later; 100 µg Mimotope P10s-PADRE will be given id alongside the control antigens to 
determine subjects’ DTH response to vaccine. 
• 
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays.  
 
48 – 72 hours later 
• 
DTH Skin test reading. 
 
Week 19 

 
Version: 19.120 
Date: 11/13/28/2013 
      
18 
• 
Administration of the P10s-PADRE/MONTANIDETM ISA 51 VG vaccine. The vaccine will 
be administered subcutaneously (SC) by nurses in the Infusion Center at the Cancer 
Institute. 
• 
Post vaccine administration, the following vital signs will be monitored every 15 (+/- 5 
minutes) minutes for one hour:  blood pressure, temperature, and pulse. 
• 
Complete medical history and physical examination 
• 
Complete blood count with differential  
• 
Other blood tests include: 
o Collection of serum samples used to measure the immune response to the vaccine. 
If vaccine is given on these days, the study labs will be drawn before the vaccine 
is administered.  
o SGOT (Serum glutamic-oxaloacetic transaminase)  
o Alkaline Phosphatase Level 
o LDH (lactate dehydrogenase) 
o GGT (gamma-glutamyl transferase) 
o Creatinine 
o Calcium 
o Albumin 
o Amylase 
o TSH (thyroid stimulating hormone) 
o T4  
o ANA (Anti-nuclear antibody) 
o PT/PTT 
o Bilirubin 
• 
Toxicity evaluations using the NCI CTCAE (National Cancer Institute Common 
Terminology Criteria for Adverse Events) Version 4.0. Toxicities to be assessed are the 
laboratory parameters listed in the study calendar, as well as any sign or symptom found 
during the history and physical examination not noted at prestudy or on the baseline 
evaluation. Special attention will be paid to signs and symptoms related to injection 
reactions, injection site reactions or symptoms or laboratory findings indicating 
autoimmune toxicities.  
• 
Concomitant medications will be collected from registration to end of treatment.  
• 
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays.  
 
Week 20 
• 
Complete blood count with differential  
 
Week 21 
• 
Complete medical history and physical examination 
• 
Complete blood count with differential  
• 
Other blood tests include: 
o Collection of serum samples used to measure the immune response to the vaccine. 
If vaccine is given on these days, the study labs will be drawn before the vaccine 
is administered.  
o SGOT (Serum glutamic-oxaloacetic transaminase)  
o Alkaline Phosphatase Level 
o LDH (lactate dehydrogenase) 

 
Version: 19.120 
Date: 11/13/28/2013 
      
19 
o GGT (gamma-glutamyl transferase) 
o Creatinine 
o Calcium 
o Albumin 
o Amylase 
o TSH (thyroid stimulating hormone) 
o T4  
o ANA (Anti-nuclear antibody) 
o PT/PTT 
o Bilirubin 
• 
Toxicity evaluations using the NCI CTCAE (National Cancer Institute Common 
Terminology Criteria for Adverse Events) Version 4.0. Toxicities to be assessed are the 
laboratory parameters listed in the study calendar, as well as any sign or symptom found 
during the history and physical examination not noted at prestudy or on the baseline 
evaluation. Special attention will be paid to signs and symptoms related to injection 
reactions, injection site reactions or symptoms or laboratory findings indicating 
autoimmune toxicities.  
• 
Concomitant medications will be collected from registration to end of treatment.  
• 
DTH Skin Test to test the subjects’ immunocompetency. Trichophyton Antigen and the 
Candida antigen will be administered intradermally (id) as control antigens at separate 
locations on the subject’s back, and the resulting induration will be read 48– 72 hours 
later; 100 µg Mimotope P10s-PADRE will be given id alongside the control antigens to 
determine subjects’ DTH response to vaccine. 
• 
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays.  
• 
CTC (circulting tumor cells) response to the vaccine will be measured in serum samples 
drawn to determine the presences of CTCs pre- and post-mimotope administration. 
 
48 – 72 hours later 
• 
DTH Skin test reading.  
 
Week 24 
• 
Complete medical history and physical examination 
• 
Complete blood count with differential  
• 
Other blood tests include: 
o Collection of serum samples used to measure the immune response to the vaccine. 
If vaccine is given on these days, the study labs will be drawn before the vaccine 
is administered.  
o SGOT (Serum glutamic-oxaloacetic transaminase)  
o Alkaline Phosphatase Level 
o LDH (lactate dehydrogenase) 
o GGT (gamma-glutamyl transferase) 
o Creatinine 
o Calcium 
o Albumin 
o Amylase 
o TSH (thyroid stimulating hormone) 
o T4  

 
Version: 19.120 
Date: 11/13/28/2013 
      
20 
o ANA (Anti-nuclear antibody) 
o PT/PTT 
o Bilirubin 
• 
Toxicity evaluations using the NCI CTCAE (National Cancer Institute Common 
Terminology Criteria for Adverse Events) Version 4.0. Toxicities to be assessed are the 
laboratory parameters listed in the study calendar, as well as any sign or symptom found 
during the history and physical examination not noted at prestudy or on the baseline 
evaluation. Special attention will be paid to signs and symptoms related to injection 
reactions, injection site reactions or symptoms or laboratory findings indicating 
autoimmune toxicities.  
• 
Concomitant medications will be collected from registration to end of treatment.  
Study lab samples. Up to 40 mLs of blood  will be collected in red and purple tubes for 
circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays. 
 
Follow up 
Follow up will occur at next scheduled standard of care visit every six months for five years and 
then yearly until death. 
 
• 
Collection of serum samples used to measure the immune response to the vaccine.  
 
• 
  
 
e. 
Prohibited Medications: Systemic steroids are prohibited. If a subject wishing to participate 
in the study has been on systemic steroids, a 6-week clearing of the drug will be required 
prior to participation in the study. 
 
f. 
Dose Limiting Criteria: Subjects who develop any grade 3 or greater toxicity as per NCI 
CTCAE v. 4 toxicity criteria, Grade 2 or greater autoimmune toxicity with the exception of 
vitiligo, or Grade 2 or greater hypersensitivity reactions will be treated and referred for 
additional care as indicated with systemic steroids, topical steroids, epinephrine or Benadryl.  
These subjects will be removed from the study and will be followed as per protocol defined 
monitoring and/or until resolution of toxicity. 
 
 
g. Dose Assignment: Subjects will be treated with vaccine admixed with MONTANIDETM ISA 
51 VG on weeks 1, 2, 3, 7 and 19 in cohorts according to the following dosing diagram: 

 
Version: 19.120 
Date: 11/13/28/2013 
      
21 
 
 
 
 
DLT and the immune-response endpoints are defined in Section 4, “Trial Objectives”. Upon 
evaluation of all subjects in a cohort (3 or 6 per dose level), the decision whether to 
escalate, de-escalate or stop will proceed according to the cohort-appropriate schedule 
shown in Tables 1, 2, and 3.   
 
 
 
 
Table 1: Toxicity Decision Rules for Initial Cohort (300 μg Vaccine Dose) 
DLTs/ Cohort1 
Action 
0/3 
Begin accrual to Escalation Cohort 
1/3 
Expand Initial Cohort to 6 subjects 
1/6 
Begin accrual to Escalation Cohort 
2/6, 3/6, or 4/6 
Begin Accrual to De-Escalation Cohort 
2/3 or 3/3 
Begin Accrual to De-Escalation Cohort 
1 DLT = Dose-Limiting Toxicity 
 
INITIAL  
COHORT 
300 µg  
Vaccine Dose  
ESCALATION  
COHORT 
500 µg 
Vaccine Dose  
DE-ESCALATION  
COHORT 
100 µg 
Vaccine Dose  

 
Version: 19.120 
Date: 11/13/28/2013 
      
22 
Table 2: Toxicity Decision Rules for Escalation Cohort (500 μg Vaccine 
Dose) 
DLTs/ Cohort1 
Action 
0/3 or 1/3 
Expand cohort to 6 subjects 
1/6 
Stop:  Declare escalation dose to be MTD2 
2/6, 3/6, or 4/6 
Stop:  Previous dose level is MTD2 
2/3 or 3/3 
Stop:  Previous dose level is MTD2 
1 DLT = Dose-Limiting Toxicity 
2 MTD = Maximum Tolerated Dose 
 
Table 3: Toxicity Decision Rules for De-Escalation Cohort (100 μg 
Vaccine Dose) 
DLTs/ Cohort1 
Action 
0/3 or 1/3 
Expand cohort to 6 subjects. 
1/6 
Stop:  De-escalation dose level is the MTD2 
2/6, 3/6, or 4/6 
Stop:  De-escalation dose level is above 
MTD2 
2/3 or 3/3 
Stop:  De-escalation dose level is above 
MTD2 
1 DLT = Dose-Limiting Toxicity 
2 MTD = Maximum Tolerated Dose 
 
 
If the initial-cohort dose of 300 μg P10s-PADRE is declared to be the MTD and only 3 
subjects were enrolled into the initial cohort, then this cohort will be expanded to 6 to assure 
that 6 subjects are treated at the MTD. The CRA for the trial will assign the subjects to 
cohorts and communicate the assignments to the research nurse, who will communicate 
them to the investigator. Only one cohort will be open to enrollment at a time.  After the last 
non-boost dose in each cohort, the study will have a 3-week hold pending the cohort’s DLT 
evaluation. Staggered dosing for the first two patients within each cohort will allow an 
interval of one week observation for toxicity following the first injection in the previous 
patient.  The research pharmacist will be notified by orders of the registration and cohort 
dose. Expansion of the cohort to 6, or enrollment to the next cohort, may start once the 
previous 3 subjects complete their week-9 serology and are evaluated for DLT. Subjects 
who withdraw from the dose-escalation study without a DLT will be replaced.  Subjects who 
withdraw from the dose- de-escalation study without a DLT will be replaced.  Subjects who 
withdraw with a DLT will be considered evaluable for MTD determination. 
 
8. Risks and Toxicities To Be Monitored 
a. 
Potential Toxicities, Risks and Precautions: 
 
Procedure 
Risks 
Measures to Minimize Risks 
Complete 
history 
and 
physical exam, 
including blood 
chemistries 
Identification 
of 
previously 
unknown 
condition 
Qualified health care provider to evaluate 
potential subject 
 
Research records are kept in a locked area 
accessible only by study personnel. 

 
Version: 19.120 
Date: 11/13/28/2013 
      
23 
Procedure 
Risks 
Measures to Minimize Risks 
Administration 
of study vaccine  
 
Experimental agent 
may be toxic or 
harmful. 
 
First time use in 
humans 
 
 
 
 
 
 
 
 
Risk of local reactions 
(i.e. swelling, redness, 
tenderness, itching, 
extravasations) 
 
 
Potential for side 
effects ranging from 
hematologic toxicities 
and hypersensitivity 
reactions to 
anaphylaxis   
 
Unanticipated risks 
  
 
 
 
Unknown risks  
 
 
Careful monitoring by clinic visits and 24 
hour, 7 days per week physicians on call for 
unexpected problems 
 
Only non-pregnant, non-lactating females 
may participate.  The use of contraception 
during the study and the use of contraception 
for 18 months post completion of the trial are 
required. 
 
Frequent laboratory tests including complete 
blood count (CBC) with differential, liver 
function tests, etc. 
 
Close and frequent monitoring of subjects by 
qualified staff 
 
DTH assay to monitor for hypersensitivity 
reactions  
 
Emergency equipment including crash carts, 
advanced 
cardiac 
life 
support 
(ACLS) 
certified staff and rescue medications such as 
Benadryl, epinephrine, high dose steroids, 
etc. will be on-site during administration. 
 
 
The Medical Monitor will review all toxicities 
on a regular basis and will be available to aid 
subjects as needed. 
 
The study drug may be discontinued. 
 
This research is being conducted at an 
experienced clinical research center. 
 
Reporting and monitoring mechanisms are in 
place for AEs, serious adverse events (SAEs) 
and unanticipated problems. 
Administration 
of 
MONTANIDE™ 
ISA 51 VG 
 
Dermatology/Skin: 
local erythema, rash, 
pruritis 
 
Gastrointestinal: 
diarrhea, anorexia, 
nausea, vomiting, 
abnormal taste 
 
Hepatic: elevated 
Careful monitoring by clinic visits and 24 
hour, 7 days per week physicians on call for 
unexpected problems 
 
Only non-pregnant, non-lactating females 
may participate.  The use of contraception 
during the study and the use of contraception 
for 18 months post completion of the trial are 
required. 
 

 
Version: 19.120 
Date: 11/13/28/2013 
      
24 
Procedure 
Risks 
Measures to Minimize Risks 
hepatic enzymes, 
hypo-albuminemia with 
prolonged treatment 
 
Neurology: confusion, 
neuropathies 
Possible CNS toxicity 
in those with known 
metastatic disease to 
brain 
 
Pulmonary: dyspnea 
(due to fluid retention 
and capillary leak 
syndrome), pleuritis 
 
Cardiovascular: 
hypertension, cardiac 
arrhythmias, atrial 
fibrillation, pericarditis 
 
Pain: headache, 
arthralgias, bone pain, 
abdominal pain, chest 
pain, myalgia 
 
 
Fever, flu-like 
syndrome (chills, 
rigors, myalgias), 
fatigue, headache, 
abnormal labs 
including blood urea 
nitrogen and albumin 
Thorough review of scans used for disease 
assessment performed  prior to participation 
in study, including brain imaging. 
 
Frequent laboratory tests including CBC with 
differential, liver function tests, etc. 
 
 
Close and frequent monitoring of subjects by 
qualified staff. 
 
 
DTH assay to monitor for hypersensitivity 
reactions  
 
 
Emergency equipment including crash carts, 
ACLS certified staff and rescue medications 
such as Benadryl, epinephrine, high dose 
steroids, 
etc. 
will 
be 
on-site 
during 
administration. 
 
 
The Medical Monitor will review all toxicities 
on a regular basis and will be available to aid 
subjects as needed. 
 
 
The study drug may be discontinued. 
 
 
This research is being conducted at an 
experienced clinical research center in the 
cancer institute. 
 
 
Reporting and monitoring mechanisms are in 
place for AEs, SAEs and unanticipated 
problems.  
Collection 
of 
blood samples  
 
 
Pain, bruising at the 
injection site and rarely 
infection 
 
 
 
Discovery of previously 
unknown conditions 
 
Possible breach of 
Experienced personnel will perform the 
phlebotomies using approved techniques. 
 
Pressure and dressings will be used to 
minimize pain, bruising and infection. 
  
Research records are kept in a locked area 
accessible only by study personnel. 
 
Subject study numbers will be used for 

 
Version: 19.120 
Date: 11/13/28/2013 
      
25 
Procedure 
Risks 
Measures to Minimize Risks 
confidentiality 
identification of samples so that they may be 
retained 
for 
future 
research 
and 
confidentiality is ensured. 
Serum 
pregnancy 
testing 
 
Discovery of previously 
unknown conditions 
 
Possible breach of 
confidentiality 
Research records are kept in a locked area 
accessible only by study personnel. 
 
 
Subjects will only be identified by study 
numbers on all research documents. 
Serum 
for 
immunologic 
evaluation 
Discovery of previously 
unknown conditions 
 
Possible breach of 
confidentiality 
Research records are kept in a locked area 
accessible only by study personnel. 
 
 
Subjects will only be identified by study 
numbers on all research documents. 
Skin test and 
DTH assay – 
performed 
at 
various 
locations 
on 
subjects’ backs 
Pain, bruising at the 
injection site, and 
rarely infection 
 
 
 
Potential for allergic 
reaction including 
anaphylaxis 
 
 
 
 
 
 
 
 
 
 
Discovery of previously 
unknown conditions 
Experienced personnel will perform the 
injections using approved techniques. 
 
Pressure and dressings will be used to 
minimize pain, bruising and infection. 
 
Emergency equipment including crash carts, 
ACLS certified staff and rescue medications 
such as Benadryl, epinephrine, high dose 
steroids, 
etc. 
will 
be 
on-site 
during 
administration. 
 
This research is being conducted at an 
experienced clinical research center. 
 
Reporting and monitoring mechanisms are in 
AEs, SAEs and unanticipated problems. 
  
Research records are kept in a locked area 
accessible only by study personnel. 
 
Subjects will only be identified by study 
numbers on all research documents. 
Collection 
of 
data 
Possible breach of 
confidentiality  
 
Research records are kept in a locked area 
accessible only by study personnel. 
 
Subjects will only be identified by study 
numbers on all research documents. 
 
Investigators will provide certification of 
completion of human subject protection 
training course. 
 

 
Version: 19.120 
Date: 11/13/28/2013 
      
26 
Procedure 
Risks 
Measures to Minimize Risks 
UAMS shall retain the records and reports for 
2 years after a marketing application is 
approved for the drug; or, if an application is 
not approved for the drug, until 2 years after 
shipment and delivery, or for 2 years after the 
IND is closed and discontinued with the Food 
and Drug Administration (FDA). . After such 
time all study records will be destroyed as 
well as the links between identifiers of the 
research subjects and their research study 
numbers 
according 
to 
UAMS’ 
record 
destruction policy  
 
Subjects will receive up to 5 planned vaccine doses unless they withdraw from the study or 
develop a dose limiting event as stated above, at which time they will discontinue the 
injections and be removed from the study. There will be no dose modifications for toxicity. 
Special attention will be given to toxicities mediated by autoimmune mechanisms, such as 
colitis, thyroiditis or systemic lupus erythematosus (SLE), as well as to injection-site local 
reactions or allergic reactions. The NCI CTCAE Version 4.0 will be used for toxicity and SAE 
reporting. A copy of the CTCAE Version 4.0 can be downloaded from the Cancer Therapy 
Evaluation Program (CTEP) home page or viewed in APPENDIX A. All appropriate 
treatment areas have access to a copy of the CTCAE Version 4.0.  
 
Any subject may voluntarily revoke consent and withdraw from the study at any time. A 
subject may be terminated early for the following conditions: (i) non-compliance, (ii) an 
unrelated intercurrent illness that may affect assessment or place the subject at risk for AEs 
or require systemic steroids, (iii) deterioration in performance status so as to make 
participation a hardship for the subject, (iv) for toxicity as determined by the DLT criteria, or 
(v) for any reason that the investigator feels it is not in the subject’s best interest to continue. 
 
b. Benefits: As this is the first time Mimotope P10s-PADRE will be administered to humans, 
there are no clearly defined benefits to subjects of this study.  However, this vaccine may 
potentiate an immune response which could improve median progression-free survival 
and overall survival of cancer patients. 
c. Injury: For DOD-funded research, participants can receive medical care at an Army 
hospital or clinic free of charge.  If participants pay out-of-pocket for medical care 
elsewhere for injuries caused by this research study, they should contact the Principal 
Investigator.  If the issue cannot be resolved, contact the U.S. Army Medical Research 
and Materiel Command (USAMRMC) Office of the Staff Judge Advocate (legal office) at 
(301) 619-7663/2221 
9. Specimen Handling 
 
Samples will be stored in the UAMS Tissue Procurement Facility, located in room B.018 of the 
hospital. Cryotubes that contain specimens will be labeled with barcodes. The coded number is 
linked to patient identification and time of blood draw. Specimens will be stored frozen at -80ºC 

 
Version: 19.120 
Date: 11/13/28/2013 
      
27 
or lower. Samples will be kept for maximum of 5 years. At the end of year 5, any unused 
specimen will be mixed with bleach and disposed in a sink. All specimen containers (tubes and 
containers) will be disposed into biohazard red plastic bags according to UAMS Occupational 
Health & Safety’s policy. Red bag waste will be picked up by occupational Health & Safety for 
final disposal.  
 
 

28 
 
10. Study Calendar 
TEST/EVENT 
WEEK 7 
 
Prestudy1 
48 hrs 
later 
48 hours 
prior to 
dosing 
1 
2 
3 
4 
5 
48 hrs 
later 
6 
7 
8 
9 
48 hrs 
later  
19 
20 
21 
48 hrs 
later  
24 
Follow 
up10 
Informed Consent 
x 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Vaccination with 
Mimotope P10s-PADRE/ 
MONTANIDETM ISA 51 
VG 
 
 
 
X9 
X9 
X9 
 
 
 
 
X9 
 
 
 
X9 
 
 
 
 
 
Adverse Events 
 
 
 
x 
x 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Concomitant medications 
8 
x 
 
 
x 
x 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
History/Physical Exam/ 
x 
 
 
x 
x 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
CBC with Differential 
x 
 
 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
 
x 
 
SGOT 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Bilirubin 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Alkaline Phosphatase 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
LDH (lactate 
dehydrogenase) 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
GGT (gamma-glutamyl 
transferase test) 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Creatinine 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Calcium 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Albumin 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Amylase 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
TSH (thyroid stimulating 
hormone) 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
T4 
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Anti-nuclear Antibody 
(ANA) 
x 
x 
x 
x 
x 
x 
 
 x 
 
PT/PTT  
x 
 
 
 
 
x 
 
 
 
 
x 
 
x 
 
x 
 
x 
 
x 
 
Serum Pregnancy Test2 
x 
 
x 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Study Lab3 
x4 
 
 
 
x 
x 
x 
 
 
 
x 
 
x 
 
x 
 
x4 
 
x4 
x 
DTH Skin Test5,6 
x5 
 
 
 
 
 
 
x6 
 
 
 
 
x6 
 
 
 
x6 
 
 
 
Read DTH Skin Test5,6 
 
x5 
 
 
 
 
 
 
x6 
 
 
 
 
x6 
 
 
 
x6 
 
 

 
Version: 19.120 
Date: 11/13/28/2013 
      
29 
1Prestudy is to be completed within 14 days before beginning injections. 
2For women of child bearing potential, a serum pregnancy test must be done at prestudy and within 48 hours prior to dosing. One test may suffice for both 
3 Study lab samples. Up to 40 mLs of blood  will be collected in purple tubes for circulating tumor cell evaluation, TACA (tumor associated carbohydrate antigens) 
assessment, and immune effector assays. Specimens will be picked up by Dr. Kieber-Emmons or a member of his research staff. Call 526-5930 for pick-up. 
4An additional 6.0 ml of blood will be taken during these study labs (Pre-Study, Week 21, and Week 24) to assay circulating tumor cells (CTCs). 
5The DTH skin test at prestudy will test the subjects’ immunocompetency. Control antigens, Trichophyton Antigen and Candida antigen, will be administered id and 
the resulting induration will be ready at 48 - 72 hours post injection. 
6 The DTH skin tests on weeks 5, 9 and 21 will test the DTH response to Mimotope P10s-PADRE. Trichophyton Antigen and Candida antigen will be administered id 
alongside Mimotope P10s-PADRE as control antigens to assess immunocompetency. DTH skin test indurations will be read at 48 – 72 hours post injection. 
7 Visits must occur at set time points mentioned in the study calendar (+/- 3 days) with the exception of reading the skin test with must occur 48-72 hours post 
administration. 
8 Concomitant medications will be collected from registration to end of treatment. 
9 Post vaccine administration, the following vital signs will be monitored every 15 minutes (+/- 5 minutes) for one hour: blood pressure, temperature, and pulse.  
10 Follow up visits shall begin with the next scheduled standard of care appointment and then shall occur every six months for five years and then yearly until death. 

30 
 
 
11. Criteria for Evaluation 
Primary Objective – Safety:   
 
• 
Determination of DLT: See Section 8, “Risks and Toxicities to be Monitored”. Any event 
that meets the dose limiting criteria will be determined a dose limiting toxicity. 
 
Secondary Objectives – Immune Response: 
 
• 
Immunological Evaluation: Serum will be collected at the Study Lab time points 
indicated in the study calendar in Section 10. Samples will be collected and stored by the 
UAMS Cancer Institute Tissue Bank following established Tissue Bank SOPs. IgM and 
IgG titers to TACAs will be evaluated by ELISA on TACA adhered to plates and FACS 
analysis of TACA adhered to liposomes.  A positive TACA-directed immune response 
will be defined as an anti-TACA serum antibody titer of 1:40 for a baseline pre-
vaccination titer of 0 or a ≥ 4-fold increase over baseline titer > 0 (1, 11).  Subjects will 
be judged to have had an adequate immune response if they have a positive TACA-
directed immune response at any one of the first five designated time points following 
vaccine administration (Weeks 2, 3, 4, 7 and 9). The value of a booster immunization will 
be determined by anti-TACA IgM and IgG titers obtained from study labs collected on 
week 21. 
 
• 
Determination of DTH Responses: To evaluate DTH responses, study subjects will be 
skin-tested according to the time points in the study calendar against the Mimotope 
P10s-PADRE, Trichophyton antigen and Candida antigen. The latter 2 antigens serve as 
control antigens. DTH responses to the control antigens will be assessed at pre-study to 
determine immunocompetency of study subjects. All antigens will be administered id at 
separate locations on the subject’s back.  Induration will be measured using calipers or a 
ruler and reported in mm across one diameter at 48 – 72 hours post injection. An 
induration > 5 mm across at least one diameter of 1 or more DTH antigens will be 
considered positive.  
 
Exploratory Objective: 
• 
Determination of presence of CTCs: To evaluate the presence of CTC’s pre- and post-
mimotope administration, 1 EDTA 6.0 mL Vacutainer test tube for blood to evaluate 
circulating tumor cell markers. Multiparameter flow cytometry analysis will be used for 
reducing false positives from background noise. Multiple reagents grouped in conjugates 
with different fluorochromes including at least 1 for negative selection (CD45, may be 
also CD48). The positive identification will be on the basis of EpCAM and/or folate-
Alexa488. Assay sensitivity will be determined beyond the scope of this protocol by 
spiking normal- non cancer patient- deidentified donor blood obtained from UAMS blood 
bank with known numbers of tumor cells (e.g. MDA 231) to determine the detection limit.  
 
 
 
 
 
 

 
Version: 19.120 
Date: 11/13/28/2013 
      
31 
12. Statistical Considerations 
 
a. 
Sample Size, Study Duration and General Considerations: Because each subject will 
receive multiple vaccine injections at a constant dose over an extended period of time, the 
second and third subject of the same dose cohort will be enrolled, as available, before the 
first subject has finished all scheduled injections.  However, if two or more subjects are 
enrolled on the same day, then their injection schedules will be staggered at least one week 
apart. The decision to escalate or de-escalate the dose, expand the cohort, or terminate the 
study will be based on assessment for DLTs, which will require 9 weeks per subject. Upon 
evaluation of all subjects in a cohort (3-6/dose), dose escalation will proceed according to 
the schedule shown in Tables 1, 2, and 3. Except for the fact that they allow for possible de-
escalation from the initial dose of 300 µg, Tables 1, 2, and 3 constitute the toxicity-based 
“traditional” design of Storer (43). The time to assess a cohort of 3 for DLTs is anticipated to 
be 13 weeks based on an accrual rate of 2 eligible Stage IV subjects per month. If no DLTs 
occur in the initial and final cohorts, then this study will require nine subjects.  Subjects who 
withdraw from the dose-escalation study without a DLT will be replaced.  Subjects who 
withdraw from the dose-de-escalation study without a DLT will be replaced. Subjects who 
withdraw with a DLT will be considered evaluable for MTD determination. Given the 
recruitment and immunization/evaluation schedule, we expect this study to be completed in 
a minimum of 12 and a maximum of 24 months. 
 
It should be noted that the primary objective of this study is to ensure the safety of the 
mimotope vaccine.  Because of sample size, these studies only provide a qualitative 
assessment of vaccine immunogenicity. However, our immunization schedule should favor 
the generation of antibodies (1, 11, 42).  A positive TACA-directed immune response will be 
defined as an anti-TACA serum antibody titer of 1:40 for a baseline pre-vaccination titer of 0 
or a ≥ 4-fold increase for a baseline titer > 0 (1, 11).  A subject will be judged to have had an 
adequate immune response if they have a positive TACA-directed immune response at any 
one of the first five indicated time points following vaccine administration on the study 
calendar (Section 10, Study Lab Weeks 2, 3, 4, 7 and 9). An additional immune-response 
endpoint will be the DTH response to Mimotope P10s-PADRE. The components of the DTH 
response, namely, the induration in mm across two diameters, will be tabulated by antigen 
(the mimotope vs. the two DTH controls) for each subject at the indicated time points on the 
study calendar. An induration of >5mm in at least one diameter will be considered a positive 
DTH response.   
 
b. 
Data Analysis Plan: Toxicity will be graded according to the NCI CTCAE Version 4.0.  All 
toxicities observed in a study subject will be enumerated and reported in terms of type 
(organ affected or laboratory determination such as absolute neutrophil count), severity (by 
NCI Common Toxicity Criteria (CTC) and nadir or maximum values for the laboratory 
measures), time of onset (i.e. dose number), duration and reversibility or outcome.  A 
toxicity meeting the dose limiting criteria as stated above will be scored as a DLT. The 
number and proportion of subjects in each dose cohort who suffer a DLT will be reported. 
The number of DLTs observed will be reported by dose cohort, both overall and also broken 
down by CTCAE category.  
 
To meet the secondary objective of determining the humoral response against TACAs, the 
anti-TACA serum titers measured from subjects’ blood samples will be collected at pre-study 
and on weeks 2, 3, 4, 7, and 9. IgM and IgG titers to TACAs will be evaluated by ELISA and 

 
Version: 19.120 
Date: 11/13/28/2013 
      
32 
FACS analysis.  Titer will be defined as the highest serum dilution yielding an OD405 ≥0.15, 
in accordance with previous studies (1) or an MFI two standard deviations higher than 
background. A positive TACA-directed immune response will be defined as an anti-TACA 
serum antibody titer of 1:40 post-vaccination when the baseline pre-vaccination titer was 0, 
or a ≥ 4-fold increase in titer post-vaccination when the baseline titer was greater than 0. 
The number and proportion of positive TACA-directed immune responses at each time point 
will be reported, both overall and by dose cohort. The number and proportion of subjects 
with at least one positive TACA-directed immune response will also be reported, both overall 
and by dose cohort.  Medians and quartiles of titer will also be reported at each time point 
for any dose cohort of size 6.   
 
To meet the secondary objective of analyzing the DTH response, we will measure the 
induration in mm across two diameters at the injection sites of the mimotope and control 
antigens. This will be done for each study subject at all indicated time points on the study 
calendar. An induration of >5mm across at least one diameter will be considered a positive 
DTH response to that antigen at that time point. The number of subjects who show at least 
one positive DTH response to mimotope during the study will be reported as the number 
and proportion in each dose cohort. The number of positive DTH responses to mimotope at 
each time point will be reported as the number and proportion in each dose cohort. Positive 
DTH responses to the control antigens will be summarized the same way. Additionally, the 
two induration measurements per injection site, as well as their product (in mm2 ), will be 
graphed as subject profiles over time, and summarized by antigen, dose group, and time as 
means and standard deviations.  
 
To meet the secondary objective of determining sustainability of the immune response, the 
subject’s TACA-directed immune titer at Week 19 will be compared to her TACA-directed 
immune titer at Week 9. The number and percentage of subjects with a sustained immune 
response will be reported in the aggregate and by dose cohort if more than one dose cohort 
is enrolled.  In addition, the ratio of week-19 and week-9 titers will be plotted as dot plots 
and summarized as the mean, median and range. 
                  
 
To meet the secondary objective of determining the immune response to the week-19 
booster immunization, the subject’s TACA-directed immune titer at Week 21 will be 
expressed as a ratio relative to her TACA-directed immune titer at Week 19.  This ratio will 
be summarized as the mean, median and range, and plotted as dot plots.  Subjects in whom 
the week-21 titer is more than 2-fold higher than the week-19 titer will be considered as 
having shown a boosted response to the booster immunization. The number and 
percentage of subjects showing a boosted response will be reported in the aggregate and 
by dose cohort if more than one cohort is enrolled.   
To meet the exploratory objective of determining the vaccine’s effect on the presence of 
CTCs at datable levels, the subject will be classified as Yes versus No for CTCs Detected, 
both at baseline and at Week 21, based on the outcomes of the assay described in 
paragraph d. of Section 9 above. The number and proportion of subjects with CTCs 
detected at each time will be reported in the form of a 2x2 contingency table for paired data, 
in order to facilitate the visualization of how many subjects change from Yes to No after 
completing the vaccination schedule. 
 
 
Any deviations from the above analysis plan will be reported to the U.S. Army Medical 

 
Version: 19.120 
Date: 11/13/28/2013 
      
33 
Research and Material Command Human Subjects Research Review Board (HSRRB), the 
Food and Drug Administration (FDA), and the UAMS Institutional Review Board  
 
c. 
Missing, Unused and Spurious Data: Missing data will be treated as missing, and will not 
be imputed.  All data collected will necessarily be reported to the FDA.  Spurious data will be 
corrected at the source document.  Any data documented as spurious that is unable to be 
corrected at the source will be treated as missing. 
13. Registration Guidelines  
 
Screening logs will be maintained by the study nurses. Subject registration will occur after the 
IRB-approved consent is signed and eligibility has been confirmed. The subjects will be 
registered in the CCTO and assigned a study number by the CRA. The study number will be 
used for identification of the research subject during the study.  
 
14. Data Submission Schedule 
 
Data must be submitted according to protocol requirements for ALL subjects registered, whether 
or not assigned treatment is administered. This includes subjects deemed to be ineligible to 
participate in the study or for whom documentation is inadequate to determine eligibility.  Data 
obtained during the study will be collected at each subject visit and entered into the protocol 
database. Subjects will be registered in C3PR, a cancer Biomedical Informatics Grid (caBIG®, 
NCI) application. Data will be entered into OpenClinica through electronic web-based case report 
forms (CRFs) which replicate the paper CRFs attached to this protocol. OpenClinica is a secure 
open source system for electronic data capture and clinical data management. 
 
UAMS shall retain the records and reports for 2 years after a marketing application is approved 
for the drug; or, if an application is not approved for the drug, until 2 years after shipment and 
delivery, or for 2 years after the IND is closed and discontinued with the Food and Drug 
Administration (FDA). . After such time all study records will be destroyed as well as the links 
between identifiers of the research subjects and their research study numbers according to 
UAMS’ record destruction policy.  
 
15. Ethical and Regulatory Considerations 
 
The following must be observed to comply with FDA regulations for the conduct and monitoring 
of clinical investigations. The following also represents sound research practice: 
All study personnel must have completed training in good clinical practice (GCP) and protection 
of human subjects. 
 
a. 
Recruitment and Informed Consent: Research subjects will be recruited from the breast 
cancer clinics (Medical Oncology and Ladies Oncology Clinics) at the Winthrop P. 
Rockefeller Cancer Institute on the UAMS campus. The research subjects will be identified 
by the research nurse and/or physician. Prior to any research activities, the research subject 
will be approached for participation in the study by her physician, who will discuss the 

 
Version: 19.120 
Date: 11/13/28/2013 
      
34 
protocol along with the risks and potential benefits of participating in it. A clear statement will 
be made concerning the voluntary nature of her participation and that her decision will have 
no effect on her remaining care.  The research nurse will follow with a detailed review of the 
informed consent document. The research subject will be encouraged to have family or 
friends participate in any or all of the process.  The research subject will be given time to ask 
questions, will be questioned to be certain she understands the information, and if she 
agrees to proceed, will sign consent. In general, registration and prestudy work will begin 
the next business day, allowing additional time for the research subject to reflect and 
request additional questions or withdraw. The consent process will be documented in the 
medical record. A copy of the informed consent document will be given to the research 
subject, and additional copies will be sent to the medical records department for distribution 
to the research pharmacy. The original informed consent will be filed with the subject file in 
CCTO. The consent process will occur in a private exam room or in the private office of the 
research nurse. There will be no additional recruitment materials. The principles of informed 
consent are described by Federal Regulatory Guidelines (21CFR50) and the Office for 
Human Research Protections: Protection of Human Subjects (Code of Federal Regulations 
45CFR46).  These principles must be followed to comply with FDA regulations for the 
conduct and monitoring of clinical investigations.  
 
b. 
Institutional Review: This study will be approved by the UAMS IRB as defined by Federal 
Regulatory Guidelines 21CFR56 and the Office for Human Research Protections: Protection 
of Human Subjects 45CFR46. This study will also undergo scientific review by the Cancer 
Institute’s Protocol Review and Monitoring Committee (PRMC). Approval by both the IRB 
and PRMC is required before the clinical trial can be activated.  An active IND is also 
required before the clinical trial can commence. 
 
  Approval will also be obtained by the U.S. Army Medical Research and Materiel       
 Command, Human Subjects Research Review Board (HSRRB) prior to implementation. A 
 copy of all continuing review and final reports will be submitted to the HSRRB.  
 
c. 
Investigational Agent Accountability: For each investigational drug, drug disposition 
(drug receipt, dispensing, transfer or return) will be maintained on the UAMS Investigational 
Agent Accountability Record. Drug supplies will be kept in a secure, limited access storage 
area under the recommended storage conditions in the research pharmacy in the Winthrop 
P. Rockefeller Cancer Institute under the direction of the research pharmacist.  During the 
course of the study, the following information will be noted on the Investigational Agent 
Accountability Record; the study number, the research subject’s initials, the research 
subject’s assigned number, the dose of drug, the date(s) and quantity of drug dispensed to 
the subject, the balance forward, the lot number and the recorder’s initials. These 
Investigational Agent Accountability Records will be readily available for inspection and are 
open to FDA inspection at any time. 
 
 
 
 
16. Adverse Events 
 
a. 
Adverse Event (AE): Any unfavorable and unintended sign, symptom or disease temporally 
associated with the use of a medical treatment or procedure regardless of whether it is 
considered related to the medical treatment or procedure.  Each AE is a unique 

 
Version: 19.120 
Date: 11/13/28/2013 
      
35 
representation of a specific event used for medical documentation and scientific analysis.  
[ICH E6 1.2] 
 
b. 
Serious Adverse Event (SAE): Any adverse drug experience occurring at any dose that 
results in any of the following outcomes: death, a life-threatening adverse drug experience, 
inpatient hospitalization, prolongation of existing hospitalization, a persistent or significant 
disability/incapacity or a congenital anomaly/birth defect.  Important medical events that may 
not result in death, be life threatening or require hospitalization may be considered a serious 
adverse drug experience when, based upon medical judgment, they may jeopardize the 
subject and may require medical or surgical intervention to prevent one of the outcomes 
listed in this definition.  FDA requires IND sponsors to report qualified AEs and SAEs 
through the expedited reporting system.  [21CFR312.32 (a)   ICH E6 1.50 Partially IRB 
Handbook policy 10.3] 
 
Any instance of a Guillain-Barre Syndrome event will require expedited reporting to the FDA, 
IRB, DoD and IND Sponsor. 
 
Any adverse experience that meets reporting guidelines for SAEs must be reported to the 
CRA for the study in the CCTO within 24 hours of knowledge of the event.  The CRA will 
follow AE reporting plans per institutional policies and applicable regulations. All qualified 
AEs will be reported to the Investigators, U. S. Army Medical Research and Material 
Command Human Subjects Research Review Board (HSRRB), FDA, IRB, Medical Monitor, 
and the UAMS Research Support Center according to this plan.  
 
All AEs and SAEs must be recorded in the appropriate section of the CRF.  The report 
should include, whenever possible, the investigator’s written medical judgment as to the 
relationship of the AE/SAE to study medications(s) (i.e., “probable”, “possible” or 
“unrelated”).  
 
Unanticipated problems involving risk to subjects or others, serious adverse events related 
to participation in the study and all subject deaths should be promptly reported by phone 
(301-619-2165), by email (hsrrb@det.amedd.army.mil), or by facsimile (301-619-7803) to 
the U.S. Army Medical Research and Materiel Command, Human Subjects Research 
Review Board (HSRRB).  A complete written report should follow the initial notification.  In 
addition to the methods above, the complete report can be sent to the U.S. Army Medical 
Research and Materiel Command, ATTN: MCMR-ZB-QH, 504 Scott Street, Fort Detrick, 
Maryland 21702-5012. 
 
17. Monitoring 
 
a. 
Medical Monitor: The Medical Monitor, Principal Investigator and study staff will meet to 
review safety data after each cohort has been enrolled and prior to enrollment of the next 
cohort.  Any death will trigger an additional urgent meeting.  
 
b. 
Data Monitor: UAMS is the IND Sponsor. One (or more) Data Monitor(s) will be appointed 
by the monitoring division of the UAMS Research Support Center (RSC) to assure that the 
rights and well-being of human subjects are protected, that the data are accurate, complete 
and verifiable from source documents and that the trial is conducted in compliance with 

 
Version: 19.120 
Date: 11/13/28/2013 
      
36 
currently approved protocol/amendments, with GCP, and with the applicable regulatory 
requirements set forth in 21 CFR 312. 
 
The Data Monitor(s) will be familiar with the investigational products, the protocol, the 
informed consent form, any other information provided to the subjects, SOPs, GCP and 
applicable regulatory requirements. 
 
Data Monitor(s) will have access to research subjects’ medical records and other study-
related records. The investigator agrees to cooperate with the study coordinator and Medical 
Monitor to ensure that any problems detected in the course of these monitoring visits are 
resolved.  Personal contact between the Data Monitor, Medical Monitor, study staff and the 
investigator will be maintained throughout the clinical trial to assure that the investigator is 
fulfilling his/her obligations and that the facilities used in the clinical trial remain acceptable. 
 
1) Pre-investigation Site Visit: A pre-investigation site visit will be performed by the 
Data Monitor in order to inspect the facility where the study is going to be 
conducted, and to assure that the investigator and his/her staff understand the 
protocol and agree to comply with the current regulations for clinical trial conduct in 
human subjects (21 CFR 312, 21 CFR 50, 21 CFR 56, 21 CFR 11, 21 CFR 21). 
The Data Monitor will document the IRB approval and generate a special report that 
will allow subject enrollment on the trial to begin.  
 
2) Periodic Site Visits: The first visit of the Data Monitor will occur as soon as 
possible after the first research subject has enrolled in the study. Subsequent 
monitoring visits will take place approximately quarterly based on enrollment. The 
Data Monitor will review the CRFs, source data/documents and other trial-related 
records for accuracy, consistency and completeness. Enrollment of research 
subjects after meeting eligibility criteria and signing a consent form will be 
documented. Missing visits, withdrawals and subject recruitment rate will be 
monitored.  
 
3) Investigational Products: The Data Monitor will verify that the storage conditions 
are appropriate and that the investigational drug is being dispensed to eligible 
subjects according to the study protocol. The Data Monitor will verify that there are 
accurate records of the receipt, use and return of the investigational product. 
 
4) Monitoring Report: After each monitoring visit a separate monitoring report will be 
generated and submitted to the investigator, U. S. Army Medical Research and 
Material Command Human Subjects Research Review Board, research staff and 
the Medical Monitor. This report will include significant findings related to 
deficiencies and deviations from the protocol, SOPs, GCP and the applicable 
regulatory requirements and actions taken to prevent recurrence of the detected 
deviations. The report will make recommendations for actions to be taken to secure 
compliance. The study team, which includes the PI, subinvestigators, biostatistician, 
Medical Monitor and the study coordinator, will meet after each cohort completes 
the protocol to review AEs and review monitoring reports in order to make 
adjustments necessary to protect the research subjects and the integrity of the trial. 
 
5) Research Subject Safety and Stopping Rules: If 3 SAEs occur with attribution to 
the study drug, the trial will be suspended until further review is completed by the 

 
Version: 19.120 
Date: 11/13/28/2013 
      
37 
Medical Monitor, PI, sponsor and FDA. This will be accomplished by the study 
team, the PI, Co-PI, biostatistician, Medical Monitor and CRA, either at the regular 
meeting or a special meeting called by the Medical Monitor or the PI because of the 
SAEs.  The study will be stopped upon Death not related to cancer or two Grade 4 
Toxicities related to the study article. 
 
6) Audits: An audit by UAMS will be scheduled after the completion of the first cohort. 
The audit will follow UAMS’ standard auditing procedure. 

 
Version: 19.120 
Date: 11/13/28/2013 
      
38 
 
18. Bibliography 
 
1. 
Sabbatini, P. J., Kudryashov, V., Ragupathi, G., Danishefsky, S. J., Livingston, P. O., 
Bornmann, W., Spassova, M., Zatorski, A., Spriggs, D., Aghajanian, C., Soignet, S., Peyton, 
M., O'Flaherty, C., Curtin, J., and Lloyd, K. O. Immunization of ovarian cancer patients with 
a synthetic Lewis(y)-protein conjugate vaccine: a phase 1 trial. Int J Cancer, 87: 79-85., 
2000. 
2.  Takahashi, T., Johnson, T. D., Nishinaka, Y., Morton, D. L., and Irie, R. F. IgM anti-
ganglioside antibodies induced by melanoma cell vaccine correlate with survival of 
melanoma patients. J Invest Dermatol, 112: 205-209, 1999. 
3. 
Vollmers, H. P. and Brandlein, S. The "early birds": natural IgM antibodies and immune 
surveillance. Histol Histopathol, 20: 927-937., 2005. 
4. 
Hakomori, S. Carbohydrate-to-carbohydrate interaction, through glycosynapse, as a basis of 
cell recognition and membrane organization. Glycoconj J, 21: 125-137., 2004. 
5. 
Hakomori, S. Tumor-associated carbohydrate antigens defining tumor malignancy: basis for 
development of anti-cancer vaccines. Adv Exp Med Biol, 491: 369-402., 2001. 
6. 
Kyte, J. A., Kvalheim, G., Aamdal, S., Saeboe-Larssen, S., and Gaudernack, G. Preclinical 
full-scale evaluation of dendritic cells transfected with autologous tumor-mRNA for 
melanoma vaccination. Cancer Gene Ther, 12: 579-591., 2005. 
7. 
Ragupathi, G., Liu, N. X., Musselli, C., Powell, S., Lloyd, K., and Livingston, P. O. Antibodies 
against tumor cell glycolipids and proteins, but not mucins, mediate complement-dependent 
cytotoxicity. J Immunol, 174: 5706-5712., 2005. 
8. 
Zhang, H., Zhang, S., Cheung, N. K., Ragupathi, G., and Livingston, P. O. Antibodies 
against GD2 ganglioside can eradicate syngeneic cancer micrometastases. Cancer Res, 58: 
2844-2849., 1998. 
9. 
Monzavi-Karbassi, B., Artaud, C., Jousheghany, F., Hennings, L., Carcel-Trullols, J., Shaaf, 
S., Korourian, S., and Kieber-Emmons, T. Reduction of spontaneous metastases through 
induction of carbohydrate cross-reactive apoptotic antibodies. J Immunol, 174: 7057-7065., 
2005. 
10. McCool, T. L., Harding, C. V., Greenspan, N. S., and Schreiber, J. R. B- and T-cell immune 
responses to pneumococcal conjugate vaccines: divergence between carrier- and 
polysaccharide-specific immunogenicity. Infect Immun, 67: 4862-4869., 1999. 
11. Gilewski, T., Ragupathi, G., Bhuta, S., Williams, L. J., Musselli, C., Zhang, X. F., Bornmann, 
W. G., Spassova, M., Bencsath, K. P., Panageas, K. S., Chin, J., Hudis, C. A., Norton, L., 
Houghton, A. N., Livingston, P. O., and Danishefsky, S. J. Immunization of metastatic breast 
cancer patients with a fully synthetic globo H conjugate: a phase I trial. Proc Natl Acad Sci U 
S A, 98: 3270-3275., 2001. 
12. Guthmann, M. D., Bitton, R. J., Carnero, A. J., Gabri, M. R., Cinat, G., Koliren, L., Lewi, D., 
Fernandez, L. E., Alonso, D. F., Gomez, D. E., and Fainboim, L. Active specific 
immunotherapy of melanoma with a GM3 ganglioside-based vaccine: a report on safety and 
immunogenicity. J Immunother, 27: 442-451., 2004. 
13. Krug, L. M., Ragupathi, G., Hood, C., Kris, M. G., Miller, V. A., Allen, J. R., Keding, S. J., 
Danishefsky, S. J., Gomez, J., Tyson, L., Pizzo, B., Baez, V., and Livingston, P. O. 
Vaccination of patients with small-cell lung cancer with synthetic fucosyl GM-1 conjugated to 
keyhole limpet hemocyanin. Clin Cancer Res, 10: 6094-6100., 2004. 
14. Ragupathi, G., Livingston, P. O., Hood, C., Gathuru, J., Krown, S. E., Chapman, P. B., 
Wolchok, J. D., Williams, L. J., Oldfield, R. C., and Hwu, W. J. Consistent antibody response 

 
Version: 19.120 
Date: 11/13/28/2013 
      
39 
against ganglioside GD2 induced in patients with melanoma by a GD2 lactone-keyhole 
limpet hemocyanin conjugate vaccine plus immunological adjuvant QS-21. Clin Cancer Res, 
9: 5214-5220., 2003. 
15. Krug, L. M., Ragupathi, G., Ng, K. K., Hood, C., Jennings, H. J., Guo, Z., Kris, M. G., Miller, 
V., Pizzo, B., Tyson, L., Baez, V., and Livingston, P. O. Vaccination of small cell lung cancer 
patients with polysialic acid or N-propionylated polysialic acid conjugated to keyhole limpet 
hemocyanin. Clin Cancer Res, 10: 916-923., 2004. 
16. Slovin, S. F., Ragupathi, G., Fernandez, C., Jefferson, M. P., Diani, M., Wilton, A. S., 
Powell, S., Spassova, M., Reis, C., Clausen, H., Danishefsky, S., Livingston, P., and Scher, 
H. I. A bivalent conjugate vaccine in the treatment of biochemically relapsed prostate 
cancer: a study of glycosylated MUC-2-KLH and Globo H-KLH conjugate vaccines given 
with the new semi-synthetic saponin immunological adjuvant GPI-0100 OR QS-21. Vaccine, 
23: 3114-3122., 2005. 
17. Holmberg, L. A. and Sandmaier, B. M. Vaccination with Theratope (STn-KLH) as treatment 
for breast cancer. Expert Rev Vaccines, 3: 655-663., 2004. 
18. Chapman, P. B., Wu, D., Ragupathi, G., Lu, S., Williams, L., Hwu, W. J., Johnson, D., and 
Livingston, P. O. Sequential immunization of melanoma patients with GD3 ganglioside 
vaccine and anti-idiotypic monoclonal antibody that mimics GD3 ganglioside. Clin Cancer 
Res, 10: 4717-4723., 2004. 
19. Foon, K. A., Lutzky, J., Baral, R. N., Yannelli, J. R., Hutchins, L., Teitelbaum, A., Kashala, O. 
L., Das, R., Garrison, J., Reisfeld, R. A., and Bhattacharya-Chatterjee, M. Clinical and 
immune responses in advanced melanoma patients immunized with an anti-idiotype 
antibody mimicking disialoganglioside GD2. J Clin Oncol, 18: 376-384., 2000. 
20. Qiu, J., Luo, P., Wasmund, K., Steplewski, Z., and Kieber-Emmons, T. Towards the 
development of peptide mimotopes of carbohydrate antigens as cancer vaccines. 
Hybridoma, 18: 103-112., 1999. 
21. Bolesta, E., Kowalczyk, A., Wierzbicki, A., Rotkiewicz, P., Bambach, B., Tsao, C. Y., 
Horwacik, I., Kolinski, A., Rokita, H., Brecher, M., Wang, X., Ferrone, S., and Kozbor, D. 
DNA vaccine expressing the mimotope of GD2 ganglioside induces protective GD2 cross-
reactive antibody responses. Cancer Res, 65: 3410-3418., 2005. 
22. Forster-Waldl, E., Riemer, A. B., Dehof, A. K., Neumann, D., Bramswig, K., Boltz-Nitulescu, 
G., Pehamberger, H., Zielinski, C. C., Scheiner, O., Pollak, A., Lode, H., and Jensen-
Jarolim, E. Isolation and structural analysis of peptide mimotopes for the disialoganglioside 
GD2, a neuroblastoma tumor antigen. Mol Immunol, 42: 319-325., 2005. 
23. Willers, J., Lucchese, A., Kanduc, D., and Ferrone, S. Molecular mimicry of phage displayed 
peptides mimicking GD3 ganglioside. Peptides, 20: 1021-1026., 1999. 
24. O, I., Kieber-Emmons, T., Otvos, L., Jr., and Blaszczyk-Thurin, M. Peptides mimicking sialyl-
Lewis A isolated from a random peptide library and peptide array. Ann N Y Acad Sci, 886: 
276-279., 1999. 
25. Kieber-Emmons, T., Luo, P., Qiu, J., Chang, T. Y., O, I., Blaszczyk-Thurin, M., and 
Steplewski, Z. Vaccination with carbohydrate peptide mimotopes promotes anti-tumor 
responses. Nat Biotechnol, 17: 660-665., 1999. 
26. Monzavi-Karbassi, B., Luo, P., Jousheghany, F., Torres-Quinones, M., Cunto-Amesty, G., 
Artaud, C., and Kieber-Emmons, T. A mimic of tumor rejection antigen-associated 
carbohydrates mediates an antitumor cellular response. Cancer Res, 64: 2162-2166., 2004. 
27. Dabelsteen, E. Cell surface carbohydrates as prognostic markers in human carcinomas. J 
Pathol, 179: 358-369., 1996. 
28. Davidson, B., Gotlieb, W. H., Ben-Baruch, G., Kopolovic, J., Goldberg, I., Nesland, J. M., 
Berner, A., Bjamer, A., and Bryne, M. Expression of carbohydrate antigens in advanced-

 
Version: 19.120 
Date: 11/13/28/2013 
      
40 
stage ovarian carcinomas and their metastases-A clinicopathologic study. Gynecol Oncol, 
77: 35-43., 2000. 
29. Nakagoe, T., Fukushima, K., Tanaka, K., Sawai, T., Tsuji, T., Jibiki, M., Nanashima, A., 
Yamaguchi, H., Yasutake, T., Ayabe, H., and Arisawa, K. Evaluation of sialyl Lewis(a), sialyl 
Lewis(x), and sialyl Tn antigens expression levels as predictors of recurrence after curative 
surgery in node-negative colorectal cancer patients. J Exp Clin Cancer Res, 21: 107-113., 
2002. 
30. Ravn, V., Mandel, U., Svenstrup, B., and Dabelsteen, E. Expression of type-2 histo-blood 
group carbohydrate antigens (Le(x), Le(y), and H) in normal and malignant human 
endometrium. Virchows Arch, 424: 411-419., 1994. 
31. Zhang, S., Zhang, H. S., Reuter, V. E., Slovin, S. F., Scher, H. I., and Livingston, P. O. 
Expression of potential target antigens for immunotherapy on primary and metastatic 
prostate cancers. Clin Cancer Res, 4: 295-302., 1998. 
32. Steplewski, Z., Blaszczyk-Thurin, M., Lubeck, M., Loibner, H., Scholz, D., and Koprowski, H. 
Oligosaccharide Y specific monoclonal antibody and its isotype switch variants. Hybridoma, 
9: 201-210., 1990. 
33. Marquina G, Waki H, Fernandez LE, Kon K, Carr A, Valiente O, Perez R, Ando S 
Gangliosides expressed in human breast cancer. Cancer Res. 1996;56:5165-71 
34. Luo, P., Agadjanyan, M., Qiu, J., Westerink, M. A., Steplewski, Z., and Kieber-Emmons, T. 
Antigenic and immunological mimicry of peptide mimotopes of Lewis carbohydrate antigens. 
Mol Immunol, 35: 865-879., 1998. 
35. Mostert, B., Sleijfer, S., Foekens, J. A., and Gratama, J. W. 2009. Circulating tumor cells 
(CTCs): Detection methods and their clinical relevance in breast cancer. Cancer Treat Rev. 
36. Sleijfer, S., Gratama, J. W., Sieuwerts, A. M., Kraan, J., Martens, J. W., and Foekens, J. A. 
2007. Circulating tumour cell detection on its way to routine diagnostic implementation? Eur 
J Cancer 43:2645-50. 
37. Paterlini-Brechot, P. and Benali, N. L. 2007. Circulating tumor cells (CTC) detection: clinical 
impact and future directions. Cancer Lett 253:180-204. 
38. Tibbe, A. G., Miller, M. C., and Terstappen, L. W. 2007. Statistical considerations for 
enumeration of circulating tumor cells. Cytometry A 71:154-62. 
39. Cristofanilli, M., Budd, G. T., Ellis, M. J., Stopeck, A., Matera, J., Miller, M. C., Reuben, J. 
M., Doyle, G. V., Allard, W. J., Terstappen, L. W., and Hayes, D. F. 2004. Circulating tumor 
cells, disease progression, and survival in metastatic breast cancer. N Engl J Med 351:781-
91. 
40. He, W., Kularatne, S. A., Kalli, K. R., Prendergast, F. G., Amato, R. J., Klee, G. G., 
Hartmann, L. C., and Low, P. S. 2008. Quantitation of circulating tumor cells in blood 
samples from ovarian and prostate cancer patients using tumor-specific fluorescent ligands. 
Int J Cancer 123:1968-73. 
41. Biris, A. S., Galanzha, E. I., Li, Z., Mahmood, M., Xu, Y., and Zharov, V. P. 2009. In vivo 
Raman flow cytometry for real-time detection of carbon nanotube kinetics in lymph, blood, 
and tissues. J Biomed Opt 14:021006. 
42. Gilewski, T., Adluri, S., Ragupathi, G., Zhang, S., Yao, T. J., Panageas, K., Moynahan, M., 
Houghton, A., Norton, L., and Livingston, P. O. Vaccination of high-risk breast cancer 
patients with mucin-1 (MUC1) keyhole limpet hemocyanin conjugate plus QS-21. Clin 
Cancer Res, 6: 1693-1701., 2000. 
43. Storer, B. E. 1989. Design and analysis of phase I clinical trials. Biometrics 45:925-37. 
44. Müller V, Hayes DF, Pantel K. Breast Cancer Res. 2006;8(5):110)